WorldWideScience

Sample records for radioiodinated human recombinant

  1. Recombinant human thyrotropin-stimulated radioiodine therapy of nodular goiter allows major reduction of the radiation burden with retained efficacy

    DEFF Research Database (Denmark)

    Fast, Søren; Hegedüs, Laszlo; Grupe, Peter

    2010-01-01

    Context and Objective: Stimulation with recombinant human TSH (rhTSH) before radioiodine ((131)I) therapy augments goiter volume reduction (GVR). Observations indicate that rhTSH has a preconditioning effect beyond increasing thyroid (131)I uptake. We test the hypothesis that an equivalent GVR mi....... This approach is attractive in terms of minimizing posttherapeutic restrictions and in reducing the potential risk of radiation-induced malignancy....

  2. Comparison between thyroid hormone withdrawal and recombinant human TSH administration before radioiodine treatment for advanced thyroid cancer

    International Nuclear Information System (INIS)

    Coelho, Sabrina M.; Corbo, Rossana; Buescu, Alexandru; Carvalho, Denise P.; Vaisman, Mario

    2005-01-01

    Full text: Radioiodine treatment is traditionally performed after thyroid hormone withdrawal. However, induction of hypothyroidism is associated with physical and psychological symptoms and a possible induction of tumor growth. This is particularly harmful in patients with advanced thyroid cancer (ATC). The objective of this study was to compare the thyroxine withdrawal and the recombinant human TSH (rh TSH) administration in patients with non-radioiodine responsive ATC after retinoic acid (RA) therapy for induction of iodine uptake. Patients were treated with isotretinoin (1.0 to 1.5 mg/kg/d) for 5 weeks, then, thyroxine (LT 4 ) was discontinued 4 weeks before therapeutic dose (150 mCi). Based on the presence of a satisfactory response to RA (increased iodine uptake, reduction of serum thyroglobulin and tumor regression), another cycle of RA was offered, then rh TSH was used (0.9 mg in two consecutive days). A total of 8 patients (1 follicular, 1 poorly differentiated and 6 papillary carcinomas) were treated. In a patient with pituitary adenoma the endogenous TSH did not rise after T 4 withdrawal, and rh TSH was administered before radioiodine therapy. Although an increase in iodine uptake was observed after RA therapy in the patient with poorly differentiated cancer, the tumor continued to progress and patient died of respiratory insufficiency. Four out of 7 patients had at least a partial response and were selected for re-treatment. Post-therapeutic whole body scan was similar using both protocols, but patients had fewer side effects with rh TSH. One patient who had no compressive symptoms during LT 4 withdrawal did present dysphagia and dysphonia secondary to tumor swelling, 6 hours after the last rh TSH injection. Glucocorticoid was administered and symptoms were reversed after 10 days. Conclusion: Radioiodine uptake using rh TSH was comparable to T 4 withdrawal and is particularly useful when endogenous TSH cannot rise. However, the possibility of compressive

  3. Usefulness of recombinant human TSH aided radioiodine doses administered in patients with differentiated thyroid carcinoma

    International Nuclear Information System (INIS)

    Pitoia, Fabian; El Tamer, Elias; Schere, Daniel B.; Passerieu, Mariano; Bruno, Oscar D.; Niepominiszcze, Hugo

    2006-01-01

    The published studies confirming the safety and efficacy of rh TSH for diagnostic purposes have led to an increased interest in its use for preparation for radioiodine (RI) dose administration in patients with recurrent or persistent differentiated thyroid carcinoma (DTC). In order to establish the efficacy of RI therapy after rh TSH, we have reviewed 39 rh TSH-aided radioiodine treatments in a series of 28 DTC patients. Patients were divided into two groups: GI (n=17), with previous thyroid bed uptake and undetectable thyroglobulin (Tg) levels under levothyroxine treatment and GII (n=11), with proven metastatic local or distant disease. Median follow-up after the first rh TSH-aided radioiodine treatment was 32 ± 13 months (range 8 to 54 months). Sixteen patients (94%) in GI were rendered disease free and one patient was shown to have persistent disease. In GII, the post therapy whole body scan showed pathological uptakes in all cases: in four patients in lungs, in four in mediastinum and in three in lateral neck. In two patients with mediastinum uptake, Tg levels were undetectable after rh TSH. In the follow-up, two patients with lateral neck uptake were rendered disease free, four patients died (three due to thyroid cancer) and five out of the remaining patients have persistent disease. In conclusion, rh TSH aided therapy was helpful to eliminate normal thyroid bed remnants in 16/17 (94%) patients (GI). rh TSH stimulated Tg was undetectable in two patients with mediastinal metastasis. We believe that rh TSH is a good alternative to levothyroxine withdrawal for the treatment of DTC with radioactive iodine, increasing the quality of life in these patients. Caution should be recommended in the follow-up of unselected DTC patients only with stimulated Tg levels. (author) [es

  4. Radioiodine plus recombinant human thyrotropin do not cause acute airway compression and are effective in reducing multinodular goiter

    Energy Technology Data Exchange (ETDEWEB)

    Albino, C.C., E-mail: ccalbino@uol.com.b [Instituto de Diabetes e Endocrinologia de Maringa, PR (Brazil); Graf, H.; Paz-Filho, G. [Universidade Federal do Parana (UFPR), Curitiba, PR (Brazil). Hospital das Clinicas. Servico de Endocrinologia e Metabologia; Diehl, L.A. [Universidade Estadual de Londrina (UEL), PR (Brazil); Olandoski, M.; Sabbag, A. [Pontificia Univ. Catolica do Parana (PUCPR), Curitiba, PR (Brazil). Nucleo de Bioestatistica; Buchpiguel, C. [Universidade de Sao Paulo (USP), SP (Brazil). Dept. de Radiologia

    2006-03-15

    Recombinant human thyrotropin (rhTSH) reduces the activity of radioiodine required to treat multinodular goiter (MNG), but acute airway compression can be a life-threatening complication. In this prospective, randomized, double-blind, placebo-controlled study, we assessed the efficacy and safety (including airway compression) of different doses of rhTSH associated with a fixed activity of {sup 131}I for treating MNG. Euthyroid patients with MNG (69.3 +- 62.0 mL, 20 females, 2 males, 64 +- 7 years) received 0.1 mg (group I, N = 8) or 0.01 mg (group II, N = 6) rhTSH or placebo (group III, N = 8), 24 h before 1.11 GBq {sup 131}I. Radioactive iodine uptake was determined at baseline and 24 h after rhTSH and thyroid volume (TV, baseline and 6 and 12 months after treatment) and tracheal cross-sectional area (TCA, baseline and 2, 7, 180, and 360 days after rhTSH) were determined by magnetic resonance; antithyroid antibodies and thyroid hormones were determined at frequent intervals. After 6 months, TV decreased significantly in groups I (28.5 +- 17.6%) and II (21.6 +- 17.8%), but not in group III (2.7 +- 15.3%). After 12 months, TV decreased significantly in groups I (36.7 +- 18.1%) and II (37.4 +- 27.1%), but not in group III (19.0 +- 24.3%). No significant changes in TCA were observed. T3 and free T4 increased transiently during the first month. After 12 months, 7 patients were hypothyroid (N 3 in group I and N = 2 in groups II and III). rhTSH plus a 1.11-GBq fixed {sup 131}I activity did not cause acute or chronic changes in TCA. After 6 and 12 months, TV reduction was more pronounced among patients treated with rhTSH plus {sup 131}I (author)

  5. Radioiodine plus recombinant human thyrotropin do not cause acute airway compression and are effective in reducing multinodular goiter

    International Nuclear Information System (INIS)

    Albino, C.C.; Graf, H.; Paz-Filho, G.; Olandoski, M.; Sabbag, A.; Buchpiguel, C.

    2006-01-01

    Recombinant human thyrotropin (rhTSH) reduces the activity of radioiodine required to treat multinodular goiter (MNG), but acute airway compression can be a life-threatening complication. In this prospective, randomized, double-blind, placebo-controlled study, we assessed the efficacy and safety (including airway compression) of different doses of rhTSH associated with a fixed activity of 131 I for treating MNG. Euthyroid patients with MNG (69.3 ± 62.0 mL, 20 females, 2 males, 64 ± 7 years) received 0.1 mg (group I, N = 8) or 0.01 mg (group II, N = 6) rhTSH or placebo (group III, N = 8), 24 h before 1.11 GBq 131 I. Radioactive iodine uptake was determined at baseline and 24 h after rhTSH and thyroid volume (TV, baseline and 6 and 12 months after treatment) and tracheal cross-sectional area (TCA, baseline and 2, 7, 180, and 360 days after rhTSH) were determined by magnetic resonance; antithyroid antibodies and thyroid hormones were determined at frequent intervals. After 6 months, TV decreased significantly in groups I (28.5 ± 17.6%) and II (21.6 ± 17.8%), but not in group III (2.7 ± 15.3%). After 12 months, TV decreased significantly in groups I (36.7 ± 18.1%) and II (37.4 ± 27.1%), but not in group III (19.0 ± 24.3%). No significant changes in TCA were observed. T3 and free T4 increased transiently during the first month. After 12 months, 7 patients were hypothyroid (N 3 in group I and N = 2 in groups II and III). rhTSH plus a 1.11-GBq fixed 131 I activity did not cause acute or chronic changes in TCA. After 6 and 12 months, TV reduction was more pronounced among patients treated with rhTSH plus 131 I (author)

  6. Cost-effectiveness of using recombinant human thyroid-stimulating hormone before radioiodine ablation for thyroid cancer treatment in Spanish hospitals.

    Science.gov (United States)

    Vallejo, J A; Muros, M A

    In thyroid cancer treatment, the thyroid-stimulating hormone (TSH) must be elevated before radioiodine ablation, either by exogenous (with recombinant human thyrotropin [rhTSH]) or endogenous stimulation by thyroid hormone withdrawal (THW). The use of rhTSH avoids hypothyroidism and favours the subsequent elimination of radioiodine, but involves the cost of the product. For this reason, a cost-effectiveness analysis was performed, taking into account all costs involved and the benefits associated with the use of this therapy. Using a Markov modelling with two analysis arms (rhTSH and THW), stratified into high (100mCi/3700 MBq) and low (30mCi/1110 MBq) radioiodine doses, and using 17 weekly cycles, the incremental cost per quality-adjusted life-year (QALY) related to the use of rhTSH was determined. The clinical inputs included in the model were based on published studies and in a treatment survey conducted in Spain. Radioablation preparation with rhTSH is superior to THW, showing additional benefits (0.048 AVAC), as well as cost savings (-€614.16), with an incremental cost-effectiveness rate (ICER) of -€12,795/QALY. The univariate and multivariate sensitivity analyses showed the result to be robust. The use of rhTSH previous to radioablation in Spain has cost savings, as well as a series of health benefits for the patient, making it highly cost-effective. Copyright © 2017 Elsevier España, S.L.U. y SEMNIM. All rights reserved.

  7. Characterization of the binding of radioiodinated hybrid recombinant IFN-alpha A/D to murine and human lymphoid cell lines

    International Nuclear Information System (INIS)

    Faltynek, C.R.; Princler, G.L.; Schwabe, M.; Shata, M.T.; Lewis, G.K.; Kamin-Lewis, R.M.

    1990-01-01

    The hybrid recombinant human interferon (IFN) rIFN-alpha A/D was radioiodinated. Specific binding of [125I]rIFN-alpha A/D was observed with both human and murine cell lines. The binding of [125I]rIFN-alpha A/D to human Daudi cells had similar characteristics to the previously described binding of [125I]rIFN-alpha A or -alpha 2. The following lines of evidence demonstrated that [125I]rIFN-alpha A/D bound with high affinity to the same receptor on murine cells as murine IFN-alpha and -beta: (i) the binding of [125I]rIFN-alpha A/D to murine LBRM cells was inhibited to a similar extent by natural murine IFN-alpha, natural murine IFN-beta, and rIFN-A/D; (ii) the Kd (approximately 2 X 10(-10) M) obtained from both competition experiments and saturation binding experiments with [125I]rIFN-alpha A/D was comparable to the previously reported Kd for the binding of natural murine IFN-alpha and -beta to other murine cell lines; (iii) the size of the cross-linked [125I]rIFN-alpha A/D receptor complex formed on murine LBRM cells was similar to the previously reported cross-linked complex formed after binding radioiodinated natural murine IFN-beta to other murine cell lines. Due to the current lack of readily available recombinant murine IFN-alpha or -beta for radiolabeling and the previously demonstrated biological activity of rIFN-alpha A/D on murine cells, [125I]rIFN-alpha A/D should prove to be a useful reagent for further studies of murine IFN receptors

  8. Quality of life changes and clinical outcomes in thyroid cancer patients undergoing radioiodine remnant ablation (RRA) with recombinant human TSH (rhTSH): a randomized controlled study.

    Science.gov (United States)

    Taïeb, D; Sebag, F; Cherenko, M; Baumstarck-Barrau, K; Fortanier, C; Farman-Ara, B; De Micco, C; Vaillant, J; Thomas, S; Conte-Devolx, B; Loundou, A; Auquier, P; Henry, J F; Mundler, O

    2009-07-01

    Recombinant human TSH (rhTSH) has become the modality of choice for radioiodine remnant ablation (RRA) in low-risk thyroid cancer patients. The aims of the present prospective randomized study were to evaluate the impact of TSH stimulation procedure (hypothyroidism vs. rhTSH) on quality of life (QoL) of thyroid cancer patients undergoing RRA and to evaluate efficacy of both procedures. L-T4 was initiated in both groups after thyroidectomy. After randomization, L-T4 was discontinued in hypothyroid (hypo) group and continued in rhTSH group. A measure of 3.7 GBq of radioiodine was given to both groups. The functional assessment of chronic illness therapy-fatigue (FACIT-F) was administered from the early postoperative period to 9 months. Socio-demographic parameters, anxiety and depression scales were also evaluated (CES-D, BDI and Spielberger state-trait questionnaires). At 9 months, patients underwent an rhTSH stimulation test, diagnostic (131)I whole body scan (dxWBS) and neck ultrasonography. A total of 74 patients were enrolled for the study. There was a significant decrease in QoL from baseline (t0) to t1 (RRA period) in the hypothyroid group with significant differences in FACIT-F TOI (P hypothyrodism. However, there is a wide heterogeneity in the clinical impact of hypothyroidism.

  9. Time to reconsider nonsurgical therapy of benign nontoxic multinodular goitre. Focus on recombinant human TSH (rhTSH) augmented radioiodine therapy

    DEFF Research Database (Denmark)

    Fast, Søren; Nielsen, Viveque; Bonnema, Steen

    2009-01-01

    alternatives are needed. Until recently, levothyroxine therapy was the preferred non-surgical alternative, but due to low efficacy and potential side-effects, it is not recommended for routine use in recent international guidelines. Conventional radioiodine (131I)-therapy has been used for two decades......, which makes 131I-therapy less feasible. Another challenge is the negative correlation between initial goitre size and goitre volume reduction (GVR). With its ability to more than double the thyroid 131I-uptake, recombinant human TSH (rhTSH) increases the absorbed radiation dose and thus enhances the GVR...... efficacy. Thus, although in its infancy, and still experimental, rhTSH-augmented 131I-therapy may profoundly alter the nonsurgical treatment of benign non-toxic MNG....

  10. Use of recombinant human thyrotropin (rh TSH) as a method of preparation for radioiodine therapy in thyroid disorders; Utilisation de la thyreostimuline humaine recombinante dans la preparation au traitement par iode-131 des pathologies thyroidiennes

    Energy Technology Data Exchange (ETDEWEB)

    Taieb, D.; Guillet, B.A.; Tessonnier, L.; Mundler, O. [Centre Hospitalo-Universitaire de la Timone, Service Central de Biophysique et de Medecine Nucleaire, 13 - Marseille (France)

    2008-02-15

    The introduction of recombinant human TSH (rh TSH) as a method of preparation for radioiodine therapy of follicular-derived thyroid tumors (benign and malignant) is a significant medical advance. Rh TSH has been approved for use in remnants ablation after total thyroidectomy for carcinoma. There are other potential uses for rh TSH that have not yet been licensed. The use of rh TSH allows to reduce administrated doses in goiters through an increase of iodine uptake and a more homogeneous distribution of radioiodine in the gland. Rh TSH also improves thyroid cancer patients quality of life by avoiding hypothyroidism. (authors)

  11. Recombinant human TSH-aided radioiodine treatment of advanced differentiated thyroid carcinoma: a single-centre study of 54 patients

    International Nuclear Information System (INIS)

    Jarzab, Barbara; Handkiewicz-Junak, Daria; Roskosz, Jozef; Puch, Zbigniew; Wygoda, Zbigniew; Kukulska, Aleksandra; Jurecka-Lubieniecka, Beata; Hasse-Lazar, Kornelia; Turska, Maria; Zajusz, Aleksander

    2003-01-01

    In 54 consecutive patients who had retained bulky metastatic and/or locoregional lesions of DTC despite the exhaustion of other therapeutic options, we gave one to four courses of two consecutive daily intramuscular injections of rhTSH, 0.9 mg, followed by a therapeutic activity of 131 I per os on day 3. Fifty patients had received prior radioiodine treatment aided by l-thyroxine (T 4 ) withdrawal. We included in the study 23 patients who had received a trial of isotretinoin therapy for re-differentiation of confirmed de-differentiated metastases. In a blinded, within-patient comparison of post-therapy whole-body scans after the first rhTSH-aided and latest withdrawal-aided treatments in patients with functional metastases at baseline, 18 of 27 (67%) scan pairs were concordant, four (15%) were discordant in favour of the rhTSH-aided scan and five (19%) were discordant in favour of the withdrawal-aided scan. In total, 37 (74%) of 50 paired scans were concordant, eight (16%) favoured rhTSH and five (10%) favoured withdrawal. All differences appeared to be attributable to clinical causes, not to any difference between endogenous and exogenous TSH stimulation. Reflecting the biochemical activity of rhTSH and the release of thyroglobulin (Tg) due to tumour destruction, median serum Tg concentration rose approximately fourfold between baseline and day 6 of the rhTSH-aided treatment course. rhTSH was well tolerated, with mostly minor, transient toxicity, except for neck oedema in three patients with neck infiltrates and pathological spine fracture in one patient with a large vertebral metastasis. At 6 months, complete response occurred in one (2%), partial response in 12 (26%) and disease stabilisation in 19 (40%) of 47 evaluable patients. The rate of complete + partial response was 41% and that of disease stabilisation, 30%, in the 27 evaluable patients with functional metastases at baseline; the corresponding rates were 10% and 55% in the 20 evaluable patients with non

  12. Effect of 30 mCi radioiodine on multinodular goiter previously treated with recombinant human thyroid-stimulating hormone

    Energy Technology Data Exchange (ETDEWEB)

    Paz-Filho, G.J.; Mesa-Junior, C.O.; Boguszewski, C.L.; Carvalho, G.A.; Graf, H. [Universidade Federal do Parana (UFPR), Curitiba, PR (Brazil). Hospital de Clinicas. Servico de Endocrinologia e Metabologia; Olandoski, M. [Pontificia Univ. Catolica do Parana, Curitiba, PR (Brazil). Nucleo de Bioestatistica; Woellner, L.C. [Centro de Medicina Nuclear, Curitiba, PR (Brazil); Goedert, C.A. [Centro de Tomografia Computadorizada, Curitiba, PR (Brazil)

    2007-12-15

    Recombinant human thyroid-stimulating hormone (rhTSH) enhances {sup 131}I uptake, permitting a decrease in radiation for the treatment of multinodular goiter (MNG). Our objective was to evaluate the safety and efficacy of a single 0.1-mg dose of rhTSH, followed by 30 mCi {sup 131}I, in patients with MNG. Seventeen patients (15 females, 59.0 {+-} 13.1 years), who had never been submitted to {sup 131}I therapy, received a single 0.1-mg injection of rhTSH followed by 30 mCi {sup 131}I on the next day. Mean basal thyroid volume measured by computed tomography was 106.1 {+-} 64.4 mL. {sup 131}I 24-h uptake, TSH, free-T4, T3, thyroglobulin, anti-thyroid antibodies, and thyroid volume were evaluated at regular intervals of 12 months. Mean {sup 131}I 24-h uptake increased from 18.1 {+-} 9.7 to 49.6 {+-} 13.4% (P < 0.001), a median 2.6-fold increase (1.2 to 9.2). Peak hormonal levels were 10.86 {+-} 5.44 mU/L for TSH (a median 15.5-fold increase), 1.80 {+-} 0.48 ng/dL for free-T4, 204.61 {+-} 58.37 ng/dL for T3, and a median of 557.0 ng/mL for thyroglobulin. The adverse effects observed were hyperthyroidism (17.6%), painful thyroiditis (29.4%) and hypothyroidism (52.9%). Thyroid volume was reduced by 34.3 {+-} 14.3% after 6 months (P < 0.001) and by 46.0 {+-} 14.6% after 1 year (P < 0.001). Treatment of MNG with a single 0.1-mg dose of rhTSH, followed by a fixed amount of radioactivity of {sup 131}I, leads to an efficacious decrease in thyroid volume for the majority of the patients, with a moderate incidence of non-serious and readily treatable adverse effects. (author)

  13. Tumour dosimetry and response in patients with metastatic differentiated thyroid cancer using recombinant human thyrotropin before radioiodine therapy

    Energy Technology Data Exchange (ETDEWEB)

    Keizer, Bart de; Hoekstra, Anne; Rijk, Peter P. van; Klerk, John M.H. de [Department of Nuclear Medicine, Room E02.222, University Medical Center Utrecht, Heidelberglaan 100, 3584 CX, Utrecht (Netherlands); Brans, Boudewijn; Dierckx, Rudi A. [Department of Nuclear Medicine, Ghent University Hospital, Ghent (Belgium); Zelissen, Pierre M.J.; Koppeschaar, Hans P.F.; Lips, Cees J.M. [Department of Endocrinology, University Medical Center Utrecht (Netherlands)

    2003-03-01

    The development of recombinant human thyrotropin (rhTSH) has given clinicians new options for diagnostic follow-up and treatment of patients with differentiated thyroid cancer (DTC). This paper evaluates the tumour dosimetry and response following -iodine-131 treatment of metastatic thyroid cancer patients after rhTSH stimulation instead of classical hormone withdrawal-induced hypothyroidism. Nineteen consecutive {sup 131}I treatments in 16 patients were performed after rhTSH stimulation. All patients had undergone a near-total thyroidectomy followed by an ablative dosage of {sup 131}I. They all suffered from metastatic or recurrent disease showing tumoral {sup 131}I uptake on previous post-treatment scintigraphy. Dosimetric calculations were performed using {sup 131}I tumour uptake measurements from post-treatment {sup 131}I scintigrams and tumour volume estimations from radiological images. Response was assessed by comparing pre-treatment serum thyroglobulin (Tg) level with the Tg level 3 months post treatment. In 18 out of 19 treatments, uptake of {sup 131}I in metastatic or recurrent lesions was seen. The median tumour radiation dose was 26.3 Gy (range 1.3-368 Gy), and the median effective half-life was 2.7 days (range 0.5-6.5 days). Eleven of 19 treatments (10/16 patients) were evaluable for response after 3 months. {sup 131}I therapy with rhTSH resulted in a biochemical partial response in 3/11 or 27% of treatments (two patients), biochemical stable disease in 2/11 or 18% of treatments and biochemical progressive disease in 6/11 or 55% of treatments. Our study showed that although tumour doses in DTC patients treated with {sup 131}I after rhTSH were highly variable, 45% of treatments led to disease stabilisation or partial remission when using rhTSH in conjunction with {sup 131}I therapy, without serious side-effects and with minimal impact on quality of life. RhTSH is therefore adequately satisfactory as an adjuvant tool in therapeutic settings and is

  14. Effect of 30 mCi radioiodine on multinodular goiter previously treated with recombinant human thyroid-stimulating hormone

    Directory of Open Access Journals (Sweden)

    G.J. Paz-Filho

    2007-12-01

    Full Text Available Recombinant human thyroid-stimulating hormone (rhTSH enhances 131I uptake, permitting a decrease in radiation for the treatment of multinodular goiter (MNG. Our objective was to evaluate the safety and efficacy of a single 0.1-mg dose of rhTSH, followed by 30 mCi 131I, in patients with MNG. Seventeen patients (15 females, 59.0 ± 13.1 years, who had never been submitted to 131I therapy, received a single 0.1-mg injection of rhTSH followed by 30 mCi 131I on the next day. Mean basal thyroid volume measured by computed tomography was 106.1 ± 64.4 mL. 131I 24-h uptake, TSH, free-T4, T3, thyroglobulin, anti-thyroid antibodies, and thyroid volume were evaluated at regular intervals of 12 months. Mean 131I 24-h uptake increased from 18.1 ± 9.7 to 49.6 ± 13.4% (P < 0.001, a median 2.6-fold increase (1.2 to 9.2. Peak hormonal levels were 10.86 ± 5.44 mU/L for TSH (a median 15.5-fold increase, 1.80 ± 0.48 ng/dL for free-T4, 204.61 ± 58.37 ng/dL for T3, and a median of 557.0 ng/mL for thyroglobulin. The adverse effects observed were hyperthyroidism (17.6%, painful thyroiditis (29.4% and hypothyroidism (52.9%. Thyroid volume was reduced by 34.3 ± 14.3% after 6 months (P < 0.001 and by 46.0 ± 14.6% after 1 year (P < 0.001. Treatment of MNG with a single 0.1-mg dose of rhTSH, followed by a fixed amount of radioactivity of 131I, leads to an efficacious decrease in thyroid volume for the majority of the patients, with a moderate incidence of non-serious and readily treatable adverse effects.

  15. Radioiodine Thyroid Remnant Ablation after Recombinant Human Thyrotropin or Thyroid Hormone Withdrawal in Patients with High-Risk Differentiated Thyroid Cancer

    Directory of Open Access Journals (Sweden)

    Fabián Pitoia

    2012-01-01

    Full Text Available To supplement limited relevant literature, we retrospectively compared ablation and disease outcomes in high-risk differentiated thyroid carcinoma (DTC patients undergoing radioiodine thyroid remnant ablation aided by recombinant human thyrotropin (rhTSH versus thyroid hormone withdrawal/withholding (THW. Our cohort was 45 consecutive antithyroglobulin antibody- (TgAb- negative, T3-T4/N0-N1-Nx/M0 adults ablated with high activities at three referral centers. Ablation success comprised negative (<1 μg/L stimulated serum thyroglobulin (Tg and TgAb, with absent or <0.1% scintigraphic thyroid bed uptake. “No evidence of disease” (NED comprised negative unstimulated/stimulated Tg and no suspicious neck ultrasonography or pathological imaging or biopsy. “Persistent disease” was failure to achieve NED, “recurrence,” loss of NED status. rhTSH patients (n=18 were oftener ≥45 years old and higher stage (P=0.01, but otherwise not different than THW patients (n=27 at baseline. rhTSH patients were significantly oftener successfully ablated compared to THW patients (83% versus 67%, P<0.02. After respective 3.3 yr and 4.5 yr mean follow-ups (P=0.02, NED was achieved oftener (72% versus 59% and persistent disease was less frequent in rhTSH patients (22% versus 33% (both comparisons P=0.03. rhTSH stimulation is associated with at least as good outcomes as is THW in ablation of high-risk DTC patients.

  16. Radioiodine thyroid remnant ablation after recombinant human thyrotropin or thyroid hormone withdrawal in patients with high-risk differentiated thyroid cancer.

    Science.gov (United States)

    Pitoia, Fabián; Marlowe, Robert J; Abelleira, Erika; Faure, Eduardo N; Bueno, Fernanda; Schwarzstein, Diego; Lutfi, Rubén Julio; Niepomniszcze, Hugo

    2012-01-01

    To supplement limited relevant literature, we retrospectively compared ablation and disease outcomes in high-risk differentiated thyroid carcinoma (DTC) patients undergoing radioiodine thyroid remnant ablation aided by recombinant human thyrotropin (rhTSH) versus thyroid hormone withdrawal/withholding (THW). Our cohort was 45 consecutive antithyroglobulin antibody- (TgAb-) negative, T3-T4/N0-N1-Nx/M0 adults ablated with high activities at three referral centers. Ablation success comprised negative (<1 μg/L) stimulated serum thyroglobulin (Tg) and TgAb, with absent or <0.1% scintigraphic thyroid bed uptake. "No evidence of disease" (NED) comprised negative unstimulated/stimulated Tg and no suspicious neck ultrasonography or pathological imaging or biopsy. "Persistent disease" was failure to achieve NED, "recurrence," loss of NED status. rhTSH patients (n = 18) were oftener ≥45 years old and higher stage (P = 0.01), but otherwise not different than THW patients (n = 27) at baseline. rhTSH patients were significantly oftener successfully ablated compared to THW patients (83% versus 67%, P < 0.02). After respective 3.3 yr and 4.5 yr mean follow-ups (P = 0.02), NED was achieved oftener (72% versus 59%) and persistent disease was less frequent in rhTSH patients (22% versus 33%) (both comparisons P = 0.03). rhTSH stimulation is associated with at least as good outcomes as is THW in ablation of high-risk DTC patients.

  17. Use of recombinant, human TSH radioiodine therapy in patients with differentiated thyroid carcinoma; Radioiodtherapie des differenzierten Schilddruesenkarzinoms nach Vorbehandlung mit rekombinantem, humanem TSH

    Energy Technology Data Exchange (ETDEWEB)

    Luster, M. [Wuerzburg Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin

    2001-12-01

    We describe the use of recombinant human TSH (rhTSH) in conjunction with ablative radioiodine therapy (RIT) in 11 patients (16 total treatments) with advanced and/or recurrent DTC (5 papillary, 6 follicular) for whom withdrawal of thyroid hormone suppressive therapy (THST) to increase serum TSH was not an option. Indications for rhTSH use in these patients included inability to tolerate withdrawal of thyroid hormones due to poor physical condition or inability to achieve sufficient serum TSH levels after THST withdrawal. All patients had undergone thyroidectomy and most (9/11) had received prior radioablative therapy after THST withdrawal. In 7 cases (5 patients), post-therapy Tg levels assessed at a mean of 4.3 months (range 2-10 months) after I-131 therapy were decreased by at least 30% compared to pre-therapy levels. In an additional 3 patients, whole body scans performed at follow-up indicated decreased or stabilized tumor burden compared to pre-therapy scans or marked clinical improvement was found. Three patients died of progressive disease within 2 months of therapy before follow-up assessments occurred. No adverse events were reported among the 8 surviving patients. The results suggest that rhTSH offers a promising alternative to THST withdrawal to allow radioablative therapy under maximal TSH stimulation in patients with advanced recurrent DTC who would not otherwise be able to receive this treatment. This therapeutic indication extends the clinical potential of the new agent, already demonstrated to be effective for use with I-131 for diagnostic purposes. However in some patients suffering from highly aggressive tumors the poor prognosis will not be improved. (orig.) [German] An unserer Klinik liegen bislang Erfahrungen mit 16 Radioiodtherapien (RIT) (z.T. mehrfache Anwendung) unter rhTSH vor. Die ueberwiegende Mehrzahl der Patienten wurde wegen einer fortgeschrittenen Tumorerkrankung mit dem Risiko einer lebensbedrohlichen Verschlechterung in

  18. Comparison of therapeutic efficacy and clinical parameters between recombinant human thyroid stimulating hormone and thyroid hormone withdrawal in high-dose radioiodine treatment with differentiated thyroid cancer

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Se Hun; Na, Chang Ju; Kim, Jeong Hun; Han, Yeon Hee; KIm, Hee Kwon; Jeong, Hwan Jeong; Sohn, Myung Hee; Lim, Seok Tae [Dept. of Nuclear Medicine, Chonbuk National University Medical School and Hospital, Jeonju (Korea, Republic of)

    2015-06-15

    High-dose radioiodine treatment (HD-RIT) after injection of recombinant human thyroid stimulating hormone (rh-TSH) has become widely used. This study compared the therapeutic efficacy of HD-RIT and clinical parameters between rh-TSH supplement and thyroid hormone withdrawal (THW) after total thyroidectomy in patients with differentiated thyroid cancer. We retrospectively reviewed 266 patients (47 male and 219 female; age, 49.0 ± 10.9 years) with differentiated thyroid cancer detected from September 2011 to September 2012. Patients comprised THW (217, 81.6 %) and rh-TSH (49, 18.4 %). Inclusion criteria were: first HD-RIT; any TN stage; absence of distant metastasis. To evaluate the complete ablation of the remnant thyroid tissue or metastasis, we reviewed stimulated serum thyroglobulin (sTg), I-123 whole-body scan (RxWBS) on T4 off-state, and thyroid ultrasonography (US) or [F-18]-fluorodeoxyglucose positron emission tomography/computed tomography (F-18 FDG PET/CT) 6–8 months after HD-RIT. We defined a complete ablation state when all three of the follow-up conditions were satisfied; <2.0 ng/ml of the sTg, I-123 RxWBS (−), and thyroid US or F-18 FDG PET/CT (−). If one of the three was positive, ablation was considered incomplete. We also compared various clinical biomarkers (body weight, body mass index, liver and kidney function) between THW and rh-TSH groups. The rates of complete ablation were 73.7 % (160/217) for the THW group and 73.5 % (36/49) for the rh-TSH group. There was no significant difference between the two groups (p = 0.970). The follow-up aspartate transaminase (p = 0.001) and alanine transaminase (p = 0.001) were significantly higher in the THW group. The renal function parameters of blood urea nitrogen (p = 0.001) and creatinine (p = 0.005) tended to increase in the THW group. The change of body weight was + Δ0.96 (±1.9) kg for the THW group and was decreased by -Δ1.39 (±1.5) kg for the rh-TSH group. The change

  19. Radioiodine therapy in non-toxic multinodular goitre. The possibility of effect-amplification with recombinant human TSH (rhTSH)

    Energy Technology Data Exchange (ETDEWEB)

    Bonnema, Steen J.; Nielsen, Viveque E.; Hegedues, Laszlo [Odense Univ. Hospital (Denmark). Dept. of Endocrinology and Metabolism

    2006-12-15

    There is no consensus regarding the optimum treatment of benign non-toxic goitre. L-thyroxine suppressive therapy is widely used, but there is poor evidence of its efficacy, and it may have serious adverse effects on health. Surgery is first choice in large goitres or if malignancy is suspected. {sup 131}I therapy results in a one-year goitre reduction of around 40% in multinodular goitres, usually with a high degree of patient satisfaction and improvement of the inspiratory capacity. The effect is attenuated with increasing goitre size. The risk of hypothyroidism is 22-58% within 5-8 years. A sufficient thyroid {sup 131}I uptake is mandatory for {sup 131}I therapy to be feasible and pre-stimulation with recombinant human TSH (rhTSH) increases this considerably. This leads to an increased absorbed thyroid dose by approx.75%, mainly in those patients with the lowest thyroid {sup 131}I uptake, and a more homogeneous intrathyroidal isotope distribution. Pre-stimulation with even a small dose of rhTSH seems to allow a reduction of the {sup 131}I activity while still achieving a mean goitre reduction of approximately 40% within a year. A significantly lower extrathyroidal radiation is achieved by this approach. With an unchanged {sup 131}I activity, rhTSH pre-stimulation improves the goitre reduction by 30-50%. However, this is at the expense of a higher rate of hypothyroidism, cervical pain and transient thyrotoxicosis. Of particular concern is the observation made in healthy persons, that rhTSH results in a transient average thyroid volume increase of 35%. A similar goitre swelling may cause problems in susceptible patients during rhTSH-augmented {sup 131}I therapy. Thus, this concept still needs a closer evaluation before routine use.

  20. Radioiodine 131I metabolism in human

    International Nuclear Information System (INIS)

    Mori, Toru

    1976-01-01

    Metabolic fate of orally administered 131 I in human was studied. Chronological observations of whole body radioactivity distribution and thyroid 131 I uptake curve revealed that 131 I metabolism was greatly affected by the amount of dietary iodine intake. Under the high iodine intake exceeding 1 mg per day, uptake curve showed biphasic descending type, that is, rapid accumulation during 3 to 6 hours and rapid fall up to 48 hours and gradual decrease afterwards. While, ascending type, monophasic and maximal at 24 hours, was found universary under low iodine intake less than 500 μg per day. Thyroid function should not be affected by the amount of iodine intake, and we analysed 131 I metabolism using a new four compartments which included intrathyroidal inorganic iodine pool. The results, especially hormone production rate, were found quite useful even under high iodine intake. Thyroidal organic iodine contents were calculated as approximately 2.5 mg and this value was much less than previously reported values from other countries. Administered radioiodine were mixed up with stable body iodine and reached equilibration by around 10 days. From seroimmunological, histological (microscopic and electron microscopic) studies, and irradiation studies to the cultured human thyroid cells, we concluded that this unexpected phenomenon was derived from chromosomal damage which induced gradual decrease in cell population because of inability to reproduce. Carcinogenic and genetic effects were not serious, and only three leukemic patients were reported in this country and 484 normal babies were born from 7,500 treated parents. Thus, therapeutic dose of 131 I was proved rather safe, and even when exposed to radioiodine, administration of perchlorate or thiocyanate, excessive iodine and TSH seemed effective to avoid radiation injuries. (auth.)

  1. Radioiodine

    International Nuclear Information System (INIS)

    Zaduban, M.

    1978-01-01

    The physical, chemical radiochemical and radiometric properties of radioiodine significant for the radiochemical analysis are summarized. The occurrence of iodine, especially in the biosphere organic part, the calculation of the amount of iodine radioisotopes formed under different conditions, the spectrometric and integral methods of measuring the radioactivity of the most important iodine radioisotopes, the methods of separating iodine and its respective chemical forms are described. (M.K.)

  2. Effect of temperature on the radioiodination of human growth hormone

    International Nuclear Information System (INIS)

    Mohammed-Ali, S.A.; Salacinski, P.R.; Landon, J.

    1981-01-01

    Studies have been undertaken to assess the effect of altering the temperature at which human growth hormone is radioiodinated on the incorporation of 125 I and the immunoreactivity and stability of the labelled hormone. Employing highly purified monomeric hormone it proved possible, by the iodogen procedure, to prepare a labelled product of high specific activity irrespective of temperature. However, in radioiodinations performed at ambient temperature (20 to 25 degrees) significant amounts of the labelled hormone were in an aggregated form which was less immunoreactive than the 125 I-labelled monomeric hormone. Such aggregation was largely prevented by radioiodinating at low temperature (0 to 4 degrees) and even the large monomeric peak was more immunoreactive (about 95% bound in antibody excess) than the monomeric peak from iodinations performed at room temperature

  3. The use of /sup 125/I recombinant DNA/sub 125/ derived human erythropoietin (R-HuEPO) as a replacement for /sup 125/I human urinary epo as tracer antigen in a radioimmunoassay for human epo

    International Nuclear Information System (INIS)

    Cotes, P.M.; Tam, R.C.; GainesDas, R.E.

    1987-01-01

    This paper represents evidence that in a radioimmunoassay for human erythropoietin, recombinant DNA derived human erythropoietin can replace highly purified human urinary erythropoietin in the preparation of radioiodinated tracer antigen

  4. HiLo: Multicentre randomized phase III clinical trial of high vs low dose radioiodine, with or without recombinant human thyroid stimulating hormone (rhTSH), for remnant ablation for differentiated thyroid cancer

    Energy Technology Data Exchange (ETDEWEB)

    Mallick, U. [Freeman Hospital, Newcastle, Newcastle upon Tyne (United Kingdom); Harmer, C.; Clarke, S.; Moss, L.; Nicol, A.; Clarke, P.; Smellie, J.; McCready, R.; Farnell, K.; Franklyn, J.; John, R.; Nutting, C.; Yap, B.; Lemon, C.; Wadlsey, J.; Gerrard, G.; Roques, T.; Macias, E.; Whitaker, S.; Abdul-Hamid, A.; Alvarez, P.; Kadalayil, L.; Hackshaw, A.

    2012-07-01

    Recommended treatment for most patients with differentiated thyroid cancer is surgery followed by radioiodine ablation. Current practice in many centres is to use a high administered activity of 3.7 GBq (100 mCi). However, a lower activity (1.1 GBq or 30 mCi) has advantages including a shorter stay in hospital isolation and lower risk of side effects, including the risk of a second cancer. Also, Thyrogen (rhTSH) allows patients to continue thyroid hormone replacement during ablation, avoiding symptoms of hypothyroidism and also reduces total body radiation dose. We conducted a large randomized factorial multi centre trial to simultaneously address whether ablation success rates are similar using (i) either 1.1 GBq or 3.7 GBq, and (ii) either Thyrogen or thyroid hormone withdrawal. It is the first ever national prospective trial in thyroid cancer in the UK. Final results will be available in 2011

  5. Usefulness of recombinant human TSH-aided radioiodine doses administered in patients with differentiated thyroid carcinoma Administración de dosis terapéuticas de radioyodo luego de TSH recombinante en pacientes con carcinoma diferenciado de tiroides

    Directory of Open Access Journals (Sweden)

    Fabián Pitoia

    2006-04-01

    Full Text Available The published studies confirming the safety and efficacy of rhTSH for diagnostic purposes have led to an increased interest in its use for preparation for radioiodine (RI dose administration in patients with recurrent or persistent differentiated thyroid carcinoma (DTC. In order to establish the efficacy of RI therapy after rhTSH, we have reviewed 39 rhTSH-aided radioiodine treatment in a series of 28 DTC patients. Patients were divided into two groups: GI (n=17, with previous thyroid bed uptake and undetectable thyroglobulin (Tg levels under levothyroxine treatment and GII (n=11, with proven metastatic local or distant disease. Median follow-up after the first rhTSH-aided radioiodine treatment was 32 ± 13 months (range 8 to 54 months. Sixteen patients (94% in GI were rendered disease free and one patient was shown to have persistent disease. In GII, the post therapy whole body scan showed pathological uptakes in all cases: in four patients in lungs, in four in mediastinum and in three in lateral neck. In two patients with mediastinum uptake, Tg levels were undetectable after rhTSH. In the follow-up, two patients with lateral neck uptake were rendered disease free, four patients died (three due to thyroid cancer and five out of the remaining patients have persistent disease. In conclusion, rhTSH aided therapy was helpful to eliminate normal thyroid bed remnants in 16/17 (94% patients (GI. rhTSH stimulated Tg was undetectable in two patients with mediastinal metastasis. We believe that rhTSH is a good alternative to levothyroxine withdrawal for the treatment of DTC with radioactive iodine, increasing the quality of life in these patients. Caution should be recommended in the follow-up of unselected DTC patients only with stimulated Tg levels.Los estudios publicados que confirman la seguridad y eficacia de la TSH recombinante (rhTSH llevaron a un incremento en el interés para su uso como adyuvante terapéutico en el CDT (ablación o tratamiento

  6. Meiotic recombination in human oocytes.

    Directory of Open Access Journals (Sweden)

    Edith Y Cheng

    2009-09-01

    Full Text Available Studies of human trisomies indicate a remarkable relationship between abnormal meiotic recombination and subsequent nondisjunction at maternal meiosis I or II. Specifically, failure to recombine or recombination events located either too near to or too far from the centromere have been linked to the origin of human trisomies. It should be possible to identify these abnormal crossover configurations by using immunofluorescence methodology to directly examine the meiotic recombination process in the human female. Accordingly, we initiated studies of crossover-associated proteins (e.g., MLH1 in human fetal oocytes to analyze their number and distribution on nondisjunction-prone human chromosomes and, more generally, to characterize genome-wide levels of recombination in the human female. Our analyses indicate that the number of MLH1 foci is lower than predicted from genetic linkage analysis, but its localization pattern conforms to that expected for a crossover-associated protein. In studies of individual chromosomes, our observations provide evidence for the presence of "vulnerable" crossover configurations in the fetal oocyte, consistent with the idea that these are subsequently translated into nondisjunctional events in the adult oocyte.

  7. Multinodular goiter treatment with radioiodine aided by recombinant human TSH in different doses: a randomized, double-blind, placebo-controlled study;Administracao previa do TSH humano recombinante, em diferentes doses, no tratamento do bocio multinodular com iodo radioativo: um estudo randomizado, duplo cego, controlado com placebo

    Energy Technology Data Exchange (ETDEWEB)

    Albino, Claudio Cordeiro

    2009-07-01

    Background: There is not an optimal treatment for multinodular goiter (MNG). Surgery is the main therapeutic option because it decreases thyroid volume, reduces compression symptoms and provide histological diagnosis. Radioiodine ({sup 131}I) is an efficient therapeutic option for the treatment of MNG mainly when surgery is not indicated or when the patient refused it. However, high activities of {sup 131}I are frequently required for clinically significant results. This procedure increases the body radiation exposure and the hospitalization costs. Recombinant human TSH (rh TSH) allows a reduction in the administered activity of {sup 131}I with effective thyroid volume (TV) reduction. However, this combination therapeutic can increase collateral effects. Objective: To evaluate the efficacy and safety of low and intermediate doses of rh TSH compared to placebo, associated with a fixed activity of {sup 131}I in MNG treatment. Patients and Methods: Thirty patients with MNG received 0.1 mg of rh TSH (group I, n=10), 0.01 mg of rh TSH (group II, n=10), or placebo (control group, n=10). After 24 hours, 30 mCi of {sup 131}I was given to all patients. Radioactive iodine uptake (RAIU) was determined before and 24 hours after rh TSH. Before and 2, 7, 180 and 360 days after the TV was measured by magnetic resonance image (MRI). The smallest cross-sectional area of tracheal lumen (Scat) was also measured with MRI before, 2 and 7 days after treatment. Antithyroid antibodies, TSH, T3 and free T4 were assessed regularly. Results: After 6 months, the decrease in TV was more significant in groups I (30.3 +- 16.5%) and II (22.6 +- 14.5%), than in control group (5.0 +- 14.6%; p=0.01). After 12 months, TV decreased more in group I (39.2 +- 16.9%) and group II (38.8 +- 24.4%) than in group III (23.4 +- 23.59%) but it was not statistically significant (p=0.205). During the first 30 days,total T3 and free T4 increased, without reaching thyrotoxic levels and TSH decreased. After 12 months

  8. Affinity purification of recombinant human plasminogen activator ...

    African Journals Online (AJOL)

    Affinity purification of recombinant human plasminogen activator from ... Screening antibody was performed using rhPA milk in an ELISA-elution assay. ... useful for purifying other tPA mutants or other novel recombinant milkderived proteins.

  9. Indirect radioiodination of human IgG with N-succinimidyl-3-iodo[125I] benzoate

    International Nuclear Information System (INIS)

    Liu Zhenfeng; Wang Yongxian; Dong Mo; Zhou Wei; Xia Jiaoyun; Yin Duanzhi; Li Linfa

    2007-01-01

    The objective of this study was to develop an acylation method for the radioiodination of monoclonal antibodies that could decrease the loss of radioiodine in vivo. Preparation of N- succinimidyl-3-iodobenzoate(S 125 IB) from the organoth precursor, N-succinimidyl-3-(tri-n-bu- tylstannyl)benzoate(ATE) proceeds in more than 95% labelling yield, when the mass of ATE and NCS are respectively 25-100 μg and 10-20 μg, and the volume of PBS is 10-20 μL, and reaction time is 5 min. IgG is labeled using S 125 IB in up to 75% conjugation efficiency and with well retained immunoreactivity to sheep anti-human IgG. Hepama-1 is also labeled using S 125 IB in more than 75% conjugation efficiency. Paired-label biodistribution studies in normal mice demonstrate that thyroid uptake(a monitor of dehalogenation) of Hepama-1 labeled by S 125 IB method is up to 87.9 times lower than that of Hepama-1 labeled with Iodogen. This result suggests that S 125 IB offers significant advantages for labeling proteins, antibodies over other conventional methods for protein radioiodination. (authors)

  10. Recombinant Human Papillomavirus (HPV) Bivalent Vaccine

    Science.gov (United States)

    This page contains brief information about recombinant human papillomavirus (HPV) bivalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  11. Recombinant Human Papillomavirus (HPV) Nonavalent Vaccine

    Science.gov (United States)

    This page contains brief information about recombinant human papillomavirus (HPV) nonavalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  12. Recombinant Human Papillomavirus (HPV) Quadrivalent Vaccine

    Science.gov (United States)

    This page contains brief information about recombinant human papillomavirus (HPV) quadrivalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  13. New perspectives on recombinant human antibodies

    NARCIS (Netherlands)

    J. de Kruif (John); A.-R. van der Vuurst de Vries (Anne); L. Cilenti (L.); E. Boel (E.); W. van Ewijk (Willem); T. Logtenberg (Ton)

    1996-01-01

    textabstractThe limited potential of murine monoclonal antibodies for human immunotherapy has driven recent progress in recombinant antibody technology. Here, de Kruif and colleagues report on advances in the development and use of phage-antibody-display libraries.

  14. Construction of retroviral recombinant containing human tissue ...

    African Journals Online (AJOL)

    USER

    2010-03-29

    Mar 29, 2010 ... Recombinant retroviral vector containing human tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) gene was ..... heavy metal ions, the protein could be express in an .... involves adhesion, degradation and movement. To.

  15. Small-scale extraction and radioiodination of human hormones for the substitution of imported radioimmunoassay reagents

    International Nuclear Information System (INIS)

    Gimbo, E.K.; Ribela, M.T.C.P.; Borghi, V.C.; Schwarz, I.; Morganti, L.; Araujo, E.A.; Bartolini, P.

    1988-01-01

    The methods for national production of radioimmunoassay reagents to substitute imported kits of: highly purified unlabelled hormones for radioiodination; 125 I-labelled hormones; and specific high titre antisera are presented. The extraction and purification of human growth hormone (hGH) and human luteinizing hormone (hGH) were done from human pituitaries. The 125 I-labelled hormones are obtained by stoichiometric methods. The 125 I-hGH, 125 I-hLH, I-hTSH and 125 I- h calcitonin were prepared and tested in internal and external quality control, in comparison with imported products. The parameters such as: maximum binding to specific antiserum (Bo), nonspecific binding (NSB), mean effective dose (ED 50), sensitivity and accuracy were evaluated. (M.C.K.) [pt

  16. Improvement of reproducibility and quality control of human growth hormone radioiodination

    International Nuclear Information System (INIS)

    Bartolini, P.; Ribela, M.T.C.P.; Camilo, M.A.

    1988-01-01

    The labelling reaction of human growth hormone (hGH) with 125 I and its chromatographic purification have been studied with emphasis on the reproducibility of the yields, quantitaTive recoveries and resulting activities. Through the accurate standardization of a monitoring technique, it is confirmed that there are no significant losses in radioactivity or protein during the labelling or purification process. By strict control of the reaction conditions a fairily good reproducibility is also obtained in the labelling of various hGH extracts with diferent 125 I shipments used after short or long storage. Finally, the specific activity (or absolute mass) of the radioiodinated protein is determined by this Analysis of the Reaction Mixture and compared to the widely used radioimmunological assay (Self-displacement). (M.A.C.) [pt

  17. Radioiodine treatment for malignant thyroid disease

    Energy Technology Data Exchange (ETDEWEB)

    Berg, Gertrud [Sahlgrenska Univ. Hospital, Goeteborg (Sweden). Dept. of Oncology

    2006-12-15

    Radioiodine treatment for thyroid disease has been given for half a decade in Sweden. The most common indication for treatment is hyperthyroidism, when iodine uptake is high. The situation in which radioiodine treatment is used in thyroid cancer is less favourable and measures therefore have to be taken to optimize the treatment. Treatment should be performed early in the course of the disease to achieve the highest possible differentiation. Before treatment the iodine and goitrogen intake should be kept low. Stimulation of the thyrocytes by thyroid-stimulating hormone (TSH) should be high. It is conventionally achieved by thyroid hormone withdrawal rendering the patient hypothyroid, or by the recently available recombinant human TSH (rhTSH) which can be recommended for ablation of the thyroid remnant after thyroidectomy and for treatment of metastases in fragile patients unable to undergo hypothyroidism. Finally, stunning - the negative effect of a prior test dose from radioactive iodine - should be avoided.

  18. Protein Crystal Recombinant Human Insulin

    Science.gov (United States)

    1994-01-01

    The comparison of protein crystal, Recombiant Human Insulin; space-grown (left) and earth-grown (right). On STS-60, Spacehab II indicated that space-grown crystals are larger and of greater optical clarity than their earth-grown counterparts. Recombiant Human Insulin facilitates the incorporation of glucose into cells. In diabetics, there is either a decrease in or complete lack of insulin, thereby leading to several harmful complications. Principal Investigator is Larry DeLucas.

  19. Radioiodinated bleomycin

    International Nuclear Information System (INIS)

    Salmon, S.E.; Liu, R.H.

    1976-01-01

    Radioiodinated bleomycin is a useful imaging agent for body tissues. Its production by iodination of bleomycin with radioactive iodide ions in the presence of an oxidizing agent is described. 7 claims, no drawings

  20. Radioiodine dosimetry

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, J R [Biomedical Research Branch, Health Sciences Div., Chalk River, Ontario (Canada). Chalk River Nuclear Lab.

    1981-01-01

    The estimation of individual doses for radiation protection and for risk assessment purposes from a radioiodine intake requires a knowledge of the distribution and retention of the radioiodine (primarily in the thyroid), and a knowledge of the average energy deposited in each organ of interest per radioactive decay (S-factors). This paper reviews a model for distribution and retention used previously for adults, and extends the model to include all ages. The extended model also includes the effects of stable iodine intakes on radioiodine uptakes explicitly. Included in the paper is a tabulation of existing adult S-factors for selected radioiodines and the extension of S-factors for the thyroid to all age groups. Finally, doses per unit intake are calculated and tabulated. A discussion and some calculations of the effects of stable iodine intake on committed doses are given.

  1. Radioiodine dosimetry

    International Nuclear Information System (INIS)

    Johnson, J.R.

    1981-01-01

    The estimation of individual doses for radiation protection and for risk assessment purposes from a radioiodine intake requires a knowledge of the distribution and retention of the radioiodine (primarily in the thyroid), and a knowledge of the average energy deposited in each organ of interest per radioactive decay (S-factors). This paper reviews a model for distribution and retention used previously for adults, and extends the model to include all ages. The extended model also includes the effects of stable iodine intakes on radioiodine uptakes explicitly. Included in the paper is a tabulation of existing adult S-factors for selected radioiodines and the extension of S-factors for the thyroid to all age groups. Finally, doses per unit intake are calculated and tabulated. A discussion and some calculations of the effects of stable iodine intake on committed doses are given. (author)

  2. Standardization and clinical application of human serum somotomedin B radioimmunoassay. Development of the enzimatic radioiodination

    International Nuclear Information System (INIS)

    Silva Gomes, E.N.D. da.

    1979-01-01

    The various steps that are necessary for the setting up of Somatomedin B radioimunoassay are presented. Radioiodination was carried out through the enzimatic method of Lactoperoxidase while the purification of the labelled was performed first of Sephadex G-25 fine and then on Sephadex G-75. It was found that a preparation of 125 I labelled Somatomedin B presented a radiochemical purity of 99,8%, a mean specific activity determined by the self-displacement method of 54.1 +-3.6μCi/μg and a binding of 98% in the presence of antibody excess. It was also shown that the high stability of this preparation allows its use for up to one month after labelling, if it is stored in refrigerator at + 4 0 C. The separation between the free and the bound antigen was achieved through the method of the second antibody after a period of incubation of 4 days at 4 0 C. The statistic analysis of the method showed also its specificity, which is proved through a study of the dilution effect on human seruns where endogenous Somatomedin B is determined. The sensitivity of the method determined by discrete analysis of the first values on the standard curve was of 0.31 ng/ml. Finally are presented the serum levels of Somatomedin B that have been found in normal subjects and in individuals with alteration HGH secretion [pt

  3. Study on biodistribution and imaging of radioiodinated antisense oligonucleotides in nude mice bearing human lymphoma

    International Nuclear Information System (INIS)

    Wang, R.F.; Shen, J.; Zhang, C.L.; Liu, M.; Guo, F.Q.

    2005-01-01

    The incidence of sporadic lymphoma has risen due to an increase in immunosuppressed patients, particularly those with human immunodeficiency virus (HIV) infection. Sometimes suspect lymphoma has an undetectable location and we can not get the pathological specimen. Management of lymphoma is also difficult because the persistence of a significant number of residual tumor cells after intensive treatment. These relative failures can be attributed to make us choose this study for opening a new diagnostic and therapeutic field of lymphoma from molecular level. Immunoglobulin (Ig) heavy chain framework region (FR) of V1 family have been verified to be a major determinant of malignant phenotype of V1 family B-cell lymphoma. Most of targets for tumor antisense therapy study are protooncogenes, such as c-myc, bc1-2, which are broad -spectrum tumor imaging agents. The aim of this study was to investigate the possibility of using radioiodine labeled FR antisense oligonucleotides (ASONs) as an imaging agent or antisense therapeutic radiopharmaceutical in lymphoma. A 18-mer partial phosphorothioate oligonucleotide sequence was synthesized and grafted in 5 ' with a tyramine group which was further labeled with 125 I or 131 I using the chloramine T method. Normal CD-1 mice were injected via a tail vein with 148 kBq of 125 I-FR-ASON (2∼3 μ g). Animals were sacrificed at 1, 2, 4 and 24 h and tissue samples were studied. Liposome-mediated 3.33 MBq of 131 I-FR-ASON (7 ∼ 9μ g) was injected intratumorally into tumor-bearing BALB/c mice (6 weeks after inoculation of 10 7 Namalwa cells) meanwhile liposome-mediated 131 I labeled sense oligonucleotides served as controls. Biodistribution was monitored by sequential scintigraphy and organ radioactivity measurement 24 h after injection. The percentage of the injected dose per gram (%ID/g) of tumor and tumor/ non-tumor tissue ratios (T/NT) were calculated for each group of mice and the difference between two groups was assessed. The 5

  4. Bayesian inference of shared recombination hotspots between humans and chimpanzees.

    Science.gov (United States)

    Wang, Ying; Rannala, Bruce

    2014-12-01

    Recombination generates variation and facilitates evolution. Recombination (or lack thereof) also contributes to human genetic disease. Methods for mapping genes influencing complex genetic diseases via association rely on linkage disequilibrium (LD) in human populations, which is influenced by rates of recombination across the genome. Comparative population genomic analyses of recombination using related primate species can identify factors influencing rates of recombination in humans. Such studies can indicate how variable hotspots for recombination may be both among individuals (or populations) and over evolutionary timescales. Previous studies have suggested that locations of recombination hotspots are not conserved between humans and chimpanzees. We made use of the data sets from recent resequencing projects and applied a Bayesian method for identifying hotspots and estimating recombination rates. We also reanalyzed SNP data sets for regions with known hotspots in humans using samples from the human and chimpanzee. The Bayes factors (BF) of shared recombination hotspots between human and chimpanzee across regions were obtained. Based on the analysis of the aligned regions of human chromosome 21, locations where the two species show evidence of shared recombination hotspots (with high BFs) were identified. Interestingly, previous comparative studies of human and chimpanzee that focused on the known human recombination hotspots within the β-globin and HLA regions did not find overlapping of hotspots. Our results show high BFs of shared hotspots at locations within both regions, and the estimated locations of shared hotspots overlap with the locations of human recombination hotspots obtained from sperm-typing studies. Copyright © 2014 by the Genetics Society of America.

  5. Binding of radioiodinated human. beta. -endorphin to serum proteins from rats and humans, determined by several methods

    Energy Technology Data Exchange (ETDEWEB)

    Sato, H.; Sugiyama, Y.; Sawada, Y.; Iga, T.; Hanano, M.

    1985-10-07

    Binding of immunoreactive radioiodinated human ..beta..-endorphin (/sup 125/I-..beta..-EP) to rat serum was demonstrated by gel filtration of /sup 125/I-..beta..-EP in pooled rat serum on Sephadex G-200. Two radioactive peaks associated with proteins eluted from the column. The first peak eluted at the void volume containing lipoproteins, ..cap alpha../sub 2/- and ..beta../sub 2/-macroglobulins, and the second peak at the fraction of albumin. Binding of /sup 125/I-..beta..-EP to albumin was directly proved by gel filtration of /sup 125/I-..beta..-EP in buffer containing 4% human serum albumin on Sephadex G-200. Equilibrium dialysis was not applicable to investigating the interaction of /sup 125/I-..beta..-EP with serum proteins, because of the intense nonspecific adsorption to the semi-permeable membrane and the degradation of the peptide during dialysis. Therefore, in order to quantitatively evaluate the binding of /sup 125/I-..beta..-EP in sera from rats and humans, the authors utilized four other methods (ultrafiltration, charcoal adsorption, polyethylene glycol precipitation and equilibrium gel filtration). These methods corresponded well with each other and indicated 35-44% binding of /sup 125/I-..beta..-EP in rat serum. Binding of /sup 125/I-..beta..-EP in normal human serum was 36%, determined by ultrafiltration. Serum protein binding of /sup 125/I-..beta..-EP was concentration independent over the concentration range studied (1-1000 nM). 23 references, 4 figures, 1 table.

  6. Radioiodine therapy

    International Nuclear Information System (INIS)

    Torres, J.F. Jr.; Deliso, H.B.

    1992-01-01

    For over 40 years now, radioiodine ( 131 I) has remained one of the most useful radionuclide for diagnosis and therapy in Nuclear Medicine. The wide application of radioiodine in the study of the thyroid gland and in the management of its disorders has been most rewarding. The medical literature is replete with reports of its efficacy, failures, and complications, but most of these studies have been conducted among Caucasian persons and in relatively affluent societies. Very few reports are available from the less developed and economically depressed areas of the world where thyroid disorders abound or and are even endemic. This chapter is an attempt to highlight the use of radioactive iodine therapy in the developing countries, particularly those in the Asian region

  7. Radioiodine therapy

    Energy Technology Data Exchange (ETDEWEB)

    Torres, Jr, J F; Deliso, H B

    1993-12-31

    For over 40 years now, radioiodine ({sup 131}I) has remained one of the most useful radionuclide for diagnosis and therapy in Nuclear Medicine. The wide application of radioiodine in the study of the thyroid gland and in the management of its disorders has been most rewarding. The medical literature is replete with reports of its efficacy, failures, and complications, but most of these studies have been conducted among Caucasian persons and in relatively affluent societies. Very few reports are available from the less developed and economically depressed areas of the world where thyroid disorders abound or and are even endemic. This chapter is an attempt to highlight the use of radioactive iodine therapy in the developing countries, particularly those in the Asian region

  8. Recombinant human thyrotropin stimulation prior to 131I therapy in toxic multinodular goitre with low radioactive iodine uptake.

    Science.gov (United States)

    Azorín Belda, M J; Martínez Caballero, A; Figueroa Ardila, G C; Martínez Ramírez, M; Gómez Jaramillo, C A; Dolado Ardit, J I; Verdú Rico, J

    Stimulation with recombinant human thyrotropin (rhTSH) increases thyroid radioiodine uptake, and is an aid to 131 I therapy in non-toxic multinodular goitre (MNG). However, there are not many studies using rhTSH prior to 131 I in toxic multinodular goitre to improve hyperthyroidism and compressive symptoms. A prospective study was conducted on patients with MNG and hyperthyroidism. Patients were recruited consecutively and divided into group I, stimulated with 0.3mg of rhTSH before radioiodine therapy, and a control group or group II, without stimulation. Thyroid function, radioiodine thyroid uptake, thyroid weight, and compressive symptoms were measured, and patients were followed-up for 9 months. Group I consisted of 16 patients (14 women), with a mean age 69.7 years, and group II with 16 patients (12 women), with a mean age 70.7 years. After stimulation with 0.3mg rhTSH in group I, 131 I uptake (RAIU) at 24h increased by 78.4%, and the estimated absorbed dose by 89.3%. In group II, the estimated absorbed dose was lower than group I after stimulation with rhTSH (29.8Gy vs. 56.4Gy; P=0.001). At 9 months of follow-up, hyperthyroidism was controlled in 87.5% of patients in group I, and 56.2% in group II (P=0.049). The mean reduction in thyroid weight was higher in group I than in group II (39.3% vs. 26.9%; P=0.017), with a tendency towards subjective improvement of compressive symptoms in group I, although non-significant. Only 2 patients described tachycardias after rhTSH administration, which were resolved with beta-blockers. Stimulation with 0.3mg of recombinant human thyrotropin prior to radioiodine therapy achieves a reduction in thyroid weight and functional improvement in patients with hyperthyroidism and multinodular goitre with low uptake, and with no need for hospital admission. Copyright © 2016 Elsevier España, S.L.U. y SEMNIM. All rights reserved.

  9. Comparative studies of Iodo-bead and chloramine-T methods for the radioiodination of human alpha-fetoprotein

    International Nuclear Information System (INIS)

    Lee, D.S.C.; Griffiths, B.W.

    1984-01-01

    Human alpha-fetoprotein (h-AFP) of iodination grade was radiolabeled with Na 125 I by chloramine-T (conventional method) and by Iodo-bead (solid-phase method) under standard and varied conditions, such as quantity of h-AFP and volume of the reaction mixture. The authors have modified the chloramine-T method and developed a protocol for the Iodo-bead method of iodination. The results show that while the chloramine-T method yields radiolabeled h-AFP of apparently 40% higher specific radioactivity (SR) than those of the Iodo-bead method, the latter appears to be a simpler, more controllable and milder method, giving rise to a more stable population of [ 125 I]h-AFP. This is supported by the results of radioactive counting of [ 125 I]h-AFP collected from a Sephadex G-50 column after radioiodination, storage stability and immunoreactivity with anti-h-AFP. (Auth.)

  10. Immunoreactivity, stability, pharmacokinetics and biodistribution of a monoclonal antibody to human leukemic B cells after three different methods of radioiodination

    International Nuclear Information System (INIS)

    Zhenping Zhu; Ghose, T.; Kralovec, Y.; Chunzheng Yang

    1994-01-01

    Dal B02, a murine monoclonal antibody against human chronic lymphocytic leukemia (CLL) was radioiodinated using chloramine T (Chl.T), Bolton-Hunter (B-H) or N-succinimidyl-p-iodobenzoate (PIB). The preparations had comparable radiochemical purity (>97%) and immunoreactive fraction (65-80%) but the Chl.T-based product was most susceptible to deiodination and loss of immunoreactivity. After i.v. injection into CLL-xenografted nude mice, the preparations had identical patterns of clearance from the blood but the PIB-based product led to more radioactivity in liver and spleen and less in the thyroid compared to the other preparations. The Chl.T-based product showed loss of immunoreactivity in circulation and less tumor-localized radioactivity 168 h after administration. The differences between the B-H-based and PIB-based products were less impressive than between PIB-based and Chl.T-based products. (author)

  11. Recombinant human bone morphogenetic protein induces bone formation

    International Nuclear Information System (INIS)

    Wang, E.A.; Rosen, V.; D'Alessandro, J.S.; Bauduy, M.; Cordes, P.; Harada, T.; Israel, D.I.; Hewick, R.M.; Kerns, K.M.; LaPan, P.; Luxenberg, D.P.; McQuaid, D.; Moutsatsos, I.K.; Nove, J.; Wozney, J.M.

    1990-01-01

    The authors have purified and characterized active recombinant human bone morphogenetic protein (BMP) 2A. Implantation of the recombinant protein in rats showed that a single BMP can induce bone formation in vivo. A dose-response and time-course study using the rat ectopic bone formation assay revealed that implantation of 0.5-115 μg of partially purified recombinant human BMP-2A resulted in cartilage by day 7 and bone formation by day 14. The time at which bone formation occurred was dependent on the amount of BMP-2A implanted; at high doses bone formation could be observed at 5 days. The cartilage- and bone-inductive activity of the recombinant BMP-2A is histologically indistinguishable from that of bone extracts. Thus, recombinant BMP-2A has therapeutic potential to promote de novo bone formation in humans

  12. A Placebo-Controlled, Blinded and Randomised Study on the Effects of Recombinant Human Thyrotropin on Quality of Life in the Treatment of Thyroid Cancer

    DEFF Research Database (Denmark)

    Nygaard, Birte; Bastholt, Lars; Bennedbæk, Finn Noe

    2013-01-01

    BACKGROUND: It is well known that thyroid hormone withdrawal (THW) in thyroid cancer patients can induce a decrease in quality of life (QOL). Recombinant human thyrotropin (rh-TSH) has been used to avoid this; however, no blinded studies have ever documented the effect. OBJECTIVE: To compare QOL...... in patients with differentiated thyroid cancer (DTC) treated with either rh-TSH or liothyronine (L-T3) THW for 10 days. STUDY DESIGN: Double-blind, randomised cross-over. PATIENTS: Fifty-six patients with DTC treated by total thyroidectomy and indication for postsurgery radioiodine (RI) ablation therapy...

  13. Purification of recombinant C-terminus polyhistidine tagged human ...

    African Journals Online (AJOL)

    Dell

    2012-05-03

    May 3, 2012 ... this research, C-terminus polyhistidine tagged human recombinant calcitonin which was ... range protein molecular weight marker was from SIGMA. PCR- ... supernatant was stored at -80°C until needed for further assays.

  14. Recombinant human erythropoietin in sports: a review

    Directory of Open Access Journals (Sweden)

    Rafael Maia de Almeida Bento

    2003-06-01

    Full Text Available Erythropoietin is an endogenous hormone of glicoproteic nature secreted by the kidneys and is the main regulator of the erythropoiesis. An alteration in its production generates a disturbance in the plasmatic concentration giving rise to several types of pathologies related to the hematopoietic system. The recombinant forms of erythropoietin have indiscriminately been used by athletes, mainly in endurance sports, by increasing the erythrocytes concentration, generating a better delivery of oxygen to the muscle tissue. The administration of recombinant erythropoietin was prohibited by the International Olympic Committee and its use considered as doping. This review has the intention to describe the physical, biological and pharmacokinetic properties of the endogenous erythropoietin, as well as its recombinant form, describing also its use in sports and the process of searching methodologies for its detection in doping control.

  15. The influence of recombination on human genetic diversity.

    Directory of Open Access Journals (Sweden)

    Chris C A Spencer

    2006-09-01

    Full Text Available In humans, the rate of recombination, as measured on the megabase scale, is positively associated with the level of genetic variation, as measured at the genic scale. Despite considerable debate, it is not clear whether these factors are causally linked or, if they are, whether this is driven by the repeated action of adaptive evolution or molecular processes such as double-strand break formation and mismatch repair. We introduce three innovations to the analysis of recombination and diversity: fine-scale genetic maps estimated from genotype experiments that identify recombination hotspots at the kilobase scale, analysis of an entire human chromosome, and the use of wavelet techniques to identify correlations acting at different scales. We show that recombination influences genetic diversity only at the level of recombination hotspots. Hotspots are also associated with local increases in GC content and the relative frequency of GC-increasing mutations but have no effect on substitution rates. Broad-scale association between recombination and diversity is explained through covariance of both factors with base composition. To our knowledge, these results are the first evidence of a direct and local influence of recombination hotspots on genetic variation and the fate of individual mutations. However, that hotspots have no influence on substitution rates suggests that they are too ephemeral on an evolutionary time scale to have a strong influence on broader scale patterns of base composition and long-term molecular evolution.

  16. Preparation of a recombinant adenoviral encoding human NIS gene and its specific expression in cardiomyocytes

    International Nuclear Information System (INIS)

    Wang Lihua; Zhang Miao; Guo Rui; Shi Shuo; Li Biao

    2012-01-01

    Objective: To construct a recombinant adenovirus vector containing the human NIS gene with the myosin light chain-2(MLC-2v) gene as the promoter and evaluate its specific expression and feasibility as a reporter gene in cardiomyocytes. Methods: MLC-2v promoter and NIS were subcloned into an adenovirus shuttle vector, and forwarded by homologous recombination in the bacteria BJ5183 containing AdEasy-1 plasmid. Positive recombinant adenovirus vector was selected, packaged and amplified in the HEK293 cells to obtain recombinant adenovirus Ad-MLC-NIS. Ad-cytomegalovirus (CMV)-NIS with cytomegalovirus as the promoter, Ad-MLC without NIS and Ad-NIS without promoter were constructed as the controls. Cardiomyocytes and non-cardiomyocytes were then infected by the adenovirus. The protein expression was tested by Western blot analysis. The function and features of NIS protein were evaluated by dynamic iodide uptake and NaClO 4 iodine uptake inhibition test in vitro. The viability and proliferation of cardiomyocytes after adenovirus transfection and radioiodine incubation were checked by trypan blue staining. Results: Recombinant NIS adenovirus was successfully constructed. Western blot analysis showed that the NIS protein was highly expressed in cardiomyocytes transfected with Ad-MLC-NIS, and all cells transfected with Ad-CMV-NIS. However, in non-cardiomyocytes transfected with Ad-MLC-NIS, little NIS protein was detected. Dynamic iodine uptake tests showed that the peaks of iodide uptake of the three different cell lines (H9C2, A549, U87 cell) transfected with Ad-MLC-NIS were 5844.0, 833.6 and 846.0 counts · min -1 , respectively. The iodide uptake function of H9C2 was inhibited by NaClO 4 . There was almost no change in cell viability and proliferation when the MOI was 100. Conclusions: Ad-MLC-NIS allows myocardial specific expression of an external gene, and the cardiomyocytes with NIS expression are capable of iodine uptake. Further research of NIS as a reporter gene in

  17. Production of Recombinant Adenovirus Containing Human Interlukin-4 Gene

    OpenAIRE

    Mojarrad, Majid; Abdolazimi, Yassan; Hajati, Jamshid; Modarressi, Mohammad Hossein

    2011-01-01

    Objective(s) Recombinant adenoviruses are currently used for a variety of purposes, including in vitro gene transfer, in vivo vaccination, and gene therapy. Ability to infect many cell types, high efficiency in gene transfer, entering both dividing and non dividing cells, and growing to high titers make this virus a good choice for using in various experiments. In the present experiment, a recombinant adenovirus containing human IL-4 coding sequence was made. IL-4 has several characteristics ...

  18. Recombinational DNA repair and human disease

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, Larry H.; Schild, David

    2002-11-30

    We review the genes and proteins related to the homologous recombinational repair (HRR) pathway that are implicated in cancer through either genetic disorders that predispose to cancer through chromosome instability or the occurrence of somatic mutations that contribute to carcinogenesis. Ataxia telangiectasia (AT), Nijmegen breakage syndrome (NBS), and an ataxia-like disorder (ATLD), are chromosome instability disorders that are defective in the ataxia telangiectasia mutated (ATM), NBS, and Mre11 genes, respectively. These genes are critical in maintaining cellular resistance to ionizing radiation (IR), which kills largely by the production of double-strand breaks (DSBs). Bloom syndrome involves a defect in the BLM helicase, which seems to play a role in restarting DNA replication forks that are blocked at lesions, thereby promoting chromosome stability. The Werner syndrome gene (WRN) helicase, another member of the RecQ family like BLM, has very recently been found to help mediate homologous recombination. Fanconi anemia (FA) is a genetically complex chromosomal instability disorder involving seven or more genes, one of which is BRCA2. FA may be at least partially caused by the aberrant production of reactive oxidative species. The breast cancer-associated BRCA1 and BRCA2 proteins are strongly implicated in HRR; BRCA2 associates with Rad51 and appears to regulate its activity. We discuss in detail the phenotypes of the various mutant cell lines and the signaling pathways mediated by the ATM kinase. ATM's phosphorylation targets can be grouped into oxidative stress-mediated transcriptional changes, cell cycle checkpoints, and recombinational repair. We present the DNA damage response pathways by using the DSB as the prototype lesion, whose incorrect repair can initiate and augment karyotypic abnormalities.

  19. Recombinational DNA repair and human disease

    International Nuclear Information System (INIS)

    Thompson, Larry H.; Schild, David

    2002-01-01

    We review the genes and proteins related to the homologous recombinational repair (HRR) pathway that are implicated in cancer through either genetic disorders that predispose to cancer through chromosome instability or the occurrence of somatic mutations that contribute to carcinogenesis. Ataxia telangiectasia (AT), Nijmegen breakage syndrome (NBS), and an ataxia-like disorder (ATLD), are chromosome instability disorders that are defective in the ataxia telangiectasia mutated (ATM), NBS, and Mre11 genes, respectively. These genes are critical in maintaining cellular resistance to ionizing radiation (IR), which kills largely by the production of double-strand breaks (DSBs). Bloom syndrome involves a defect in the BLM helicase, which seems to play a role in restarting DNA replication forks that are blocked at lesions, thereby promoting chromosome stability. The Werner syndrome gene (WRN) helicase, another member of the RecQ family like BLM, has very recently been found to help mediate homologous recombination. Fanconi anemia (FA) is a genetically complex chromosomal instability disorder involving seven or more genes, one of which is BRCA2. FA may be at least partially caused by the aberrant production of reactive oxidative species. The breast cancer-associated BRCA1 and BRCA2 proteins are strongly implicated in HRR; BRCA2 associates with Rad51 and appears to regulate its activity. We discuss in detail the phenotypes of the various mutant cell lines and the signaling pathways mediated by the ATM kinase. ATM's phosphorylation targets can be grouped into oxidative stress-mediated transcriptional changes, cell cycle checkpoints, and recombinational repair. We present the DNA damage response pathways by using the DSB as the prototype lesion, whose incorrect repair can initiate and augment karyotypic abnormalities

  20. Reproducibility of whole-body {sup 131}I scan and serum thyrotropin and stimulated thyroglobulin values in patients studied twice after injection of recombinant human thyrotropin

    Energy Technology Data Exchange (ETDEWEB)

    Niederkohr, Ryan D.; McDougall, I.R. [Stanford University Medical Center, Division of Nuclear Medicine, Stanford, CA (United States)

    2007-03-15

    Recombinant human TSH (rhTSH) is used to increase radioiodine uptake during imaging of thyroid cancer, obviating the need to render the patient hypothyroid. We assessed the reproducibility of radioiodine uptake, serum thyrotropin (TSH), and stimulated serum thyroglobulin (Tg) levels after rhTSH administration. A retrospective review was performed of patients at Stanford who underwent whole-body {sup 131}I scanning for surveillance of thyroid cancer twice after thyroidectomy and {sup 131}I ablation, with rhTSH prior to each scan. Forty-eight hour radioiodine uptake, peak serum TSH, and stimulated serum Tg levels for each study were recorded. Paired t tests and correlation analysis were used to assess interexamination repeatability. Twenty-three patients underwent two scintiscans with rhTSH, for a total of 46 exams. There was no significant difference between percent uptake at 48 h in the paired exams (p=0.40). Serum TSH level was measured in 45 of 46 exams; TSH exceeded 50 mIU/l in all cases, and there was no significant difference between paired TSH levels (p=0.93). All patients had stimulated serum Tg levels measured, with no significant difference between paired Tg levels (p=0.40); after excluding one patient whose Tg changed from 15.8 ng/ml to undetectable between scans without interval treatment, the p value rose to 0.95. There was a strong correlation among paired uptake values (r=0.85, p<0.0001), peak serum TSH (r=0.69, p=0.0003), and stimulated Tg levels (r=0.81, p<0.0001). No discordant scan interpretations were reported. Forty-eight hour radioiodine uptake, peak serum TSH, and stimulated serum Tg levels after administration of rhTSH are repeatable between studies, demonstrating reproducibility of diagnostic results without rendering patients hypothyroid. (orig.)

  1. Humanizing recombinant glycoproteins from Chinese hamster ovary cells

    DEFF Research Database (Denmark)

    Hansen, Anders Holmgaard; Amann, Thomas; Kol, Stefan

    With new tools for gene-editing like zinc-fingers, TALENS and CRISPR, it is now feasible totailor-make the N-Glycoforms for therapeutic glycoproteins that have previously been almost impossible. We here demonstrate a case of humanizing a recombinant human glycoprotein that in Wild type (WT) Chinese...

  2. Physiologically based pharmacokinetics of radioiodinated human beta-endorphin in rats. An application of the capillary membrane-limited model

    Energy Technology Data Exchange (ETDEWEB)

    Sato, H.; Sugiyama, Y.; Sawada, Y.; Iga, T.; Hanano, M.

    1987-07-01

    In order to simulate the distribution and elimination of radioiodinated human beta-endorphin (/sup 125/I-beta-EP) after iv bolus injection in rats, we proposed a physiologically based pharmacokinetic model incorporating diffusional transport of /sup 125/I-beta-EP across the capillary membrane. This model assumes that the distribution of /sup 125/I-beta-EP is restricted only within the blood and the tissue interstitial fluid, and that a diffusional barrier across the capillary membrane exists in each tissue except the liver. The tissue-to-blood partition coefficients were estimated from the ratios of the concentration in tissues to that in arterial plasma at the terminal (pseudoequilibrium) phase. The total body plasma clearance (9.0 ml/min/kg) was appropriately assigned to the liver and kidney. The transcapillary diffusion clearances of /sup 125/I-beta-EP were also estimated and shown to correlate linearly with that of inulin in several tissues. Numerically solving the mass-balance differential equations as to plasma and each tissue simultaneously, simulated concentration curves of /sup 125/I-beta-EP corresponded well with the observed data. It was suggested by the simulation that the initial rapid disappearance of /sup 125/I-beta-EP from plasma after iv injection could be attributed in part to the transcapillary diffusion of the peptide.

  3. Ovarian response to recombinant human follicle-stimulating hormone

    DEFF Research Database (Denmark)

    Arce, Joan-Carles; Andersen, Anders Nyboe; Fernández-Sánchez, Manuel

    2014-01-01

    OBJECTIVE: To evaluate the dose-response relationship of a novel recombinant human FSH (rhFSH; FE 999049) with respect to ovarian response in patients undergoing IVF/intracytoplasmic sperm injection treatment; and prospectively study the influence of initial antimüllerian hormone (AMH) concentrat......OBJECTIVE: To evaluate the dose-response relationship of a novel recombinant human FSH (rhFSH; FE 999049) with respect to ovarian response in patients undergoing IVF/intracytoplasmic sperm injection treatment; and prospectively study the influence of initial antimüllerian hormone (AMH...

  4. Therapeutic implications of recombinant human erythropoietin in ...

    African Journals Online (AJOL)

    AJB SERVER

    2006-12-29

    Dec 29, 2006 ... quence of both, RHUEPO has achieved the highest annual sales ... analysis of the US Medicare database (Ma et al., 1999) ... blood transfusions and improves quality of life (Eschbach, ... Large doses of EPO results increase in blood pressure .... human erythropoietin was obtained from human genomic.

  5. Termini of human chromosomes display elevated rates of mitotic recombination.

    Science.gov (United States)

    Cornforth, M N; Eberle, R L

    2001-01-01

    The strand-specific in situ hybridization technique of CO-FISH was used to probe telomeres of human mitotic cells in order to determine the spontaneous frequency of crossover. This approach allowed the detection of recombinational crossovers occurring anywhere along the length of individual chromosomes, including reciprocal events taking place between sister chromatids. Although the process of sister chromatid exchange (SCE) is the most prominent type of recombination in somatic mammalian cells, our results show that SCEs accounted for less than a third of the recombinational events revealed by CO-FISH. It is concluded that chromosomal regions near the termini of chromosome arms undergo extraordinarily high rates of spontaneous recombination, producing terminal crossovers whose small size precludes detection by standard cytogenetic methods. That similar results were observed for transformed epithelial cells, as well as primary fibroblasts, suggests that the phenomenon is a common characteristic of human cells. These findings are noteworthy because, although telomeric and subtelomeric DNA is known to be preferentially involved in certain types of recombination, the tips of somatic mammalian chromosomes have not previously been identified as preferred sites for crossover. Implications of these results are discussed in terms of limitations imposed on CO-FISH for its proposed use in directional hybridization mapping.

  6. Recombination in the human Pseudoautosomal region PAR1.

    Directory of Open Access Journals (Sweden)

    Anjali G Hinch

    2014-07-01

    Full Text Available The pseudoautosomal region (PAR is a short region of homology between the mammalian X and Y chromosomes, which has undergone rapid evolution. A crossover in the PAR is essential for the proper disjunction of X and Y chromosomes in male meiosis, and PAR deletion results in male sterility. This leads the human PAR with the obligatory crossover, PAR1, to having an exceptionally high male crossover rate, which is 17-fold higher than the genome-wide average. However, the mechanism by which this obligatory crossover occurs remains unknown, as does the fine-scale positioning of crossovers across this region. Recent research in mice has suggested that crossovers in PAR may be mediated independently of the protein PRDM9, which localises virtually all crossovers in the autosomes. To investigate recombination in this region, we construct the most fine-scale genetic map containing directly observed crossovers to date using African-American pedigrees. We leverage recombination rates inferred from the breakdown of linkage disequilibrium in human populations and investigate the signatures of DNA evolution due to recombination. Further, we identify direct PRDM9 binding sites using ChIP-seq in human cells. Using these independent lines of evidence, we show that, in contrast with mouse, PRDM9 does localise peaks of recombination in the human PAR1. We find that recombination is a far more rapid and intense driver of sequence evolution in PAR1 than it is on the autosomes. We also show that PAR1 hotspot activities differ significantly among human populations. Finally, we find evidence that PAR1 hotspot positions have changed between human and chimpanzee, with no evidence of sharing among the hottest hotspots. We anticipate that the genetic maps built and validated in this work will aid research on this vital and fascinating region of the genome.

  7. Potential use of recombinant human interleukin-6 in clinical oncology

    NARCIS (Netherlands)

    Veldhuis, GJ; Willemse, PHB; Mulder, NH; Limburg, PC; deVries, EGE

    Recombinant human IL-6 (rhIL-6) is a pleiotropic cytokine with stimulatory actions on the hematopoietic system, the immune system and hepatocytes. Clinical interest in the use of this cytokine was raised because of its thrombopoietic properties and also because of its anti-tumor activity, which was

  8. Immunological aspects of antibody formation against recombinant human therapeutics

    NARCIS (Netherlands)

    Sauerborn, M.S.

    2010-01-01

    With about 200 new products in the pipeline, recombinant human (rh) therapeutics are becoming the most dominant class of drugs. One of the reasons to create rh therapeutics was to avoid recognition by the immune system due to foreign origin. Nevertheless, rh therapeutics induced formation of

  9. Antiproliferative activity of recombinant human interferon-λ2 ...

    African Journals Online (AJOL)

    Antiproliferative activity of recombinant human interferon-λ2 expressed in stably ... The representing 26 kDa protein band of IFN-λ2 was detected by SDS-PAGE and ... The antiproliferative activity of hIFN-λ2 was determined by MTT assay.

  10. Haemostatic aspects of recombinant human erythropoietin in colorectal surgery

    DEFF Research Database (Denmark)

    Poulsen, K A; Qvist, N; Winther, K

    1998-01-01

    OBJECTIVE: To find out whether recombinant human erythropoietin (r-HuEPO) given perioperatively has any effect on haemostatic activity in patients undergoing elective colorectal resection. DESIGN: A placebo-controlled double-blind study. SETTING: Odense university hospital, Denmark. SUBJECTS: 24...

  11. Recombiner

    International Nuclear Information System (INIS)

    Kikuchi, Nobuo.

    1983-01-01

    Purpose: To shorten the pre-heating time for a recombiner and obtain a uniform temperature distribution for the charged catalyst layer in a BWR type reactor. Constitution: A pre-heating heater is disposed to the outer periphery of a vessel for a recombiner packed with catalysts for recombining hydrogen and oxygen in gases flowing through a radioactive gaseous wastes processing system. Heat pipes for transmitting the heat applied to said container to the catalyst are disposed vertically and horizontally within the container. Different length of the heat pipes are combined. In this way, pre-heating time for the recombiner before the operation start and before the system switching can be shortened and the uniform pre-heating for the inside of the recombiner is also made possible. Further, heater control in the pre-heating can be carried out effectively and with ease. (Moriyama, K.)

  12. Expression and characterization of recombinant human serum ...

    African Journals Online (AJOL)

    C-peptide (CP), connecting the A and B chains in proinsulin, has been considered to possess physiological effects in diabetes. In order to prolong the half-life of CP in vivo, a long acting CP analog [human serum albumin (HSA-CP)] was obtained by direct gene fusion of a single-chain CP to HSA and expressed in host ...

  13. Expression and characterization of recombinant human serum ...

    African Journals Online (AJOL)

    ajl yemi

    2011-11-14

    Nov 14, 2011 ... C-peptide (CP), connecting the A and B chains in proinsulin, has been considered to possess physiological effects in diabetes. In order to prolong the half-life of CP in vivo, a long acting CP analog. [human serum albumin (HSA-CP)] was obtained by direct gene fusion of a single-chain CP to HSA and.

  14. A review of age dependent radioiodine dosimetry

    International Nuclear Information System (INIS)

    Johnson, J.R.

    1987-01-01

    Age dependent models of radioiodine metabolism in humans have been described. These models have been used to calculate age dependent committed doses to the thyroid from intakes of radioiodines. A model of fetal iodine metabolism is also described and used to calculate fetal thyroid doses from intakes of radioiodines by the mother. These doses are calculated using model parameter values thought to be representative of average for North American/European populations. Considerable variability from these results can be expected for individuals. In addition, population with significant differences in stable iodine intake, and in body parameters, will have model parameters somewhat different than the ones described in this paper. These different model parameters will result in different doses from intakes of radioiodines, but it is doubtful if the differences in population would be as large as the variation between individuals. 25 refs.; 11 figs.; 1 table

  15. Structural analysis of recombinant human protein QM

    International Nuclear Information System (INIS)

    Gualberto, D.C.H.; Fernandes, J.L.; Silva, F.S.; Saraiva, K.W.; Affonso, R.; Pereira, L.M.; Silva, I.D.C.G.

    2012-01-01

    Full text: The ribosomal protein QM belongs to a family of ribosomal proteins, which is highly conserved from yeast to humans. The presence of the QM protein is necessary for joining the 60S and 40S subunits in a late step of the initiation of mRNA translation. Although the exact extra-ribosomal functions of QM are not yet fully understood, it has been identified as a putative tumor suppressor. This protein was reported to interact with the transcription factor c-Jun and thereby prevent c-Jun actives genes of the cellular growth. In this study, the human QM protein was expressed in bacterial system, in the soluble form and this structure was analyzed by Circular Dichroism and Fluorescence. The results of Circular Dichroism showed that this protein has less alpha helix than beta sheet, as described in the literature. QM protein does not contain a leucine zipper region; however the ion zinc is necessary for binding of QM to c-Jun. Then we analyzed the relationship between the removal of zinc ions and folding of protein. Preliminary results obtained by the technique Fluorescence showed a gradual increase in fluorescence with the addition of increasing concentration of EDTA. This suggests that the zinc is important in the tertiary structure of the protein. More studies are being made for better understand these results. (author)

  16. 76 FR 65210 - Certain Products and Pharmaceutical Compositions Containing Recombinant Human Erythropoetin...

    Science.gov (United States)

    2011-10-20

    ... INTERNATIONAL TRADE COMMISSION [Investigation No. 337-TA-568] Certain Products and Pharmaceutical Compositions Containing Recombinant Human Erythropoetin; Termination of Investigation on the Basis of... after importation of certain products and pharmaceutical compositions containing recombinant human...

  17. Variation in human recombination rates and its genetic determinants.

    Directory of Open Access Journals (Sweden)

    Adi Fledel-Alon

    Full Text Available Despite the fundamental role of crossing-over in the pairing and segregation of chromosomes during human meiosis, the rates and placements of events vary markedly among individuals. Characterizing this variation and identifying its determinants are essential steps in our understanding of the human recombination process and its evolution.Using three large sets of European-American pedigrees, we examined variation in five recombination phenotypes that capture distinct aspects of crossing-over patterns. We found that the mean recombination rate in males and females and the historical hotspot usage are significantly heritable and are uncorrelated with one another. We then conducted a genome-wide association study in order to identify loci that influence them. We replicated associations of RNF212 with the mean rate in males and in females as well as the association of Inversion 17q21.31 with the female mean rate. We also replicated the association of PRDM9 with historical hotspot usage, finding that it explains most of the genetic variance in this phenotype. In addition, we identified a set of new candidate regions for further validation.These findings suggest that variation at broad and fine scales is largely separable and that, beyond three known loci, there is no evidence for common variation with large effects on recombination phenotypes.

  18. Recombiner

    International Nuclear Information System (INIS)

    Osumi, Morimichi.

    1979-01-01

    Purpose: To provide a recombiner which is capable of converting hydrogen gas into water by use of high-frequency heating at comparatively low temperatures and is safe and cheap in cost. Constitution: Hydrogen gas is introduced from an outer pipeline to the main structure of a recombiner, and when it passes through the vicinity of the central part of the recombiner, it is reacted with copper oxide (CuO 2 ) heated to a temperature more than 300 0 C by a high-frequency heater, and converted gently into water by reduction operation (2H 2 + CuO 2 → Cu + 2H 2 O). The thus prepared water is exhausted through the outer pipeline to a suppression pool. A part of hydrogen gas which has not been converted completely into water by the reaction and is remaining as hydrogen is recovered through exhaust nozzles and again introduced into the main structure of the recombiner. (Yoshino, Y.)

  19. Similarity of recombinant human perlecan domain 1 by alternative expression systems bioactive heterogenous recombinant human perlecan D1

    DEFF Research Database (Denmark)

    Ellis, April L; Pan, Wensheng; Yang, Guang

    2010-01-01

    BACKGROUND: Heparan sulfate glycosaminoglycans are diverse components of certain proteoglycans and are known to interact with growth factors as a co-receptor necessary to induce signalling and growth factor activity. In this report we characterize heterogeneously glycosylated recombinant human...... perlecan domain 1 (HSPG2 abbreviated as rhPln.D1) synthesized in either HEK 293 cells or HUVECs by transient gene delivery using either adenoviral or expression plasmid technology. RESULTS: By SDS-PAGE analysis following anion exchange chromatography, the recombinant proteoglycans appeared to possess...... glycosaminoglycan chains ranging, in total, from 6 kDa to >90 kDa per recombinant. Immunoblot analysis of enzyme-digested high Mr rhPln.D1 demonstrated that the rhPln.D1 was synthesized as either a chondroitin sulfate or heparan sulfate proteoglycan, in an approximately 2:1 ratio, with negligible hybrids. Secondary...

  20. Human DNA repair and recombination genes

    International Nuclear Information System (INIS)

    Thompson, L.H.; Weber, C.A.; Jones, N.J.

    1988-09-01

    Several genes involved in mammalian DNA repair pathways were identified by complementation analysis and chromosomal mapping based on hybrid cells. Eight complementation groups of rodent mutants defective in the repair of uv radiation damage are now identified. At least seven of these genes are probably essential for repair and at least six of them control the incision step. The many genes required for repair of DNA cross-linking damage show overlap with those involved in the repair of uv damage, but some of these genes appear to be unique for cross-link repair. Two genes residing on human chromosome 19 were cloned from genomic transformants using a cosmid vector, and near full-length cDNA clones of each gene were isolated and sequenced. Gene ERCC2 efficiently corrects the defect in CHO UV5, a nucleotide excision repair mutant. Gene XRCC1 normalizes repair of strand breaks and the excessive sister chromatid exchange in CHO mutant EM9. ERCC2 shows a remarkable /approximately/52% overall homology at both the amino acid and nucleotide levels with the yeast RAD3 gene. Evidence based on mutation induction frequencies suggests that ERCC2, like RAD3, might also be an essential gene for viability. 100 refs., 4 tabs

  1. Recombinant human TSH in differentiated thyroid cancer: a nuclear medicine perspective

    Energy Technology Data Exchange (ETDEWEB)

    Zanotti-Fregonara, P. [CEA, DSV, I2BM, SHFJ, LMNRB, Orsay (France); Rubello, D. [Osped S Maria Misericordia, IRCCS, IOV, Dept Nucl Med, PET Ctr, I-45100 Rovigo (Italy); Hindie, E. [Hop St Louis, Dept Nucl Med, Paris (France)

    2008-07-01

    The use of recombinant human thyroid-stimulating hormone (rhTSH) in differentiated thyroid cancer (DTC) is widely discussed in the literature with regard to the diagnostic and therapeutic aspects of the management of DTC patients. However, some controversy about the appropriate indications, advantages and potential disadvantages of the use of rhTSH may still exist within the community of nuclear medicine physicians. In our opinion, the clinical benefits of rhTSH in avoiding hypothyroidism outweigh its somewhat lesser diagnostic accuracy. However, we disagree on designating rhTSH as the 'golden standard' to obtain TSH stimulation, as suggested by some authors. Thus, the first follow-up examination after ablation, which is determinant for patients' prognostic classification, can be either done under rhTSH stimulation or after hormone withdrawal. In our practice, and for higher risk patients, we still favour performing the initial follow-up after thyroid hormone withdrawal. rhTSH also shows the ability to enhance radioiodine concentration into thyroid cells. This characteristic is obviously of great interest among the nuclear medicine community. In clinical practice, it seems preferable to perform {sup 131}I treatment for metastatic disease during hypothyroidism. rhTSH may find its utility for the treatment of specific populations of patients, i.e. those in whom hormone withdrawal is medically contraindicated or in whom adequate endogenous TSH levels cannot be obtained due to reduced pituitary reserve or continued thyroxine production by metastatic tissue. In conclusion, rhTSH has demonstrated to be a reliable alternative to hypothyroidism for the stimulation of Tg in the follow-up of thyroid cancer patients. However, its use must be more carefully chosen in the therapeutic setting. Our feeling is that rhTSH should no tbe used for remnant ablation in high-risk patients and for the treatment of metastatic disease, except for specific populations of

  2. Recombinant human TSH in differentiated thyroid cancer: a nuclear medicine perspective

    International Nuclear Information System (INIS)

    Zanotti-Fregonara, P.; Rubello, D.; Hindie, E.

    2008-01-01

    The use of recombinant human thyroid-stimulating hormone (rhTSH) in differentiated thyroid cancer (DTC) is widely discussed in the literature with regard to the diagnostic and therapeutic aspects of the management of DTC patients. However, some controversy about the appropriate indications, advantages and potential disadvantages of the use of rhTSH may still exist within the community of nuclear medicine physicians. In our opinion, the clinical benefits of rhTSH in avoiding hypothyroidism outweigh its somewhat lesser diagnostic accuracy. However, we disagree on designating rhTSH as the 'golden standard' to obtain TSH stimulation, as suggested by some authors. Thus, the first follow-up examination after ablation, which is determinant for patients' prognostic classification, can be either done under rhTSH stimulation or after hormone withdrawal. In our practice, and for higher risk patients, we still favour performing the initial follow-up after thyroid hormone withdrawal. rhTSH also shows the ability to enhance radioiodine concentration into thyroid cells. This characteristic is obviously of great interest among the nuclear medicine community. In clinical practice, it seems preferable to perform 131 I treatment for metastatic disease during hypothyroidism. rhTSH may find its utility for the treatment of specific populations of patients, i.e. those in whom hormone withdrawal is medically contraindicated or in whom adequate endogenous TSH levels cannot be obtained due to reduced pituitary reserve or continued thyroxine production by metastatic tissue. In conclusion, rhTSH has demonstrated to be a reliable alternative to hypothyroidism for the stimulation of Tg in the follow-up of thyroid cancer patients. However, its use must be more carefully chosen in the therapeutic setting. Our feeling is that rhTSH should no tbe used for remnant ablation in high-risk patients and for the treatment of metastatic disease, except for specific populations of patients. (O.M.)

  3. Biophysical characterisation of GlycoPEGylated recombinant human factor VIIa

    DEFF Research Database (Denmark)

    Plesner, Bitten; Westh, Peter; Nielsen, Anders D.

    2011-01-01

    The effects of GlycoPEGylation on the structural, kinetic and thermal stability of recombinant human FVIIa were investigated using rFVIIa and linear 10 kDa and branched 40 kDa GlycoPEGylated® recombinant human FVIIa derivatives. The secondary and tertiary structure of rFVIIa measured by circular...... dichroism (CD) was maintained upon PEGylation. In contrast, the thermal and kinetic stability of rFVIIa was affected by GlycoPEGylation, as the apparent unfolding temperature Tm measured by differential scanning calorimetry (DSC) and the temperature of aggregation, Tagg, measured by light scattering (LS......) both increased with GlycoPEGylation. Both Tm and Tagg were independent of the molecular weight and the shape of the PEG chain. From the present biophysical characterisation it is concluded that after GlycoPEGylation, rFVIIa appears to be unaffected structurally (secondary and tertiary structure...

  4. Radioimmunoassay for the middle region of human parathyroid hormone: comparison of two radioiodinated synthetic peptides

    International Nuclear Information System (INIS)

    Sharp, M.E.; Marx, S.J.

    1985-01-01

    Two synthetic peptides were evaluated to develop radioligands for midregion-specific radioimmunoassay (RIA) of human parathyroid hormone (hPTH). Both radioligands were tested using three anti-PTH sera of proven clinical utility. While each of these midregion-directed antisera showed unique specificity, they all reacted with high affinity with both radioligands and none of them discriminated significantly between the two synthetic midregion peptides. Analysis of data on the relation of serum calcium and hPTH midregion immunoreactivity showed a useful separation of groups (all nonazotemic) with primary hyperparathyroidism, secondary hyperparathyroidism, primary hypoparathyroidism and secondary hypoparathyroidism. (Auth.)

  5. Posttranslational modifications in human plasma MBL and human recombinant MBL

    DEFF Research Database (Denmark)

    Jensen, Pia Hønnerup; Laursen, Inga; Matthiesen, Finn

    2007-01-01

    the intact protein in its active conformation. For the first time, positions and occupation frequency of partial hydroxylations and partial glycosylations are reported in MBL. Hydroxylation and glycosylation patterns of both recombinant and plasma derived MBL were determined, using a combination of mass......Mannan-binding lectin (MBL) is a complex serum protein that plays an important role in innate immunity. In addition to assuming several different oligomeric forms, the polypeptide itself is highly heterogeneous. This heterogeneity is due to post-translational modifications, which help to stabilize......(202)) was modified in trace amounts to dehydroalanine, as detected by a 34 Da mass loss. This impairs the formation of a disulphide bond in the carbohydrate recognition domain. The dehydroalanine was identified in similar small amounts in both recombinant and plasma-derived MBL....

  6. The rise and fall of a human recombination hot spot.

    Science.gov (United States)

    Jeffreys, Alec J; Neumann, Rita

    2009-05-01

    Human meiotic crossovers mainly cluster into narrow hot spots that profoundly influence patterns of haplotype diversity and that may also affect genome instability and sequence evolution. Hot spots also seem to be ephemeral, but processes of hot-spot activation and their subsequent evolutionary dynamics remain unknown. We now analyze the life cycle of a recombination hot spot. Sperm typing revealed a polymorphic hot spot that was activated in cis by a single base change, providing evidence for a primary sequence determinant necessary, though not sufficient, to activate recombination. This activating mutation occurred roughly 70,000 y ago and has persisted to the present, most likely fortuitously through genetic drift despite its systematic elimination by biased gene conversion. Nonetheless, this self-destructive conversion will eventually lead to hot-spot extinction. These findings define a subclass of highly transient hot spots and highlight the importance of understanding hot-spot turnover and how it influences haplotype diversity.

  7. Chemical forms of radioiodine

    International Nuclear Information System (INIS)

    Tachikawa, Enzo

    1979-01-01

    Release of radioiodine built-up during reactor operations presents a potential problem from the standpoint of environmental safety. Among the chemical forms of radioiodine, depending upon the circumstances, organic iodides cast a most serious problem because of its difficulties in the trapping and because of its stability compared to other chemical forms. Furthermore, pellet-cladding interaction (PCl) fuel failures in LWR fuel rods are believed to be stress corrosion cracks caused by embrittling fission product species, radioiodine. To deal with these problems, knowledge is required on the chemical behaviors of radioiodine in and out of fuels, as well as the release behaviors from fuels. Here a brief review is given of these respects, in aiming at clearing-up the questions still remaining unknown. The data seem to indicate that radioiodine exists as a combined form in fuels. upon heating slightly irradiated fuels, the iodine atoms are released in a chemical form associated with uranium atoms. Experiments, however, as needed with specimen of higher burnup, where the interactions of radioiodine with metallic fission products could be favored. The dominant release mechanism of radioiodine under normal operating temperatures will be diffusion to grain boundaries leading to open surfaces. Radiation-induced internal traps, however, after the rate of diffusion significantly. The carbon sources of organic iodides formed under various conditions and its formation mechanisms have also been considered. (author)

  8. Production of biologically active recombinant human factor H in Physcomitrella.

    Science.gov (United States)

    Büttner-Mainik, Annette; Parsons, Juliana; Jérôme, Hanna; Hartmann, Andrea; Lamer, Stephanie; Schaaf, Andreas; Schlosser, Andreas; Zipfel, Peter F; Reski, Ralf; Decker, Eva L

    2011-04-01

    The human complement regulatory serum protein factor H (FH) is a promising future biopharmaceutical. Defects in the gene encoding FH are associated with human diseases like severe kidney and retinal disorders in the form of atypical haemolytic uremic syndrome (aHUS), membranoproliferative glomerulonephritis II (MPGN II) or age-related macular degeneration (AMD). There is a current need to apply intact full-length FH for the therapy of patients with congenital or acquired defects of this protein. Application of purified or recombinant FH (rFH) to these patients is an important and promising approach for the treatment of these diseases. However, neither protein purified from plasma of healthy individuals nor recombinant protein is currently available on the market. Here, we report the first stable expression of the full-length human FH cDNA and the subsequent production of this glycoprotein in a plant system. The moss Physcomitrella patens perfectly suits the requirements for the production of complex biopharmaceuticals as this eukaryotic system not only offers an outstanding genetical accessibility, but moreover, proteins can be produced safely in scalable photobioreactors without the need for animal-derived medium compounds. Transgenic moss lines were created, which express the human FH cDNA and target the recombinant protein to the culture supernatant via a moss-derived secretion signal. Correct processing of the signal peptide and integrity of the moss-produced rFH were verified via peptide mapping by mass spectrometry. Ultimately, we show that the rFH displays complement regulatory activity comparable to FH purified from plasma. © 2010 The Authors. Plant Biotechnology Journal © 2010 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  9. Recombiner

    International Nuclear Information System (INIS)

    Saalfrank, H.

    1985-01-01

    Air containing hydrogen can be oxidized by heating in a container called a recombiner, in order to avoid the collection of hydrogen. The container is long and a large number of straight heating bars are arranged in parallel in it and they are flanged to a lid. The heating bars are surrounded by tubes, in order to obtain good heat transfer by a narrow annular gap. (orig.) [de

  10. Radioiodine in soils

    International Nuclear Information System (INIS)

    Szabova, T.

    1981-01-01

    Behaviour was studied of radioiodine in different soil subtypes sampled in the vicinity of the A-1 nuclear power plant at Jaslovske Bohunice. Radioiodine sorption is mainly affected by the amount of humus and by the clay fraction of soil. The highest sorption was recorded for meadow chernozem and the lowest for rendzina soils. At the same pH, soils with a higher level of organic matter adsorb more radioiodine. Upon applying radioiodate in soil samples, reduction to iodide takes place. Under the action of anions, radioiodine sorption decreases, the effect decreasing as follows: H 2 PO 4 - >SO 2- >Cl - >NO 3 - . The highest desorption was caused by 0.1 N NaOH solution. (author)

  11. Broad-scale recombination patterns underlying proper disjunction in humans.

    Directory of Open Access Journals (Sweden)

    Adi Fledel-Alon

    2009-09-01

    Full Text Available Although recombination is essential to the successful completion of human meiosis, it remains unclear how tightly the process is regulated and over what scale. To assess the nature and stringency of constraints on human recombination, we examined crossover patterns in transmissions to viable, non-trisomic offspring, using dense genotyping data collected in a large set of pedigrees. Our analysis supports a requirement for one chiasma per chromosome rather than per arm to ensure proper disjunction, with additional chiasmata occurring in proportion to physical length. The requirement is not absolute, however, as chromosome 21 seems to be frequently transmitted properly in the absence of a chiasma in females, a finding that raises the possibility of a back-up mechanism aiding in its correct segregation. We also found a set of double crossovers in surprisingly close proximity, as expected from a second pathway that is not subject to crossover interference. These findings point to multiple mechanisms that shape the distribution of crossovers, influencing proper disjunction in humans.

  12. Recombinant human prion protein inhibits prion propagation in vitro.

    Science.gov (United States)

    Yuan, Jue; Zhan, Yi-An; Abskharon, Romany; Xiao, Xiangzhu; Martinez, Manuel Camacho; Zhou, Xiaochen; Kneale, Geoff; Mikol, Jacqueline; Lehmann, Sylvain; Surewicz, Witold K; Castilla, Joaquín; Steyaert, Jan; Zhang, Shulin; Kong, Qingzhong; Petersen, Robert B; Wohlkonig, Alexandre; Zou, Wen-Quan

    2013-10-09

    Prion diseases are associated with the conformational conversion of the cellular prion protein (PrP(C)) into the pathological scrapie isoform (PrP(Sc)) in the brain. Both the in vivo and in vitro conversion of PrP(C) into PrP(Sc) is significantly inhibited by differences in amino acid sequence between the two molecules. Using protein misfolding cyclic amplification (PMCA), we now report that the recombinant full-length human PrP (rHuPrP23-231) (that is unglycosylated and lacks the glycophosphatidylinositol anchor) is a strong inhibitor of human prion propagation. Furthermore, rHuPrP23-231 also inhibits mouse prion propagation in a scrapie-infected mouse cell line. Notably, it binds to PrP(Sc), but not PrP(C), suggesting that the inhibitory effect of recombinant PrP results from blocking the interaction of brain PrP(C) with PrP(Sc). Our findings suggest a new avenue for treating prion diseases, in which a patient's own unglycosylated and anchorless PrP is used to inhibit PrP(Sc) propagation without inducing immune response side effects.

  13. Genetic recombination between human and animal parasites creates novel strains of human pathogen.

    Directory of Open Access Journals (Sweden)

    Wendy Gibson

    2015-03-01

    Full Text Available Genetic recombination between pathogens derived from humans and livestock has the potential to create novel pathogen strains, highlighted by the influenza pandemic H1N1/09, which was derived from a re-assortment of swine, avian and human influenza A viruses. Here we investigated whether genetic recombination between subspecies of the protozoan parasite, Trypanosoma brucei, from humans and animals can generate new strains of human pathogen, T. b. rhodesiense (Tbr responsible for sleeping sickness (Human African Trypanosomiasis, HAT in East Africa. The trait of human infectivity in Tbr is conferred by a single gene, SRA, which is potentially transferable to the animal pathogen Tbb by sexual reproduction. We tracked the inheritance of SRA in crosses of Tbr and Tbb set up by co-transmitting genetically-engineered fluorescent parental trypanosome lines through tsetse flies. SRA was readily transferred into new genetic backgrounds by sexual reproduction between Tbr and Tbb, thus creating new strains of the human pathogen, Tbr. There was no evidence of diminished growth or transmissibility of hybrid trypanosomes carrying SRA. Although expression of SRA is critical to survival of Tbr in the human host, we show that the gene exists as a single copy in a representative collection of Tbr strains. SRA was found on one homologue of chromosome IV in the majority of Tbr isolates examined, but some Ugandan Tbr had SRA on both homologues. The mobility of SRA by genetic recombination readily explains the observed genetic variability of Tbr in East Africa. We conclude that new strains of the human pathogen Tbr are being generated continuously by recombination with the much larger pool of animal-infective trypanosomes. Such novel recombinants present a risk for future outbreaks of HAT.

  14. Genetic recombination between human and animal parasites creates novel strains of human pathogen.

    Science.gov (United States)

    Gibson, Wendy; Peacock, Lori; Ferris, Vanessa; Fischer, Katrin; Livingstone, Jennifer; Thomas, James; Bailey, Mick

    2015-03-01

    Genetic recombination between pathogens derived from humans and livestock has the potential to create novel pathogen strains, highlighted by the influenza pandemic H1N1/09, which was derived from a re-assortment of swine, avian and human influenza A viruses. Here we investigated whether genetic recombination between subspecies of the protozoan parasite, Trypanosoma brucei, from humans and animals can generate new strains of human pathogen, T. b. rhodesiense (Tbr) responsible for sleeping sickness (Human African Trypanosomiasis, HAT) in East Africa. The trait of human infectivity in Tbr is conferred by a single gene, SRA, which is potentially transferable to the animal pathogen Tbb by sexual reproduction. We tracked the inheritance of SRA in crosses of Tbr and Tbb set up by co-transmitting genetically-engineered fluorescent parental trypanosome lines through tsetse flies. SRA was readily transferred into new genetic backgrounds by sexual reproduction between Tbr and Tbb, thus creating new strains of the human pathogen, Tbr. There was no evidence of diminished growth or transmissibility of hybrid trypanosomes carrying SRA. Although expression of SRA is critical to survival of Tbr in the human host, we show that the gene exists as a single copy in a representative collection of Tbr strains. SRA was found on one homologue of chromosome IV in the majority of Tbr isolates examined, but some Ugandan Tbr had SRA on both homologues. The mobility of SRA by genetic recombination readily explains the observed genetic variability of Tbr in East Africa. We conclude that new strains of the human pathogen Tbr are being generated continuously by recombination with the much larger pool of animal-infective trypanosomes. Such novel recombinants present a risk for future outbreaks of HAT.

  15. The value of recombinant human TSH-aided 131I treatment in differentiated thyroid carcinoma patients

    International Nuclear Information System (INIS)

    Ding Yong; Long Yahong; Tian Jiahe; Xu Baixuan; Xing Jialiu; Fang Yi; Wei Lijing; Zong Zhaoyi

    2013-01-01

    Objective: To evaluate the efficacy and safety of recombinant human TSH(rhTSH)-aided 131 I treatment for DTC. Methods: A total of 144 patients with DTC who underwent total or near total thyroidectomy were retrospectively analyzed. The rhTSH-aided 131 I treatment of 3.7 GBq was performed in 72 cases (Group Ⅰ: euthyroid). Another 72 cases received radioiodine ablation treatment of 3.7 GBq after 4 to 6 weeks of thyroxine withdrawal (Group Ⅱ: hypothyroidism). Serum endogenous TSH, FT 3 , FT 4 and Tg were measured. The life qualities of both groups were observed, such as intolerance to cold, weight gain, constipation, motor retardation, skin dryness, periorbital edema and bone pain. Absence of visible uptake or uptake rate less than 1% was taken as complete ablation. The efficacy of 131 I treatment was evaluated. The life quality of both groups was evaluated by χ 2 test, and the effect of 131 I treatment was analyzed by t test. Results: Serum TSH was effectively improved in both groups before 131 I treatment. In group Ⅰ, TSH was higher than that of group Ⅱ ((141.26 ± 27.30) mU/L vs (70.57 ± 51.13) mU/L; t=2.435, P<0.05), and FT 3 , FT 4 were not significantly different before or after the injection of rhTSH. Tg was well stimulated in both groups with no statistical difference. Group Ⅱ exhibited more side effects, which included intolerance to cold 80.56% (58/72), weight gain 86.11% (62/72), constipation 15.28% (11/72), motor retardation 22.22% (16/72), skin dryness 56.94% (41/72), bone pain 2.78% (2/72), and no periorbital edema was found. Group Ⅰ had a higher quality of life than group Ⅱ, only few side effects were observed including dizziness and nausea 2.78% (2/72), bone pain 2.78% (2/72), and transient tachycardia 1.39% (1/72). The effect of 131 I treatment was evaluated by whole body scans with a diagnostic dose of 131 I. The complete ablation rate was 70.83% (51/72) in group Ⅰ and 66.67% (48/72) in group Ⅱ (χ 2 =0.58, P>0.05). Conclusion: The

  16. High versus low radioiodine activity in patients with differentiated thyroid cancer - A meta-analysis

    International Nuclear Information System (INIS)

    Valachis, Antonis; Nearchou, Andreas

    2013-01-01

    Background: The purpose of the meta-analysis was to estimate the effectiveness and toxicity of low activity radioiodine ablation versus high activity in patients with differentiated thyroid cancer (DTC). Design: A systematic review and meta-analysis was performed by including all randomized trials of low activity versus high activity radioiodine ablation after thyroidectomy. Standard meta-analytic procedures were used to analyze the study outcomes. Results: Ten trials were considered eligible and were further analyzed. The pooled risk ratio (RR) of having a successful ablation for an activity of 1100 MBq versus 3700 MBq (seven trials, 1772 patients) was 0.94 (95% CI 0.85 - 1.04, p-value 0.21). The RR for successful ablation when only thyroid hormone withdrawal was used (five trials, 1116 patients) was 0.87 (95% CI 0.72 - 1.06, p-value 0.17) and it was comparable to RR when only recombinant-human TSH (rec-hTSH) (two trials, 812 patients) was used (1.00, 95% CI 0.93 - 1.07, p-value 0.92). Salivary dysfunction, nausea, and neck pain were significantly more frequent among patients with higher dose for ablation. Conclusion: Our meta-analysis provides some evidence from randomized trials that a lower activity of radioiodine ablation is as effective as higher dose after surgery in patients with DTC with lower toxicity

  17. High versus low radioiodine activity in patients with differentiated thyroid cancer - A meta-analysis

    Energy Technology Data Exchange (ETDEWEB)

    Valachis, Antonis [Dept. of Oncology, Maelarsjukhuset., Eskilstuna (Sweden); Univ. of Uppsala,, (Sweden)], e-mail: Valachis@hotmail.com, Antonis.Valachis@akademiska.uu.se; Nearchou, Andreas [Dept, of Oncology, Maelarsjukhuset., Eskilstuna (Sweden); Univ. of Uppsala., Uppsala (Sweden)

    2013-08-15

    Background: The purpose of the meta-analysis was to estimate the effectiveness and toxicity of low activity radioiodine ablation versus high activity in patients with differentiated thyroid cancer (DTC). Design: A systematic review and meta-analysis was performed by including all randomized trials of low activity versus high activity radioiodine ablation after thyroidectomy. Standard meta-analytic procedures were used to analyze the study outcomes. Results: Ten trials were considered eligible and were further analyzed. The pooled risk ratio (RR) of having a successful ablation for an activity of 1100 MBq versus 3700 MBq (seven trials, 1772 patients) was 0.94 (95% CI 0.85 - 1.04, p-value 0.21). The RR for successful ablation when only thyroid hormone withdrawal was used (five trials, 1116 patients) was 0.87 (95% CI 0.72 - 1.06, p-value 0.17) and it was comparable to RR when only recombinant-human TSH (rec-hTSH) (two trials, 812 patients) was used (1.00, 95% CI 0.93 - 1.07, p-value 0.92). Salivary dysfunction, nausea, and neck pain were significantly more frequent among patients with higher dose for ablation. Conclusion: Our meta-analysis provides some evidence from randomized trials that a lower activity of radioiodine ablation is as effective as higher dose after surgery in patients with DTC with lower toxicity.

  18. Biological evaluation of recombinant human erythropoietin in pharmaceutical products

    Directory of Open Access Journals (Sweden)

    Ramos A.S.

    2003-01-01

    Full Text Available The potencies of mammalian cell-derived recombinant human erythropoietin pharmaceutical preparations, from a total of five manufacturers, were assessed by in vivo bioassay using standardized protocols. Eight-week-old normocythemic mice received a single subcutaneous injection followed by blood sampling 96 h later or multiple daily injections with blood sampling 24 h after the last injection. Reticulocyte counting by microscopic examination was employed as the end-point using the brilliant cresyl blue or selective hemolysis methods, together with automated flow cytometry. Different injection schedules were investigated and dose-response curves for the European Pharmacopoeia Biological Reference Preparation of erythropoietin were compared. Manual and automated methods of reticulocyte counting were correlated with respect to assay validity and precision. Using 8 mice per treatment group, intra-assay precision determined for all of the assays in the study showed coefficients of variation of 12.1-28.4% for the brilliant cresyl blue method, 14.1-30.8% for the selective hemolysis method and 8.5-19.7% for the flow cytometry method. Applying the single injection protocol, a combination of at least two independent assays was required to achieve the precision potency and confidence limits indicated by the manufacturers, while the multiple daily injection protocol yielded the same acceptable results within a single assay. Although the latter protocol using flow cytometry for reticulocyte counting gave more precise and reproducible results (intra-assay coefficients of variation: 5.9-14.2%, the well-characterized manual methods provide equally valid alternatives for the quality control of recombinant human erythropoietin therapeutic products.

  19. Different behaviour of radioiodinated human recombinant interleukin-1 and its receptor antagonist in an animal model of infection

    International Nuclear Information System (INIS)

    Laken, C.J. van der; Boerman, O.C.; Oyen, W.J.G.; Ven, M.T.P. van den; Claessens, R.A.M.J.; Meer, J.W.M. van der; Corstens, F.H.M.

    1996-01-01

    Recently, we demonstrated that radiolabelled interleukin-1α (IL-1) specifically accumulates in focal infection in mice through interaction with its receptor. Unfortunately, systemic side-effects of IL-1 limit its clinical application. We investigated whether this problem could be circumvented by using the interleukin-1 receptor antagonist (IL-1ra), an equally sized protein that binds to the same receptors as IL-1 without induction of biological effects. Biodistribution of 125 I-IL-1 and 125 I-IL-1ra was determined in Swiss mice with Staphylococcus aureus-induced abscesses in the left calf muscle at 4, 12, 24 and 48 h after injection of either 0.4 MBq 125 I-IL-1 or 0.4 MBq 125 I-IL-1ra. In vitro, the proteins displayed similar binding characteristics. High-performance liquid chromatographic analysis revealed a tendency for IL-1ra to associate with serum proteins. Both proteins rapidly cleared from most organs. However, the abscess uptake of 125 I-IL-1ra was significantly lower than that of 125 I-IL-1 at all time points (48 h p.i.: 0.06±0.01%ID/g vs 0.60±0.04%ID/g; P 125 I-IL-1ra, while the ratios for 125 I-IL-1 reached 46.9±5.7 at 48 h p.i. Despite similar in vitro receptor binding, the abscess uptake of IL-1ra was much lower than that of IL-1. The interaction of IL-1ra with serum proteins in vivo may reduce its availability for receptor binding in the infection. Although on theoretical grounds IL-1ra is very interesting, these characteristics will prevent its development as a clinically useful radiopharmaceutical to image infection. (orig.). With 4 figs., 2 tabs

  20. Short-term effects of recombinant human growth hormone and feeding on gluconeogenesis in humans

    Science.gov (United States)

    After a short-term fast, lactating women have increased rates of glucose production but not gluconeogenesis (GNG) despite relative hypoinsulinemia. We explored the effects of non-insulin-dependent increase in glucose utilization and recombinant human growth hormone (rhGH) on glucose production, glyc...

  1. High-Resolution Patterns of Meiotic Recombination across the Human Major Histocompatibility Complex

    Science.gov (United States)

    Cullen, Michael; Perfetto, Stephen P.; Klitz, William; Nelson, George; Carrington, Mary

    2002-01-01

    Definitive characteristics of meiotic recombination events over large (i.e., >1 Mb) segments of the human genome remain obscure, yet they are essential for establishing the haplotypic structure of the genome and for efficient mapping of complex traits. We present a high-resolution map of recombination at the kilobase level across a 3.3-Mb interval encompassing the major histocompatibility complex (MHC). Genotyping of 20,031 single sperm from 12 individuals resulted in the identification and fine mapping of 325 recombinant chromosomes within genomic intervals as small as 7 kb. Several principal characteristics of recombination in this region were observed: (1) rates of recombination can differ significantly between individuals; (2) intense hot spots of recombination occur at least every 0.8 Mb but are not necessarily evenly spaced; (3) distribution in the location of recombination events can differ significantly among individuals; (4) between hot spots, low levels of recombination occur fairly evenly across 100-kb segments, suggesting the presence of warm spots of recombination; and (5) specific sequence motifs associate significantly with recombination distribution. These data provide a plausible model for recombination patterns of the human genome overall. PMID:12297984

  2. A survey of owners' perceptions and experiences of radioiodine treatment of feline hyperthyroidism in the UK.

    Science.gov (United States)

    Boland, Lara A; Murray, Jane K; Bovens, Catherine Pv; Hibbert, Angie

    2014-08-01

    The efficacy of radioiodine treatment of feline hyperthyroidism is well established; however, limited information is known about owners' perceptions or experiences of radioiodine. This study aimed to examine factors that influence owner treatment choices and their opinions following radioiodine. Surveys were sent to owners of cats referred for radioiodine treatment between 2002 and 2011 (radioiodine group; 264 cats) and owners of non-radioiodine-treated hyperthyroid cats seen at first-opinion practices (control group; 199 cats). The response rate was 67.0% (310 returned: 175 radioiodine, 135 control). Of 135 controls, 72 (53.3%) were unaware of radioiodine as a treatment option. Owners of cats ⩾15 years old and uninsured cats were less likely to pursue radioiodine. Cost of treatment, travel distance, potential human or animal health risks and waiting periods for radioiodine had a low impact on owners' treatment choice. Owners reported a moderate level of concern about treatment hospitalisation length, which included (158 respondents) the possibility of the cat being unhappy 130 (82.3%), owner missing the cat 102 (64.6%), inappetence 50 (31.6%), other pets missing the cat 32 (20.3%), development of co-morbid disease 28 (17.7%) and side effects 25 (15.8%). Owners assessed their cat's quality of life on a scale of 1 (very poor) to 10 (excellent), as 4 (4) (median [interquartile range]) pre-radioiodine (134 respondents) and 9 (2) post-radioiodine (131 respondents). Of 132 respondents, 121 (91.7%) were happy with their decision to choose radioiodine. The results of this questionnaire may assist veterinarians in addressing common owner concerns when discussing radioiodine as a treatment option for hyperthyroidism. © ISFM and AAFP 2014.

  3. Genetic analysis of variation in human meiotic recombination.

    Directory of Open Access Journals (Sweden)

    Reshmi Chowdhury

    2009-09-01

    Full Text Available The number of recombination events per meiosis varies extensively among individuals. This recombination phenotype differs between female and male, and also among individuals of each gender. In this study, we used high-density SNP genotypes of over 2,300 individuals and their offspring in two datasets to characterize recombination landscape and to map the genetic variants that contribute to variation in recombination phenotypes. We found six genetic loci that are associated with recombination phenotypes. Two of these (RNF212 and an inversion on chromosome 17q21.31 were previously reported in the Icelandic population, and this is the first replication in any other population. Of the four newly identified loci (KIAA1462, PDZK1, UGCG, NUB1, results from expression studies provide support for their roles in meiosis. Each of the variants that we identified explains only a small fraction of the individual variation in recombination. Notably, we found different sequence variants associated with female and male recombination phenotypes, suggesting that they are regulated by different genes. Characterization of genetic variants that influence natural variation in meiotic recombination will lead to a better understanding of normal meiotic events as well as of non-disjunction, the primary cause of pregnancy loss.

  4. Recombinant human endostatin improves tumor vasculature and alleviates hypoxia in Lewis lung carcinoma

    International Nuclear Information System (INIS)

    Peng Fang; Wang Jin; Zou Yi; Bao Yong; Huang Wenlin; Chen Guangming; Luo Xianrong; Chen Ming

    2011-01-01

    Objective: To investigate whether recombinant human endostatin can create a time window of vascular normalization prior to vascular pruning to alleviate hypoxia in Lewis lung carcinoma in mice. Methods: Kinetic changes in morphology of tumor vasculature in response to recombinant human endostatin were detected under a confocal microscope with immunofluorescent staining in Lewis lung carcinomas in mice. The hypoxic cell fraction of different time was assessed with immunohistochemical staining . Effects on tumor growth were monitored as indicated in the growth curve of tumors . Results: Compared with the control group vascularity of the tumors was reduced over time by recombinant human endostatin treatment and significantly regressed for 9 days. During the treatment, pericyte coverage increased at day 3, increased markedly at day 5, and fell again at day 7. The vascular basement membrane was thin and closely associated with endothelial cells after recombinant human endostatin treatment, but appeared thickened, loosely associated with endothelial cells in control tumors. The decrease in hypoxic cell fraction at day 5 after treatment was also found. Tumor growth was not accelerated 5 days after recombinant human endostatin treatment. Conclusions: Recombinant human endostatin can normalize tumor vasculature within day 3 to 7, leading to improved tumor oxygenation. The results provide important experimental basis for combining recombinant human endostatin with radiation therapy in human tumors. (authors)

  5. Recombinant methods for screening human DNA excision repair proficiency

    International Nuclear Information System (INIS)

    Athas, W.F.

    1988-01-01

    A method for measuring DNA excision repair in response to ultraviolet radiation (UV)-induced DNA damage has been developed, validated, and field-tested in cultured human lymphocytes. The methodology is amenable to population-based screening and should facilitate future epidemiologic studies seeking to investigate associations between excision repair proficiency and cancer susceptibility. The impetus for such endeavors derives from the belief that the high incidence of skin cancer in the genetic disorder xeroderma pigmentosum (XP) primarily is a result of the reduced capacity of patients cells to repair UV-induced DNA damage. For assay, UV-irradiated non-replicating recombinant plasmid DNA harboring a chloramphenicol acetyltransferase (CAT) indicator gene is introduced into lymphocytes using DEAE-dextran short-term transfection conditions. Exposure to UV induces transcriptionally-inactivating DNA photoproducts in the plasmid DNA which inactivate CAT gene expression. Excision repair of the damaged CAT gene is monitored indirectly as a function of reactivated CAT enzyme activity following a 40 hour repair/expression incubation period

  6. Recombinant human erythropoietin therapy in critically ill Jehovah's Witnesses.

    Science.gov (United States)

    Ball, Amanda M; Winstead, P Shane

    2008-11-01

    Blood transfusions and blood products are often given as a life-saving measure in patients with critical illness. However, some patients, such as Jehovah's Witnesses, may refuse their administration due to religious beliefs. Jehovah's Witnesses accept most available medical treatments, but not blood transfusions or blood products due to their religion's interpretation of several passages from the Bible. Since recombinant human erythropoietin (rHuEPO) became available, several cases have been reported in which rHuEPO was successfully administered to critically ill Jehovah's Witnesses. Administration of rHuEPO in combination with other blood conservation techniques has been shown to increase hemoglobin levels and survival in patients who experienced trauma, burns, general surgery, or gastrointestinal hemorrhage. We performed a literature search of the MEDLINE and International Pharmaceutical Abstracts databases of rHuEPO therapy in the Jehovah's Witness population. Fourteen cases were identified in which rHuEPO was administered to Jehovah's Witnesses who required the drug for critical care resuscitation as an alternative to blood products. In each clinical situation, rHuEPO enhanced erythropoiesis; however, time to the start of treatment, dosages, route of administration, and treatment duration varied widely. Supplementation with adjunctive agents, such as iron, folic acid, and vitamin B12, was also beneficial. Use of rHuEPO in Jehovah's Witnesses may provide an alternative to blood transfusions or blood products. Other alternatives, such as hemoglobin-based oxygen carriers and perfluorocarbons, are also being explored.

  7. Synthesis of radioiodinated labeled peptides

    International Nuclear Information System (INIS)

    Matloobi, M.; Rafii, H.; Beigi, D.; Khalaj, A.; Kamali-Dehghan, M.

    2003-01-01

    Optimization of radioiodination of peptides is covered by both a direct method in which a constituent tyrosine residue is labeled and indirect method by using an iodinated derivative (SIB) of N succinimidyl 3-(tri-n-butylstannyl) benzoate (ATE) as the intermediate. Radioiodination of IgG and FMLF were performed by direct method using Chloramine-T as an oxidant but since Formyl-Methyl-Leucyl-Phenylalanine, FMLF, does not lend itself for direct radioiodination we performed labeling of FMLF by indirect method via radioiodined SIB at different pH. (author)

  8. Comparative pharmacology of a new recombinant FSH expressed by a human cell line

    DEFF Research Database (Denmark)

    Koechling, Wolfgang; Plaksin, Daniel; Croston, Glenn E.

    2017-01-01

    Recombinant FSH proteins are important therapeutic agents for the treatment of infertility, including follitropin alfa expressed in Chinese Hamster Ovary (CHO) cells and, more recently, follitropin delta expressed in the human cell line PER.C6. These recombinant FSH proteins have distinct glycosy...

  9. Obtention of a prosthetic group for labelling of radioiodinated proteins

    International Nuclear Information System (INIS)

    Santos, Josefina da S.; Colturato, Maria Tereza; Araujo, Elaine B. de

    2000-01-01

    Antibodies and peptides labeled with radionuclides has been extensively used in radioimmunotherapy and radioimmunodetection. The principal problem with the use of radioiodinated proteins is the in vivo dehalogenation. The use of prosthetic groups for indirect labeling of proteins with radioiodine has showed to be useful on labeling proteins with greater in vivo stability. A procedure is described for the preparation of an radioiodinated prosthetic group (N-succinimidyl 4-radioiodine-benzoate-SIB), using procedure described by Stocklin et al, with the iodination of p-bromo-benzoic acid and subsequent reaction with TSTU. Preliminary labeling results showed that the prosthetic group can be obtained in a good yield. The coupling of the SIB to the protein will be studied using human IgG as protein model. (author)

  10. (Radioiodinated free fatty acids)

    Energy Technology Data Exchange (ETDEWEB)

    Knapp, Jr., F. F.

    1987-12-11

    The traveler participated in the Second International Workshop on Radioiodinated Free Fatty Acids in Amsterdam, The Netherlands where he presented an invited paper describing the pioneering work at the Oak Ridge National Laboratory (ORNL) involving the design, development and testing of new radioiodinated methyl-branched fatty acids for evaluation of heart disease. He also chaired a technical session on the testing of new agents in various in vitro and in vivo systems. He also visited the Institute for Clinical and Experimental Nuclear Medicine in Bonn, West Germany, to review, discuss, plan and coordinate collaborative investigations with that institution. In addition, he visited the Cyclotron Research Center in Liege, Belgium, to discuss continuing collaborative studies with the Osmium-191/Iridium-191m radionuclide generator system, and to complete manuscripts and plan future studies.

  11. Ligand and proton exchange dynamics in recombinant human myoglobin mutants.

    Science.gov (United States)

    Lambright, D G; Balasubramanian, S; Boxer, S G

    1989-05-05

    Site-specific mutants of human myoglobin have been prepared in which lysine 45 is replaced by arginine (K45R) and aspartate 60 by glutamate (D60E), in order to examine the influence of these residues and their interaction on the dynamics of the protein. These proteins were studied by a variety of methods, including one and two-dimensional proton nuclear magnetic resonance spectroscopy, exchange kinetics for the distal and proximal histidine NH protons as a function of pH in the met cyano forms, flash photolysis of the CO forms, and ligand replacement kinetics. The electronic absorption and proton nuclear magnetic resonance spectra of the CO forms of these proteins are virtually identical, indicating that the structure of the heme pocket is unaltered by these mutations. There are, however, substantial changes in the dynamics of both CO binding and proton exchange for the mutant K45R, whereas the mutant D60E exhibits behavior indistinguishable from the reference human myoglobin. K45R has a faster CO bimolecular recombination rate and slower CO off-rate relative to the reference. The kinetics for CO binding are independent of pH (6.5 to 10) as well as ionic strength (0 to 1 M-NaCl). The exchange rate for the distal histidine NH is substantially lower for K45R than the reference, whereas the proximal histidine NH exchange rate is unaltered. The exchange behavior of the human proteins is similar to that reported for a comparison of the exchange rates for myoglobins having lysine at position 45 with sperm whale myoglobin, which has arginine at this position. This indicates that the differences in exchange rates reflects largely the Lys----Arg substitution. The lack of a simple correlation for the CO kinetics with this substitution means that these are sensitive to other factors as well. Specific kinetic models, whereby substitution of arginine for lysine at position 45 can affect ligand binding dynamics, are outlined. These experiments demonstrate that a relatively

  12. Boundary mode lubrication of articular cartilage by recombinant human lubricin.

    Science.gov (United States)

    Gleghorn, Jason P; Jones, Aled R C; Flannery, Carl R; Bonassar, Lawrence J

    2009-06-01

    Lubrication of cartilage involves a variety of physical and chemical factors, including lubricin, a synovial glycoprotein that has been shown to be a boundary lubricant. It is unclear how lubricin boundary lubricates a wide range of bearings from tissue to artificial surfaces, and if the mechanism is the same for both soluble and bound lubricin. In the current study, experiments were conducted to investigate the hypothesis that recombinant human lubricin (rh-lubricin) lubricates cartilage in a dose-dependent manner and that soluble and bound fractions of rh-lubricin both contribute to the lubrication process. An rh-lubricin dose response was observed with maximal lubrication achieved at concentrations of rh-lubricin greater than 50 microg/mL. A concentration-response variable-slope model was fit to the data, and indicated that rh-lubricin binding to cartilage was not first order. The pattern of decrease in equilibrium friction coefficient indicated that aggregation of rh-lubricin or steric arrangement may regulate boundary lubrication. rh-lubricin localized at the cartilage surface was found to lubricate a cartilage-glass interface in boundary mode, as did soluble rh-lubricin at high concentrations (150 microg/mL); however, the most effective lubrication occurred when both soluble and bound rh-lubricin were present at the interface. These findings point to two distinct mechanisms by which rh-lubricin lubricates, one mechanism involving lubricin bound to the tissue surface and the other involving lubricin in solution. Copyright 2008 Orthopaedic Research Society

  13. Recombinant human hyaluronidase-enabled subcutaneous pediatric rehydration.

    Science.gov (United States)

    Allen, Coburn H; Etzwiler, Lisa S; Miller, Melissa K; Maher, George; Mace, Sharon; Hostetler, Mark A; Smith, Sharon R; Reinhardt, Neil; Hahn, Barry; Harb, George

    2009-11-01

    The Increased Flow Utilizing Subcutaneously-Enabled (INFUSE)-Pediatric Rehydration Study was designed to assess efficacy, safety, and clinical utility of recombinant human hyaluronidase (rHuPH20)-facilitated subcutaneous rehydration in children 2 months to 10 years of age. Patients with mild/moderate dehydration requiring parenteral treatment in US emergency departments were eligible for this phase IV, multicenter, single-arm study. They received subcutaneous injection of 1 mL rHuPH20 (150 U), followed by subcutaneous infusion of 20 mL/kg isotonic fluid over the first hour. Subcutaneous rehydration was continued as needed for up to 72 hours. Rehydration was deemed successful if it was attributed by the investigator primarily to subcutaneous fluid infusion and the child was discharged without requiring an alternative method of rehydration. Efficacy was evaluated in 51 patients (mean age: 1.9 years; mean weight: 11.2 kg). Initial subcutaneous catheter placement was achieved with 1 attempt for 46/51 (90.2%) of patients. Rehydration was successful for 43/51 (84.3%) of patients. Five patients (9.8%) were hospitalized but deemed to be rehydrated primarily through subcutaneous therapy, for a total of 48/51 (94.1%) of patients. No treatment-related systemic adverse events were reported, but 1 serious adverse event occurred (cellulitis at infusion site). Investigators found the procedure easy to perform for 96% of patients (49/51 patients), and 90% of parents (43/48 parents) were satisfied or very satisfied. rHuPH20-facilitated subcutaneous hydration seems to be safe and effective for young children with mild/moderate dehydration. Subcutaneous access is achieved easily, and the procedure is well accepted by clinicians and parents.

  14. Therapy with recombinant human erythropoietin in patients with myelodysplastic syndromes.

    Science.gov (United States)

    Stone, R M; Bernstein, S H; Demetri, G; Facklam, D P; Arthur, K; Andersen, J; Aster, J C; Kufe, D

    1994-10-01

    We conducted a Phase I-II trial of recombinant human erythropoietin-beta (rhEPO) in patients with myelodysplastic syndrome (MDS). Patients with anemia and pathologically confirmed MDS were eligible for the study. Treatment consisted of rhEPO by subcutaneous injection thrice weekly for 6 weeks at one of three dose levels (100 U/kg (three patients), 200 U/kg (three patients) and 400 U/kg (14 patients)). Ferrous sulfate (325 mg po tid) was also administered if the transferrin saturation was below 30% (two patients). Patients were monitored with weekly CBC, white cell differential, and reticulocyte counts. Bone marrow examinations were performed at the conclusion of the treatment period and after a 2 week washout period. Patients who responded to therapy were continued on rhEPO at the same dose for 6 additional months. Response criteria included: 50% reduction in transfusion requirements compared with the 6 week pre-study period; doubling of reticulocyte count that was maintained on two determinations at least 1 week apart; or an increase in hemoglobin by at least 1.2 gm/dl without transfusions. Pre-treatment factors potentially predictive of response were analyzed by univariate analysis and in a multivariate fashion by classification and regression trees. Seven of the twenty patients sustained an untransfused rise in serum hemoglobin > or = 1.2 gm/dl. Four of the sixteen patients (including three of seven patients experiencing a rise in serum hemoglobin) who were transfusion-dependent prior to the study achieved a reduction or elimination of their transfusion requirements. Five of thirteen patients who received rhEPO during the extension phase had a continued response. A low baseline erythropoietin level (< 50 mU/ml) was the best predictor of hemoglobin response when controlling for other variables. rhEPO has a role in the treatment of certain patients with MDS, particularly in those whose endogenous serum erythropoietin levels are not markedly elevated.

  15. Indirect labeling of proteins with radioiodine

    International Nuclear Information System (INIS)

    Araujo, Elaine Bortoleti de; Lavinas, Tatiana; Muramoto, Emiko; Pereira, Nilda P.S. de; Silva, Constancia P.G.; Tavares, Leoberto C.

    2000-01-01

    A procedure is described for the radioiodination of proteins using an iodinated derivative of N succinimidyl 3-(tri-n-butylstannyl)benzoate (ATE), previously described by Zalutsky. ATE was obtained in a high pure form and the iodination has been performed with 131-Iodine in 70-80% yield. Protein labeling studies performed with human IgG indicate that the ATE intermediate is an important alternative to conventional labeling methods. (author)

  16. Genetic recombination as a major cause of mutagenesis in the human globin gene clusters.

    Science.gov (United States)

    Borg, Joseph; Georgitsi, Marianthi; Aleporou-Marinou, Vassiliki; Kollia, Panagoula; Patrinos, George P

    2009-12-01

    Homologous recombination is a frequent phenomenon in multigene families and as such it occurs several times in both the alpha- and beta-like globin gene families. In numerous occasions, genetic recombination has been previously implicated as a major mechanism that drives mutagenesis in the human globin gene clusters, either in the form of unequal crossover or gene conversion. Unequal crossover results in the increase or decrease of the human globin gene copies, accompanied in the majority of cases with minor phenotypic consequences, while gene conversion contributes either to maintaining sequence homogeneity or generating sequence diversity. The role of genetic recombination, particularly gene conversion in the evolution of the human globin gene families has been discussed elsewhere. Here, we summarize our current knowledge and review existing experimental evidence outlining the role of genetic recombination in the mutagenic process in the human globin gene families.

  17. Recombinant human DNase in children with airway malacia and lower respiratory tract infection.

    NARCIS (Netherlands)

    Boogaard, R.; Jongste, J.C. de; Vaessen-Verberne, A.A.; Hop, W.C.J.; Merkus, P.J.F.M.

    2009-01-01

    BACKGROUND: Children with airway malacia often have protracted courses of airway infections, because dynamic airway collapse during coughing results in impaired mucociliary clearance. The aim of this study was to determine the effect of the mucolytic drug recombinant human deoxyribonuclease

  18. Pharmacoeconomic review of recombinant human DNase in the management of cystic fibrosis

    NARCIS (Netherlands)

    Zijlstra, Gerrit; Boersma, Cornelis; Frijlink, Henderik W.; Postma, Maarten J.

    For the treatment of patients with cystic fibrosis, recombinant human deoxyribonuclease I is widely used. Deoxyribonuclease I has a positive effect on lung function and the number of hospitalizations. Deoxyribonuclease I is currently administered by nebulization, which is an inefficient

  19. Improved Refolding Efficacy of Recombinant Human Interferon α-2b ...

    African Journals Online (AJOL)

    Different refolding buffers were employed for refolding the target protein. The refolded ... secondary structure of the protein was altered, probably due to increase in alpha-helix from 23.7 % at. pH 7.0 to 28.1 % ... One of the recombinant proteins ...

  20. Radioiodinated tracers for myocardial imaging

    International Nuclear Information System (INIS)

    Kulkarni, P.V.; Corbett, J.R.

    1990-01-01

    Recent advances in the efficient production of high purity radioiodine (123I) and new efficient radiolabeling techniques have allowed the development of new classes of cardiovascular radiopharmaceuticals. These include 123I-labeled fatty acids to assess myocardial metabolism, 123I-metaiodobenzylguanidine (MIBG) for myocardial neuronal activity, labeled monoclonal antibodies for myocardial necrosis, and labeled lipoproteins for receptor concentration. 123I-labeled fatty acids and MIBG are under clinical investigation with encouraging results. 123I- and 111In-labeled fragments of monoclonal antibodies to myosin have been used for imaging myocardial necrosis in humans. The development of radiotracers for imaging of cholinergic and adrenergic receptors is still in the experimental stage. Recent advances in imaging instrumentation and radiopharmaceuticals have resulted in cardiac imaging applications beyond blood pool ventriculography, perfusion, and infarct-avid imaging. Developments of radioiodine (123I)-labeled agents promise to play an important role in the assessment of myocardial metabolism, neuronal activity, and receptor concentration. The chemistry of iodine is well defined compared with that of 99mTc; therefore, iodine isotopes are well suited for labeling biologically important molecules. Among the iodine isotopes, 123I has nearly ideal nuclear properties for nuclear medical applications with a 13.3-hour half-life (T1/2) and 159 keV gamma emission (83%). Despite the nearly ideal chemical and nuclear properties of 123I, the widespread application of 123I-based radiopharmaceuticals in clinical practice has been limited by high production costs (123I is produced in a cyclotron), relatively limited availability, and the presence of undesirable radionuclidic impurities (124I, T1/2 = 4.2 days; 125I, T1/2 = 60 days; 126I, T1/2 = 13.1 days). 77 references

  1. Short-term effect of recombinant human growth hormone in patients with alcoholic cirrhosis

    DEFF Research Database (Denmark)

    Møller, S; Becker, U; Grønbaek, M

    1994-01-01

    As growth hormone possesses anabolic properties that are active on protein metabolism, and thus of potential benefit to patients with chronic liver disease, we determined the metabolic effects of recombinant human growth hormone on insulin-like growth factor-I (IGF-I) its specific binding proteins......, and liver function. Twenty consecutive patients with cirrhosis were randomized to recombinant human growth hormone (Norditropin, 4 I.U. twice daily) subcutaneously for 6 weeks (n = 10) or conventional medical treatment (n = 10). The serum concentrations of insulin-like growth factor-I in the recombinant...... patients as well as in controls, whereas no change in insulin-like growth factor binding protein-1 concentrations was found. No significant changes were seen in the area under the curve for biochemical liver function tests. We conclude that administration of recombinant human growth hormone induces...

  2. Structure of recombinant human carboxylesterase 1 isolated from whole cabbage looper larvae

    International Nuclear Information System (INIS)

    Greenblatt, Harry M.; Otto, Tamara C.; Kirkpatrick, Melanie G.; Kovaleva, Elena; Brown, Susan; Buchman, George; Cerasoli, Douglas M.; Sussman, Joel L.

    2012-01-01

    Large quantities of recombinant human carboxylesterase 1 have been produced in an economical whole insect larvae system. The crystal structure of this enzyme is essentially identical to that produced by cell culture techniques. The use of whole insect larvae as a source of recombinant proteins offers a more cost-effective method of producing large quantities of human proteins than conventional cell-culture approaches. Human carboxylesterase 1 has been produced in and isolated from whole Trichoplusia ni larvae. The recombinant protein was crystallized and its structure was solved to 2.2 Å resolution. The results indicate that the larvae-produced enzyme is essentially identical to that isolated from cultured Sf21 cells, supporting the use of this expression system to produce recombinant enzymes for crystallization studies

  3. Genetic recombination pathways and their application for genome modification of human embryonic stem cells.

    Science.gov (United States)

    Nieminen, Mikko; Tuuri, Timo; Savilahti, Harri

    2010-10-01

    Human embryonic stem cells are pluripotent cells derived from early human embryo and retain a potential to differentiate into all adult cell types. They provide vast opportunities in cell replacement therapies and are expected to become significant tools in drug discovery as well as in the studies of cellular and developmental functions of human genes. The progress in applying different types of DNA recombination reactions for genome modification in a variety of eukaryotic cell types has provided means to utilize recombination-based strategies also in human embryonic stem cells. Homologous recombination-based methods, particularly those utilizing extended homologous regions and those employing zinc finger nucleases to boost genomic integration, have shown their usefulness in efficient genome modification. Site-specific recombination systems are potent genome modifiers, and they can be used to integrate DNA into loci that contain an appropriate recombination signal sequence, either naturally occurring or suitably pre-engineered. Non-homologous recombination can be used to generate random integrations in genomes relatively effortlessly, albeit with a moderate efficiency and precision. DNA transposition-based strategies offer substantially more efficient random strategies and provide means to generate single-copy insertions, thus potentiating the generation of genome-wide insertion libraries applicable in genetic screens. 2010 Elsevier Inc. All rights reserved.

  4. Developing baculovirus-insect cell expression systems for humanized recombinant glycoprotein production

    International Nuclear Information System (INIS)

    Jarvis, Donald L.

    2003-01-01

    The baculovirus-insect cell expression system is widely used to produce recombinant glycoproteins for many different biomedical applications. However, due to the fundamental nature of insect glycoprotein processing pathways, this system is typically unable to produce recombinant mammalian glycoproteins with authentic oligosaccharide side chains. This minireview summarizes our current understanding of insect protein glycosylation pathways and our recent efforts to address this problem. These efforts have yielded new insect cell lines and baculoviral vectors that can produce recombinant glycoproteins with humanized oligosaccharide side chains

  5. Metabolism of ethylbenzene by human liver microsomes and recombinant human cytochrome P450s (CYP).

    Science.gov (United States)

    Sams, Craig; Loizou, George D; Cocker, John; Lennard, Martin S

    2004-03-07

    The enzyme kinetics of the initial hydroxylation of ethylbenzene to form 1-phenylethanol were determined in human liver microsomes. The individual cytochrome P450 (CYP) forms catalysing this reaction were identified using selective inhibitors and recombinant preparations of hepatic CYPs. Production of 1-phenylethanol in hepatic microsomes exhibited biphasic kinetics with a high affinity, low Km, component (mean Km = 8 microM; V(max) = 689 pmol/min/mg protein; n = 6 livers) and a low affinity, high Km, component (Km = 391 microM; V(max) = 3039 pmol/min/mg protein; n = 6). The high-affinity component was inhibited 79%-95% (mean 86%) by diethyldithiocarbamate, and recombinant CYP2E1 was shown to metabolise ethylbenzene with low Km (35 microM), but also low (max) (7 pmol/min/pmol P450), indicating that this isoform catalysed the high-affinity component. Recombinant CYP1A2 and CYP2B6 exhibited high V(max) (88 and 71 pmol/min/pmol P450, respectively) and high Km (502 and 219 microM, respectively), suggesting their involvement in catalysing the low-affinity component. This study has demonstrated that CYP2E1 is the major enzyme responsible for high-affinity side chain hydroxylation of ethylbenzene in human liver microsomes. Activity of this enzyme in the population is highly variable due to induction or inhibition by physiological factors, chemicals in the diet or some pharmaceuticals. This variability can be incorporated into the risk assessment process to improve the setting of occupational exposure limits and guidance values for biological monitoring.

  6. The remarkable frequency of human immunodeficiency virus type 1 genetic recombination.

    Science.gov (United States)

    Onafuwa-Nuga, Adewunmi; Telesnitsky, Alice

    2009-09-01

    The genetic diversity of human immunodeficiency virus type 1 (HIV-1) results from a combination of point mutations and genetic recombination, and rates of both processes are unusually high. This review focuses on the mechanisms and outcomes of HIV-1 genetic recombination and on the parameters that make recombination so remarkably frequent. Experimental work has demonstrated that the process that leads to recombination--a copy choice mechanism involving the migration of reverse transcriptase between viral RNA templates--occurs several times on average during every round of HIV-1 DNA synthesis. Key biological factors that lead to high recombination rates for all retroviruses are the recombination-prone nature of their reverse transcription machinery and their pseudodiploid RNA genomes. However, HIV-1 genes recombine even more frequently than do those of many other retroviruses. This reflects the way in which HIV-1 selects genomic RNAs for coencapsidation as well as cell-to-cell transmission properties that lead to unusually frequent associations between distinct viral genotypes. HIV-1 faces strong and changeable selective conditions during replication within patients. The mode of HIV-1 persistence as integrated proviruses and strong selection for defective proviruses in vivo provide conditions for archiving alleles, which can be resuscitated years after initial provirus establishment. Recombination can facilitate drug resistance and may allow superinfecting HIV-1 strains to evade preexisting immune responses, thus adding to challenges in vaccine development. These properties converge to provide HIV-1 with the means, motive, and opportunity to recombine its genetic material at an unprecedented high rate and to allow genetic recombination to serve as one of the highest barriers to HIV-1 eradication.

  7. Expression of active recombinant human alpha 1-antitrypsin in transgenic rabbits

    NARCIS (Netherlands)

    Massoud, M.; Bischoff, Rainer; Dalemans, W.; Pointu, H.; Attal, J.; Schultz, H.; Clesse, D.; Stinnakre, M.G.; Pavirani, A.; Houdebine, L.M.

    1991-01-01

    A DNA construct containing the human alpha 1-antitrypsin gene including 1.5 and 4 kb of 5' and 3' flanking sequences, was microinjected into the pronucleus of rabbit embryos. The recombinant human protein was (a) expressed in the blood circulation of F0 and F1 transgenic rabbits at an average

  8. Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells.

    Science.gov (United States)

    Gerashchenko, O L; Zhuravel, E V; Skachkova, O V; Khranovska, N N; Filonenko, V V; Pogrebnoy, P V; Soldatkina, M A

    2013-06-01

    The aim of the study was in vitro analysis of biological activity of recombinant human beta-defensin-4 (rec-hBD-4). hBD-4 cDNA was cloned into pGEX-2T vector, and recombinant plasmid was transformed into E. coli BL21(DE3) cells. To purify soluble fusion GST-hBD-4 protein, affinity chromatography was applied. Rec-hBD-4 was cleaved from the fusion protein with thrombin, and purified by reverse phase chromatography on Sep-Pack C18. Effects of rec-hBD-4 on proliferation, viability, cell cycle distribution, substrate-independent growth, and mobility of cultured human cancer cells of A431, A549, and TPC-1 lines were analyzed by direct cell counting technique, MTT assay, flow cytofluorometry, colony forming assay in semi-soft medium, and wound healing assay. Rec-hBD-4 was expressed in bacterial cells as GST-hBD-4 fusion protein, and purified by routine 3-step procedure (affine chromatography on glutathione-agarose, cleavage of fusion protein by thrombin, and reverse phase chromatography). Analysis of in vitro activity of rec-hBD-4 toward three human cancer cell lines has demonstrated that the defensin is capable to affect cell behaviour in concentration-dependent manner. In 1-100 nM concentrations rec-hBD-4 significantly stimulates cancer cell proliferation and viability, and promotes cell cycle progression through G2/M checkpoint, greatly enhances colony-forming activity and mobility of the cells. Treatment of the cells with 500 nM of rec-hBD-4 resulted in opposite effects: significant suppression of cell proliferation and viability, blockage of cell cycle in G1/S checkpoint, significant inhibition of cell migration and colony forming activity. Recombinant human beta-defensin-4 is biologically active peptide capable to cause oppositely directed effects toward biologic features of cancer cells in vitro dependent on its concentration.

  9. Tratamiento con eritropoyetina humana recombinante Human recombinant erythropoietin therapy

    Directory of Open Access Journals (Sweden)

    Hugo Donato

    2006-02-01

    Full Text Available La eritropoyetina recombinante (rHuEPO se ha transformado en la citoquina más utilizada terapéuticamente en el mundo. Luego del éxito obtenido en pacientes con insuficiencia renal terminal, se pudo establecer la utilidad de la terapia con rHuEPO para mejorar otras anemias, incluso en pacientes pediátricos y neonatos. El tratamiento o la prevención de la anemia del prematuro mediante el uso de rHuEPO llevó a una significativa reducción en cantidad de transfusiones y en exposición a dadores. Aún debe establecerse una clara definición sobre cuáles niños prematuros deben recibir tratamiento rutinariamente. Otras indicaciones en período neonatal incluyen anemias hiporregenerativas y hemolíticas. La eficacia de la rHuEPO en niños mayores, con excepción de la insuficiencia renal crónica, no ha sido tan exhaustivamente evaluada como en adultos. Mientras que durante los últimos años se han realizado gran cantidad de estudios en adultos con anemia asociada al cáncer o a infección por HIV, permitiendo establecer conclusiones claras sobre su eficacia, sólo escasa cantidad de estudios con pequeño número de pacientes han sido realizados en niños. Hasta la fecha, los resultados sugieren que la terapia con rHuEPO en niños es tan útil como en adultos, pero la realización de estudios aleatorizados prospectivos incluyendo gran número de pacientes es esencial para alcanzar conclusiones definitivas. Los resultados de estudios dirigidos a evaluar la eficacia de la rHuEpo para mantener una dosis adecuada de ribavirina en pacientes en tratamiento por hepatitis C son alentadores. La utilización potencial de los efectos no hemopoyéticos de la rHuEPO en neonatos es un terreno novedoso y apasionante. El rol de la Epo como citoprotector para sistema nervioso central y mucosa intestinal está bajo investigación exhaustiva.Recombinant human erythropoietin (rHuEpo has become the most widely used cytokine in the world. Following the success of

  10. Teenage hyperthyroidism and radioiodine therapy

    International Nuclear Information System (INIS)

    Hussain, F.A.; Nisa, L.; Hoque, M.; Jehan, A.H.

    2007-01-01

    Full text: Objective: To assess the efficacy of radioiodine therapy in teenage patients with hyperthyroidism. Method: The medical records of 28 patients (age range 16 -18 years) were purposely selected from a pool of 3637 hyperthyroid patients treated with radioiodine during the period January 1982 to December 2006. Data for analysis included the age, sex, clinical symptoms, type of hyperthyroidism, antithyroid medications received, doses of radioiodine therapy given and the outcome of the therapy after one and five years. Results: The mean age of the patients treated for hyperthyroidism was 17.60 ± 0.73 years and 82 % were females. The most common type of hyperthyroidism was Graves' disease (75%). All patients were pre-treated with antithyroid drugs for variable duration periods (6 months to 2 years). The mean administered dose of radioiodine was 10.69 ± 2.77 mCi. The dose of I-131 was calculated on the basis of gland size and morphology (assessment by palpation and scintigraphy) and percentage of radioactive iodine uptake. Effective control of hyperthyroidism after radioiodine treatment occurred in 60.72% patients with a single dose, 35.71% required a second dose and 3.57% required more than two doses. Frequency of early hypothyroidism within one year was 38.09 % in Graves' disease and 33.33% in toxic multinodular goiter. Most patients (66.66%) in the toxic nodule group remained euthyroid up to one year after I-131 therapy. Overall incidence rates of hypothyroidism after 1 year and 5 years of radioiodine therapy were 32.14% and 75%, respectively. Conclusion: Hyperthyroidism in the teen group of patients was effectively controlled with radioiodine. There were no early side effects. The only long-term effect was hypothyroidism. Appearance of early/late hypothyroidism showed a direct relationship with the type of hyperthyroidism and the dose of radioiodine administered. In general, patients with Graves' disease showed a greater tendency in the evolution of early

  11. Radioiodine therapy for hyperthyroidism

    International Nuclear Information System (INIS)

    Andres-Barrenechea, E.C.

    2001-01-01

    A ten year study (1988-98) was done at the Veterans Memorial Medical Center on radioiodine (RAI) therapy for hyperthyroidism. A total of 162 patients received 131-I after careful selection hence was included in this study. A predominantly female population was seen (81%) compared to only (19%) males. The most frequent age group were in the third and fourth decades of life. Those included had clinical manifestations of thyrotoxicosis aside from the abnormal thyroid function tests and elevated RAI uptake. Almost all were given antithyroid drugs and beta-blockers prior to RAI ablation. Doses ranged from 7 to 12 mCi depending on gland size and degree of toxicity. Success rate of treatment was 92% which meant that the symptoms were abated and there was shrinkage of the thyroid gland after a single dose of iodine. The most common short term complications were sialitis and local neck tenderness while hypothyroidism was the commonest long term complication. (author)

  12. Radioiodine treatment of hyperthyroidism

    International Nuclear Information System (INIS)

    Becker, D.V.; Hurley, J.R.

    1988-01-01

    Hyperthyroidism is a clinical symptom complex caused by elevated thyroid hormone levels. Of the disorders in which it appears, the most common is Graves' disease; toxic nodular goiter (Plummer's disease) is considerably less frequent. Recent studies suggest that Graves' disease is autoimmune in origin and is probably caused by a genetic defect in immunoserveillance in which thyroid-stimulating antibodies interact with thyroid-stimulating hormone (TSH) receptor sites on thyroid follicular cell membranes to increase synthesis and secretion of thyroid hormones. Toxic nodular goiter is characterized by the secretion of increased amounts of thyroid hormone by localized autonomous areas of increased function within the thyroid gland. Other forms of hyperthyroidism are discussed. Radioiodine treatment of hyperthyroidism is described

  13. High-level secretion of native recombinant human calreticulin in yeast

    DEFF Research Database (Denmark)

    Čiplys, Evaldas; Žitkus, Eimantas; Gold, Leslie I.

    2015-01-01

    , Saccharomyces cerevisiae and Pichia pastoris. RESULTS: Expression of a full-length human CRT precursor including its native signal sequence resulted in high-level secretion of mature recombinant protein into the culture medium by both S. cerevisiae and P. pastoris. To ensure the structural and functional...... by non-denaturing PAGE. Moreover, limited trypsin digestion yielded identical fragment patterns of calcium-binding recombinant and native CRT suggesting that the yeast-derived CRT was correctly folded. Furthermore, both native and recombinant CRT induced cellular proliferation (MTS assay) and migration...... recombinant CRT protein with yields reaching 75 % of total secreted protein and with production levels of 60 and 200 mg/l from S. cerevisiae and P. pastoris, respectively. Finally, cultivation of P. pastoris in a bioreactor yielded CRT secretion titer to exceed 1.5 g/l of culture medium. CONCLUSIONS: Yeasts...

  14. A 30-year perspective on radioiodine therapy of benign nontoxic multinodular goiter

    DEFF Research Database (Denmark)

    Bonnema, Steen J; Hegedüs, Laszlo

    2009-01-01

    rate of permanent hypothyroidism, whereas other side-effects are very modest when using a rhTSH dose of 0.1 mg or below. SUMMARY: RhTSH-stimulated radioiodine therapy of benign nontoxic multinodular goiter is still an off-label use but several trials have shown that this treatment is significantly more......PURPOSE OF REVIEW: There is no consensus on the ideal treatment of patients with a benign nontoxic multinodular goiter. In some European countries, (131)I therapy has replaced surgery as the treatment of choice in these patients. Recombinant human thyrotropin (rhTSH) is a very potent stimulator...... of the thyroid gland and this review focuses on rhTSH-stimulated (131)I therapy. RECENT FINDINGS: The concept of rhTSH-stimulated (131)I therapy has been tested in several trials during the last 8 years. With this treatment, the goiter reduction is improved by 35-55%, compared with (131)I therapy without rh...

  15. Use of Recombinant Human Erythropoietin in Renal Anemia in Children

    Directory of Open Access Journals (Sweden)

    Habibur Rahman

    2009-11-01

    Full Text Available Erythropoietin is a hormone highly effective as like as natural erythropoietin to maintain target hemoglobin and hematocrit level in renal anemia. Its advantage over blood transfusion has been proved by improving the quality of life and decreasing morbidity and mortality in ESRD patients. Effectiveness of r-erythropoietin depends on absences of infection, inflammation and vitamin deficiency and iron status. Iron supplementation is needed before r-erythropoietin administration and sub-cutaneous rout is better in renal anemia because of slow and sustained releases of r-erythropoietin from the site of administration. Target hemoglobin level is 11-12.5 gm/dl and hematocrit is 33% which can be achieved by this hormone therapy. Key words- Recombinant erythropoietin, renal anemia, end stage renal disease.DOI: 10.3329/bsmmuj.v2i1.3713 BSMMU J 2009; 2(1: 50-53  

  16. Conjugation of gold nanoparticles and recombinant human endostatin modulates vascular normalization via interruption of anterior gradient 2-mediated angiogenesis.

    Science.gov (United States)

    Pan, Fan; Yang, Wende; Li, Wei; Yang, Xiao-Yan; Liu, Shuhao; Li, Xin; Zhao, Xiaoxu; Ding, Hui; Qin, Li; Pan, Yunlong

    2017-07-01

    Several studies have revealed the potential of normalizing tumor vessels in anti-angiogenic treatment. Recombinant human endostatin is an anti-angiogenic agent which has been applied in clinical tumor treatment. Our previous research indicated that gold nanoparticles could be a nanoparticle carrier for recombinant human endostatin delivery. The recombinant human endostatin-gold nanoparticle conjugates normalized vessels, which improved chemotherapy. However, the mechanism of recombinant human endostatin-gold nanoparticle-induced vascular normalization has not been explored. Anterior gradient 2 has been reported to be over-expressed in many malignant tumors and involved in tumor angiogenesis. To date, the precise efficacy of recombinant human endostatin-gold nanoparticles on anterior gradient 2-mediated angiogenesis or anterior gradient 2-related signaling cohort remained unknown. In this study, we aimed to explore whether recombinant human endostatin-gold nanoparticles could normalize vessels in metastatic colorectal cancer xenografts, and we further elucidated whether recombinant human endostatin-gold nanoparticles could interrupt anterior gradient 2-induced angiogenesis. In vivo, it was indicated that recombinant human endostatin-gold nanoparticles increased pericyte expression while inhibit vascular endothelial growth factor receptor 2 and anterior gradient 2 expression in metastatic colorectal cancer xenografts. In vitro, we uncovered that recombinant human endostatin-gold nanoparticles reduced cell migration and tube formation induced by anterior gradient 2 in human umbilical vein endothelial cells. Treatment with recombinant human endostatin-gold nanoparticles attenuated anterior gradient 2-mediated activation of MMP2, cMyc, VE-cadherin, phosphorylation of p38, and extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) in human umbilical vein endothelial cells. Our findings demonstrated recombinant human endostatin-gold nanoparticles might normalize

  17. Immunotoxicity assessment of rice-derived recombinant human serum albumin using human peripheral blood mononuclear cells.

    Directory of Open Access Journals (Sweden)

    Kai Fu

    Full Text Available Human serum albumin (HSA is extensively used in clinics to treat a variety of diseases, such as hypoproteinemia, hemorrhagic shock, serious burn injuries, cirrhotic ascites and fetal erythroblastosis. To address supply shortages and high safety risks from limited human donors, we recently developed recombinant technology to produce HSA from rice endosperm. To assess the risk potential of HSA derived from Oryza sativa (OsrHSA before a First-in-human (FIH trial, we compared OsrHSA and plasma-derived HSA (pHSA, evaluating the potential for an immune reaction and toxicity using human peripheral blood mononuclear cells (PBMCs. The results indicated that neither OsrHSA nor pHSA stimulated T cell proliferation at 1x and 5x dosages. We also found no significant differences in the profiles of the CD4(+ and CD8(+ T cell subsets between OsrHSA- and pHSA-treated cells. Furthermore, the results showed that there were no significant differences between OsrHSA and pHSA in the production of cytokines such as interferon-gamma (IFN-γ, tumor necrosis factor-alpha (TNF-α, interleukin (IL-10 and IL-4. Our results demonstrated that OsrHSA has equivalent immunotoxicity to pHSA when using the PBMC model. Moreover, this ex vivo system could provide an alternative approach to predict potential risks in novel biopharmaceutical development.

  18. Graves' disease and toxic nodular goiter - radioiodine therapy

    International Nuclear Information System (INIS)

    Schicha, H.; Dietlein, M.

    2002-01-01

    At the 15th conference on the human thyroid in Heidelberg in 2001 the following aspects of the radioiodine therapy of benign thyroid disorders were presented: General strategies for therapy of benign thyroid diseases, criterions for conservative or definitive treatment of hyperthyroidism as first line therapy and finally preparation, procedural details, results, side effects, costs and follow-up care of radioiodine therapy as well as legal guidelines for hospitalization in Germany. The diagnosis Graves' hyperthyroidism needs the decision, if rather a conservative treatment or if primary radioiodine therapy is the best therapeutic approach. In the USA 70-90% of these patients are treated with radioiodine as first line therapy, whereas in Germany the conservative therapy for 1-1.5 years is recommended for 90%. This review describes subgroups of patients with Graves' disease showing a higher probability to relapse after conservative treatment. Comparing benefits, adverse effects, costs, and conveniences of both treatment strategies the authors conclude that radioiodine therapy should be preferred as first line therapy in 60-70% of the patients with Graves' hyperthyroidism. (orig.) [de

  19. Recombinant human interleukin 2 acts as a B cell growth and differentiation promoting factor

    OpenAIRE

    Emmrich, F.; Moll, Heidrun; Simon, Markus M.

    2009-01-01

    Human B cells appropriately activated by a B cell mitogen are rendered susceptible to human Interleukin 2 (IL-2) as demonstrated with recombinant human IL-2 (rec. h IL-2). They show increased proliferation and drastically enhanced immunoglobulin secretion. Susceptibility to IL-2 is accompanied with the expression of the IL-2 receptor (Tac antigen) on B cells. The data suggest that IL-2 is one of the lymphokines directly involved in the activation of B lymphocytes.

  20. Preparation of mono-radioiodinated tracers for study of the in vivo metabolism of atrial natriuretic peptide in humans

    International Nuclear Information System (INIS)

    Clerico, A.; Marastoni, M.; Del Chicca, M.G.; Giannessi, D.; Del Ry, S.; Andreassi, M.G.; Sabatino, L.; Iascone, M.R.; Biagini, A.; Donato, L.

    1995-01-01

    The authors evaluate the optimum chemical conditions for labelling atrial natriuretic peptide (ANP) and its metabolites and for preparing highly purified radiotracers which can be used for in vivo kinetic studies of ANP in humans. Synthetic α h 1-28 ANP and some hormone metabolites were iodinated with Na 125 I or Na 131 I by means of the lactoperoxidase (ANP) or the chloramine-T (ANP metabolites) technique. The biological activity of labelled ANP was tested by means of a binding study using mouse cardiac membranes. A high-performance liquid chromatography (HPLC) procedure was used to purify the labelled hormone and the principal labelled metabolites in venous plasma samples collected up to 50 min after the injection of 125 I-labelled ANP from nine healthy men. The main ANP kinetic parameters were derived from the disappearance curves of the [ 125 I]ANP, which were satisfactorily fitted by a bi-exponential function in all subjects. The main advantages of this tracer technique are high accuracy, allowing the identification of the metabolites produced in vivo under steady-state conditions after injection of the precursor (labelled hormone) high sensitivity, allowing the detection of minimal quantities of metabolites high specificity, allowing the detection of possible in vitro artefactual generation of cleavage products of ANP using an internal labelled standard. Utilizing this tracer method, it was possible to estimate the principal parameters of ANP kinetics and also to plot the appearance curves of the labelled metabolites produced in vivo after the injection of the labelled precursor. (orig.). With 5 figs

  1. The 8p23 inversion polymorphism determines local recombination heterogeneity across human populations.

    Science.gov (United States)

    Alves, Joao M; Chikhi, Lounès; Amorim, António; Lopes, Alexandra M

    2014-04-01

    For decades, chromosomal inversions have been regarded as fascinating evolutionary elements as they are expected to suppress recombination between chromosomes with opposite orientations, leading to the accumulation of genetic differences between the two configurations over time. Here, making use of publicly available population genotype data for the largest polymorphic inversion in the human genome (8p23-inv), we assessed whether this inhibitory effect of inversion rearrangements led to significant differences in the recombination landscape of two homologous DNA segments, with opposite orientation. Our analysis revealed that the accumulation of genetic differentiation is positively correlated with the variation in recombination profiles. The observed recombination dissimilarity between inversion types is consistent across all populations analyzed and surpasses the effects of geographic structure, suggesting that both structures (orientations) have been evolving independently over an extended period of time, despite being subjected to the very same demographic history. Aside this mainly independent evolution, we also identified a short segment (350 kb, inversion) in the central region of the inversion where the genetic divergence between the two structural haplotypes is diminished. Although it is difficult to demonstrate it, this could be due to gene flow (possibly via double-crossing over events), which is consistent with the higher recombination rates surrounding this segment. This study demonstrates for the first time that chromosomal inversions influence the recombination landscape at a fine-scale and highlights the role of these rearrangements as drivers of genome evolution.

  2. Radioiodinated ligands for dopamine receptors

    International Nuclear Information System (INIS)

    Kung, H.F.

    1994-01-01

    The dopamine receptor system is important for normal brain function; it is also the apparent action site for various neuroleptic drugs for the treatment of schizophrenia and other metal disorders. In the past few years radioiodinated ligands for single photon emission tomography (SPECT) have been successfully developed and tested in humans: [ 123 I]TISCH for D1 dopamine receptors; [ 123 I]IBZM, epidepride, IBF and FIDA2, four iodobenzamide derivatives, for D2/D3 dopamine receptors. In addition, [ 123 I]β-CIT (RTI-55) and IPT, cocaine derivatives, for the dopamine reuptake site are potentially useful for diagnosis of loss of dopamine neurons. The first iodinated ligand, (R)trans-7-OH-PIPAT, for D3 dopamine receptors, was synthesized and characterized with cloned cell lines (Spodoptera frugiperda, Sf9) expressing the D2 and D3 dopamine receptors and with rat basal forebrain membrane preparations. Most of the known iodobenzamides displayed similar potency in binding to both D2 and D3 dopamine receptors expressed in the cell lines. Initial studies appear to suggest that by fine tuning the structures it may be possible to develop agents specific for D2 and D3 dopamine receptors. It is important to investigate D2/D3 selectivity for this series of potent ligands

  3. Effects of recombinant human collagen VI from Escherichia coli on ...

    African Journals Online (AJOL)

    Jane

    2011-07-20

    Jul 20, 2011 ... In this study, we reported the cloning and over expression of a gene coding for human collagen peptide. (CP6) in Escherichia coli and investigated the protective effects of CP6 on UVA-irradiated human skin fibroblasts cells. The collagen peptide (CP6) was highly soluble and the expression level was.

  4. Radioiodination of antibodies for tumor imaging

    International Nuclear Information System (INIS)

    Saha, G.B.

    1983-01-01

    In view of the great potential of radioiodinated antibody for the detection and treatment of cancer, the present article deals with the various techniques of radioiodination of antibody and their uses. Topics include methods of iodination of antibody, advantages and disadvantages of different methods, and effects of radioiodination on the antibody molecules with respect to their physiochemical and immunologic reactivity. In addition, the clinical usefulness of radioiodinated antibodies is discussed. (Auth.)

  5. Characterization of receptors for recombinant human tumor necrosis factor-alpha from human placental membranes

    International Nuclear Information System (INIS)

    Aiyer, R.A.; Aggarwal, B.B.

    1990-01-01

    High affinity receptors for recombinant human tumor necrosis factor-alpha (rhTNF-alpha) were identified on membranes prepared from full term human placenta. Highly purified rhTNF-alpha iodinated by the iodogen method was found to bind placental membranes in a displaceable manner with an approximate dissociation constant (KD) of 1.9 nM. The membrane bound TNF-alpha receptor could be solubilized by several detergents with optimum extraction being obtained with 1% Triton X-100. The binding of 125I-rhTNF-alpha to the solubilized receptor was found to be time and temperature dependent, yielding maximum binding within 1 h, 24 h and 48 h at 37 degrees C, 24 degrees C and 4 degrees C, respectively. However, the maximum binding obtainable at 4 degrees C was only 40% of that at 37 degrees C. The binding 125I-rhTNF-alpha to solubilized placental membrane extracts was displaceable by unlabeled rhTNF-alpha, but not by a related protein recombinant human tumor necrosis factor-beta (rhTNF-beta; previously called lymphotoxin). This is similar to the behavior of TNF-alpha receptors derived from detergent-solubilized cell extracts, although on intact cells, both rhTNF-alpha and rhTNF-beta bind with equal affinity to TNF receptors. The Scatchard analysis of the binding data of the solubilized receptor revealed high affinity binding sites with a KD of approximately 0.5 nM and a receptor concentration of about 1 pmole/mg protein. Gel filtration of the solubilized receptor-ligand complexes on Sephacryl S-300 revealed two different peaks of radioactivity at approximate molecular masses of 50,000 Da and 400,000 Da. The 400,000 dalton peak corresponded to the receptor-ligand complex. Overall, our results suggest that high affinity receptors for TNF-alpha are present on human placental membranes and provide evidence that these receptors may be different from that of rhTNF-beta

  6. High-resolution recombination patterns in a region of human chromosome 21 measured by sperm typing.

    Directory of Open Access Journals (Sweden)

    Irene Tiemann-Boege

    2006-05-01

    Full Text Available For decades, classical crossover studies and linkage disequilibrium (LD analysis of genomic regions suggested that human meiotic crossovers may not be randomly distributed along chromosomes but are focused instead in "hot spots." Recent sperm typing studies provided data at very high resolution and accuracy that defined the physical limits of a number of hot spots. The data were also used to test whether patterns of LD can predict hot spot locations. These sperm typing studies focused on several small regions of the genome already known or suspected of containing a hot spot based on the presence of LD breakdown or previous experimental evidence of hot spot activity. Comparable data on target regions not specifically chosen using these two criteria is lacking but is needed to make an unbiased test of whether LD data alone can accurately predict active hot spots. We used sperm typing to estimate recombination in 17 almost contiguous ~5 kb intervals spanning 103 kb of human Chromosome 21. We found two intervals that contained new hot spots. The comparison of our data with recombination rates predicted by statistical analyses of LD showed that, overall, the two datasets corresponded well, except for one predicted hot spot that showed little crossing over. This study doubles the experimental data on recombination in men at the highest resolution and accuracy and supports the emerging genome-wide picture that recombination is localized in small regions separated by cold areas. Detailed study of one of the new hot spots revealed a sperm donor with a decrease in recombination intensity at the canonical recombination site but an increase in crossover activity nearby. This unique finding suggests that the position and intensity of hot spots may evolve by means of a concerted mechanism that maintains the overall recombination intensity in the region.

  7. Application of Recombinant Proteins for Serodiagnosis of Visceral Leishmaniasis in Humans and Dogs.

    Science.gov (United States)

    Farahmand, Mahin; Nahrevanian, Hossein

    2016-07-01

    Visceral leishmaniasis (VL) is a zoonotic disease caused by leishmania species. Dogs are considered to be the main reservoir of VL. A number of methods and antigen-based assays are used for the diagnosis of leishmaniasis. However, currently available methods are mainly based on direct examination of tissues for the presence of parasites, which is highly invasive. A variety of serological tests are commonly applied for VL diagnosis, including indirect fluorescence antibody test, enzyme-linked immunosorbent assay (ELISA), dot-ELISA, direct agglutination test, Western-blotting, and immunochromatographic test. However, when soluble antigens are used, serological tests are less specific due to cross-reactivity with other parasitic diseases. Several studies have attempted to replace soluble antigens with recombinant proteins to improve the sensitivity and the specificity of the immunodiagnostic tests. Major technological advances in recombinant antigens as reagents for the serological diagnosis of VL have led to high sensitivity and specificity of these serological tests. A great number of recombinant proteins have been shown to be effective for the diagnosis of leishmania infection in dogs, the major reservoir of L. infantum. Although few recombinant proteins with high efficacy provide reasonable results for the diagnosis of human and canine VL, more optimization is still needed for the appropriate antigens to provide high-throughput performance. This review aims to explore the application of different recombinant proteins for the serodiagnosis of VL in humans and dogs.

  8. Genetic battle between Helicobacter pylori and humans. The mechanism underlying homologous recombination in bacteria, which can infect human cells.

    Science.gov (United States)

    Hanada, Katsuhiro; Yamaoka, Yoshio

    2014-10-01

    Helicobacter pylori is a gram-negative pathogenic bacterium that colonises the human stomach. The chronic infection it causes results in peptic ulcers and gastric cancers. H. pylori can easily establish a chronic infection even if the immune system attacks this pathogen with oxidative stress agents and immunoglobulins. This is attributed to bacterial defence mechanisms against these stresses. As a defence mechanism against oxidative stresses, in bacterial genomes, homologous recombination can act as a repair pathway of DNA's double-strand breaks (DSBs). Moreover, homologous recombination is also involved in the antigenic variation in H. pylori. Gene conversion alters genomic structures of babA and babB (encoding outer membrane proteins), resulting in escape from immunoglobulin attacks. Thus, homologous recombination in bacteria plays an important role in the maintenance of a chronic infection. In addition, H. pylori infection causes DSBs in human cells. Homologous recombination is also involved in the repair of DSBs in human cells. In this review, we describe the roles of homologous recombination with an emphasis on the maintenance of a chronic infection. Copyright © 2014 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  9. Intraarticular Sprifermin (Recombinant Human Fibroblast Growth Factor 18) in Knee Osteoarthritis

    DEFF Research Database (Denmark)

    Lohmander, L. S.; Hellot, S.; Dreher, D.

    2014-01-01

    Objective. To evaluate the efficacy and safety of intraarticular sprifermin (recombinant human fibroblast growth factor 18) in the treatment of symptomatic knee osteoarthritis (OA). Methods. The study was a randomized, double-blind, placebo-controlled, proof-of-concept trial. Intraarticular sprif...

  10. Antigenic profile of human recombinant PrP: generation and chracterization of a versatile polyclonal antiserum

    NARCIS (Netherlands)

    Sachsamanoglou, M.; Paspaltzis, I.; Petrakis, S.; Verghese-Nikolakaki, S.; Panagiotidis, C.H.; Voitlander, T.; Budka, H.; Langeveld, J.P.M.; Sklaviadis, T.

    2004-01-01

    We describe the quality of a rabbit polyclonal antiserum (Sal1) that was raised against mature human recombinant prion protein (rhuPrP). Epitope mapping demonstrated that the Sal1 antiserum recognized six to eight linear antigenic sites, depending on the animal species. The versatility of the

  11. IgM-specific serodiagnosis of acute human cytomegalovirus infection using recombinant autologous fusion proteins

    NARCIS (Netherlands)

    Vornhagen, R; Hinderer, W; Sonneborn, HH; Bein, G; Matter, L; The, T. Hauw; Enders, G; Jahn, G; Plachter, B

    Portions of three human cytomegalovirus (HCMV) polypeptides, which were shown previously to be highly reactive with patient sera, were expressed in Escherichia coli as autologous fusion proteins. Purified recombinant polypeptides were used as antigens in enzyme linked immunosorbent assay (ELISA) and

  12. Structural Evolution of Human Recombinant alfaB-Crystallin under UV Irradiation

    DEFF Research Database (Denmark)

    Sugiyama, Masaaki; Fujii, Noriko; Morimoto, Yukio

    2008-01-01

    External stresses cause certain proteins to lose their regular structure and aggregate. In order to clarify this abnormal aggregation process, a structural evolution of human recombinant aB-crystallin under UV irradiation was observed with in situ small-angle neutron scattering. The abnormal...

  13. Human recombinant protein C for severe sepsis and septic shock in adult and paediatric patients

    DEFF Research Database (Denmark)

    Martí-Carvajal, Arturo J; Solà, Ivan; Gluud, Christian

    2012-01-01

    Sepsis is a common and frequently fatal condition. Human recombinant activated protein C (APC) has been introduced to reduce the high risk of death associated with severe sepsis or septic shock. This systematic review is an update of a Cochrane review originally published in 2007....

  14. Expression of the human multidrug transporter in insect cells by a recombinant baculovirus

    International Nuclear Information System (INIS)

    Germann, U.A.; Willingham, M.C.; Pastan, I.; Gottesman, M.M.

    1990-01-01

    The plasma membrane associated human multidrug resistance (MDR1) gene product, known as the 170-kDa P-glycoprotein or the multidrug transporter, acts as an ATP-dependent efflux pump for various cytotoxic agents. The authors expressed recombinant human multidrug transporter in a baculovirus expression system to obtain large quantities and further investigate its structure and mechanism of action. MDR1 cDNA was inserted into the genome of the Autographa californica nuclear polyhedrosis virus under the control of the polyhedrin promoter. Spodoptera frugiperda insect cells synthesized high levels of recombinant multidrug transporter 2-3 days after infection. The transporter was localized by immunocytochemical methods on the external surface of the plasma membranes, in the Golgi apparatus, and within the nuclear envelope. The human multidrug transporter expressed in insect cells is not susceptible to endoglycosidase F treatment and has a lower apparent molecular weight of 140,000, corresponding to the nonglycosylated precursor of its authentic counterpart expressed in multidrug-resistant cells. Labeling experiments showed that the recombinant multidrug transporter is phosphorylated and can be photoaffinity labeled by [ 3 H]azidopine, presumably at the same two sites as the native protein. Various drugs and reversing agents compete with the [ 3 H]azidopine binding reaction when added in excess, indicating that the recombinant human multidrug transporter expressed in insect cells is functionally similar to its authentic counterpart

  15. A Child with Local Lipohypertrophy following Recombinant Human Growth Hormone Administration

    NARCIS (Netherlands)

    Koppen, Ilan J. N.; Bakx, Roel; de Kruiff, Chris C.; van Trotsenburg, A. S. Paul

    2016-01-01

    Local lipohypertrophy due to recombinant human growth hormone (rhGH) administration is a rare phenomenon. Here, we report a case of an 11-year-old girl who presented with a paraumbilical swelling, approximately one year after the start of rhGH treatment for short stature due to the presumed

  16. RECOMBINANT HUMAN INTERLEUKIN-6 INDUCES A RAPID AND REVERSIBLE ANEMIA IN CANCER-PATIENTS

    NARCIS (Netherlands)

    NIEKEN, J; MULDER, NH; VELLENGA, E; LIMBURG, PC; PIERS, DA; DEVRIES, EGE

    1995-01-01

    Initial studies have shown that recombinant human interleukin-6 (rhIL-6) induces anemia. Until now, the pathophysiologic mechanism of this induced anemia has been unknown. To unravel the underlying mechanism, we examined 15 cancer patients receiving rhIL-6 as an antitumor immunotherapy in a phase II

  17. Development of a transgenic mouse model to study the immunogenicity of recombinant human insulin

    NARCIS (Netherlands)

    Torosantucci, Riccardo; Brinks, Vera; Kijanka, Grzegorz; Halim, Liem Andhyk; Sauerborn, Melody; Schellekens, Huub; Jiskoot, Wim

    2014-01-01

    Mouse models are commonly used to assess the immunogenicity of therapeutic proteins and to investigate the immunological processes leading to antidrug antibodies. The aim of this work was to develop a transgenic (TG) Balb/c mouse model for evaluating the immunogenicity of recombinant human insulin

  18. Short-term effect of recombinant human growth hormone in patients with alcoholic cirrhosis

    DEFF Research Database (Denmark)

    Møller, S; Becker, U; Grønbaek, M

    1994-01-01

    As growth hormone possesses anabolic properties that are active on protein metabolism, and thus of potential benefit to patients with chronic liver disease, we determined the metabolic effects of recombinant human growth hormone on insulin-like growth factor-I (IGF-I) its specific binding proteins...

  19. Recombinant human laminin isoforms can support the undifferentiated growth of human embryonic stem cells

    International Nuclear Information System (INIS)

    Miyazaki, Takamichi; Futaki, Sugiko; Hasegawa, Kouichi; Kawasaki, Miwa; Sanzen, Noriko; Hayashi, Maria; Kawase, Eihachiro; Sekiguchi, Kiyotoshi; Nakatsuji, Norio; Suemori, Hirofumi

    2008-01-01

    Human embryonic stem cells (hESCs) are thought to be a promising cell source for cell transplantation therapy. For such a clinical application, the hESCs should be manipulated using appropriate and qualified materials. In this study, we examined the efficacy of recombinant human laminin (rhLM) isoforms on the undifferentiated growth of hESCs. We first determined the major integrins expressed on the hESCs to reveal the preference of the hESCs for rhLMs, and found that the hESCs mainly expressed integrin α6β1, which binds predominantly to laminin-111, -332 and -511/-521. When the hESCs were seeded onto rhLMs, the cells indeed adhered markedly to rhLM-332, and to rhLM-511 and rhLM-111 to a lesser extent. The hESCs proliferated on these three rhLMs for several passages while preserving their pluripotency. These results show that rhLM-111, -332, and -511 are good substrates to expand undifferentiated hESCs due to their high affinity to integrin α6β1 expressed on hESCs

  20. Radioiodination of humic substances

    Energy Technology Data Exchange (ETDEWEB)

    Franke, K.; Kupsch, H. [Inst. of Interdisciplinary Isotope Research, Leipzig (Germany)

    2010-07-01

    The known IODO-GEN trademark -method was adapted for radiolabeling of humic and fulvic acids with {sup 131}I. The water insoluble oxidizing agent 1,3,4,6tetrachloro-3{alpha},6{alpha}-diphenylglycoluril (IODO-GEN trademark) forms an iodous ion species (I{sup +}), which undergoes an electrophilic I/H-substitution on aromatic moieties of the humic and fulvic acids. This method offers mild conditions with a lesser extent of oxidative alterations of the target molecule, accompanied by an easy handling due to the virtual water-insolubility of the oxidizing agent. The method was optimized and different techniques were tested for the purification of the radioiodinated humic material. The yield of the labeling procedure varies between 45 and 75% depending on the provenance of the humic material and the applied purification method. A specific activity up to 40 MBq/mg was achieved. Furthermore, the known inherent photo-susceptibility of the iodinated humic substance and the influence of reducing agents were verified. An additional release of {sup 131}I up to 20% and up to 35%, respectively were observed. (orig.)

  1. Kinetics of photodissociated oxygen recombination to human oxyhemoglobin

    International Nuclear Information System (INIS)

    Bokut', S.B.; Syakhovich, V.E.; Parul', D.A.; Lepeshkevich, S.V.; Dzhagarov, B.M.

    2001-01-01

    Oxygen binding to the tetrameric hemoglobin (Hb) is a basic reaction for study of a cooperativity and allosteric homotropic and heterotropic interactions in proteins. In tetrameric hemoglobin the certain sites in the α 1 β 2 -interface have the precise geometry and chemical reactivity to bind 2,3-diphosphoglycerate, protons, chloride and hence shift the equilibrium away from the oxyconformation, thereby favoring O 2 release. Post-translational modifications of the major hemoglobin fraction Hb A 1 with sugar moiety in the Hb central cavity leads to differences in geometry of the effectors binding region providing a useful experimental tool to study the long range relationship in the tetramer molecule. Here we present the results of the nongeminate biomolecular association of Hb and O 2 obtained by nanosecond laser flash-photolysis. All measurements were carried out in 50 mM potassium-phosphate buffer pH 7.4 with the following samples Hb A 1 , HbA 1c , HbA 1b , and HbA 1 in the presence of the tenfold excess of inositol hexaphosphate (IHP). Our results show that oxygen recombination kinetics are characterized by two processes with different decay times and Hb-form-dependent contributions. This process can be described by the following expression: A(t)=A 1 exp(-t/τ 1 )+A 2 exp(-t/τ 2 ), where A(t) is a normalized number of the deoxy-Hb molecules. The short-live component has a lifetime τ 1 , which is Hb-type dependent and changes in the intervals 30-60 μs, the second component has a lifetime τ 2 around 100 μs, and also is sample-dependent value. A(t=0) is proportional to apparent quantum yields of the photodissociation and determines by geminate stages of oxygen binding to Fe from the protein matrix areas. These results show that post-translational modifications of the major hemoglobin component HbA 1 have influence on hemoglobin transport function via the long range relationship in the tetramer molecule

  2. Limited infection upon human exposure to a recombinant raccoon pox vaccine vector

    Science.gov (United States)

    Rocke, T.E.; Dein, F.J.; Fuchsberger, M.; Fox, B.C.; Stinchcomb, D.T.; Osorio, J.G.

    2004-01-01

    A laboratory accident resulted in human exposure to a recombinant raccoon poxvirus (RCN) developed as a vaccine vector for antigens of Yersinia pestis for protection of wild rodents (and other animals) against plague. Within 9 days, the patient developed a small blister that healed within 4 weeks. Raccoon poxvirus was cultured from the lesion, and the patient developed antibody to plague antigen (F1) and RCN. This is the first documented case of human exposure to RCN.

  3. Limited infection upon human exposure to a recombinant raccoon pox vaccine vector.

    Science.gov (United States)

    Rocke, Tonie E; Dein, F Joshua; Fuchsberger, Martina; Fox, Barry C; Stinchcomb, Dan T; Osorio, Jorge E

    2004-07-29

    A laboratory accident resulted in human exposure to a recombinant raccoon poxvirus (RCN) developed as a vaccine vector for antigens of Yersinia pestis for protection of wild rodents (and other animals) against plague. Within 9 days, the patient developed a small blister that healed within 4 weeks. Raccoon poxvirus was cultured from the lesion, and the patient developed antibody to plague antigen (F1) and RCN. This is the first documented case of human exposure to RCN.

  4. Comparison of pituitary and recombinant human thyrotropin standards in an immunoradiometric system

    International Nuclear Information System (INIS)

    Blanca Fernandez, Silvia; Rodriguez Gonzalez, Julio Cesar; Nisembaum Alas, Amaparo; Sevy Gonzalez, O.

    1998-01-01

    Results of two standards of human thyrotropin of pituitaries (B) and recombinant (C) origen supplied by the Instituto of pesquisas Energeticas y Nucleares, Brazil, were compared in our immunoradiometric reference system that use an human thyrotropin pituitary standard of local production (A). This work was supported by the International Atomic Energy Agency for an inter-regional comparison and set up of a reference standard

  5. Frequency and genetic characterization of V(DD)J recombinants in the human peripheral blood antibody repertoire.

    Science.gov (United States)

    Briney, Bryan S; Willis, Jordan R; Hicar, Mark D; Thomas, James W; Crowe, James E

    2012-09-01

    Antibody heavy-chain recombination that results in the incorporation of multiple diversity (D) genes, although uncommon, contributes substantially to the diversity of the human antibody repertoire. Such recombination allows the generation of heavy chain complementarity determining region 3 (HCDR3) regions of extreme length and enables junctional regions that, because of the nucleotide bias of N-addition regions, are difficult to produce through normal V(D)J recombination. Although this non-classical recombination process has been observed infrequently, comprehensive analysis of the frequency and genetic characteristics of such events in the human peripheral blood antibody repertoire has not been possible because of the rarity of such recombinants and the limitations of traditional sequencing technologies. Here, through the use of high-throughput sequencing of the normal human peripheral blood antibody repertoire, we analysed the frequency and genetic characteristics of V(DD)J recombinants. We found that these recombinations were present in approximately 1 in 800 circulating B cells, and that the frequency was severely reduced in memory cell subsets. We also found that V(DD)J recombination can occur across the spectrum of diversity genes, indicating that virtually all recombination signal sequences that flank diversity genes are amenable to V(DD)J recombination. Finally, we observed a repertoire bias in the diversity gene repertoire at the upstream (5') position, and discovered that this bias was primarily attributable to the order of diversity genes in the genomic locus. © 2012 The Authors. Immunology © 2012 Blackwell Publishing Ltd.

  6. Expression of Recombinant Streptokinase from Streptococcus Pyogenes and its Reaction with Infected Human and Murine Sera

    Science.gov (United States)

    Molaee, Neda; Abtahi, Hamid; Mosayebi, Ghasem

    2013-01-01

    Objective(s): Streptokinase (SKa) is an antigenic protein which is secreted by Streptococcus pyogenes. Streptokinase induces inflammation by complement activation, which may play a role in post infectious diseases. In the present study, recombinant streptokinase from S. pyogenes was produced and showed that recombinant SKa protein was recognized by infected human sera using Western blot analysis. Materials and Methods: In this study, the ska gene from S. pyogenes was amplified and cloned into pET32a which is a prokaryotic expression vector. pET32a-ska was transformed to Escherichia coli BL21 (DE3) pLysS and gene expression was induced by IPTG. Protein production was improved by modification of composition of the bacterial culture media and altering the induction time by IPTG. The expressed protein was purified by affinity chromatography using the Ni-NTA resin. The integrity of the product was confirmed by Westernblot analysis using infected mice. Serum reactivity of five infected individuals was further analyzed against the recombinant SKa protein. Results: Data indicated that recombinant SKa protein from S. pyogenes can be recognized by patient and mice sera. The concentration of the purified recombinant protein was 3.2 mg/L of initial culture. The highest amount of the expressed protein after addition of IPTG was obtained in a bacterial culture without glucose with the culture optical density of 0.8 (OD600 = 0.8). Conclusion : Present data shows, recombinant SKa protein has same epitopes with natural form of this antigen. Recombinant SKa also seemed to be a promising antigen for the serologic diagnosis of S. pyogenes infections. PMID:24171077

  7. Expression of Recombinant Streptokinase from Streptococcus Pyogenes and Its Reaction with Infected Human and Murine Sera

    Directory of Open Access Journals (Sweden)

    Neda Molaee

    2013-09-01

    Full Text Available   Objective(s: Streptokinase (SKa is an antigenic protein which is secreted by Streptococcus pyogenes. Streptokinase induces inflammation by complement activation, which may play a role in post infectious diseases. In the present study, recombinant streptokinase from S. pyogenes was produced and showed that recombinant SKa protein was recognized by infected human sera using Western blot analysis.   Materials and Methods: In this study, the ska gene from S. pyogenes was amplified and cloned into pET32a which is a prokaryotic expression vector. pET32a-ska was transformed to Escherichia coli BL21 (DE3 pLysS and gene expression was induced by IPTG. Protein production was improved by modification of composition of the bacterial culture media and altering the induction time by IPTG. The expressed protein was purified by affinity chromatography using the Ni-NTA resin. The integrity of the product was confirmed by Westernblot analysis using infected mice. Serum reactivity of five infected individuals was further analyzed against the recombinant SKa protein. Results: Data indicated that recombinant SKa protein from S. pyogenes can be recognized by patient and mice sera. The concentration of the purified recombinant protein was 3.2 mg/L of initial culture. The highest amount of the expressed protein after addition of IPTG was obtained in a bacterial culture without glucose with the culture optical density of 0.8 (OD600 = 0.8. Conclusion : Present data shows, recombinant SKa protein has same epitopes with natural form of this antigen. Recombinant SKa also seemed to be a promising antigen for the serologic diagnosis of S. pyogenes infections.

  8. Recombinant production of human Aquaporin-1 to an exceptional high membrane density in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Julie Bomholt

    Full Text Available In the present paper we explored the capacity of yeast Saccharomyces cerevisiae as host for heterologous expression of human Aquaporin-1. Aquaporin-1 cDNA was expressed from a galactose inducible promoter situated on a plasmid with an adjustable copy number. Human Aquaporin-1 was C-terminally tagged with yeast enhanced GFP for quantification of functional expression, determination of sub-cellular localization, estimation of in vivo folding efficiency and establishment of a purification protocol. Aquaporin-1 was found to constitute 8.5 percent of total membrane protein content after expression at 15°C in a yeast host over-producing the Gal4p transcriptional activator and growth in amino acid supplemented minimal medium. In-gel fluorescence combined with western blotting showed that low accumulation of correctly folded recombinant Aquaporin-1 at 30°C was due to in vivo mal-folding. Reduction of the expression temperature to 15°C almost completely prevented Aquaporin-1 mal-folding. Bioimaging of live yeast cells revealed that recombinant Aquaporin-1 accumulated in the yeast plasma membrane. A detergent screen for solubilization revealed that CYMAL-5 was superior in solubilizing recombinant Aquaporin-1 and generated a monodisperse protein preparation. A single Ni-affinity chromatography step was used to obtain almost pure Aquaporin-1. Recombinant Aquaporin-1 produced in S. cerevisiae was not N-glycosylated in contrast to the protein found in human erythrocytes.

  9. Selection of LNA-containing DNA aptamers against recombinant human CD73

    DEFF Research Database (Denmark)

    Elle, Ida C; Karlsen, Kasper K; Terp, Mikkel G

    2015-01-01

    tested by surface plasmon resonance. Truncated variants of these aptamers and variants where the LNA nucleotides were substituted for the DNA equivalent also exhibited affinity for the recombinant CD73 in the low nanomolar range. In enzyme inhibition assays with recombinant CD73 the aptamer sequences......LNA-containing DNA aptamers against CD73 (human ecto-5'-nucleotidase), a protein frequently overexpressed in solid tumours, were isolated by SELEX. A pre-defined stem-loop library, containing LNA in the forward primer region, was enriched with CD73 binding sequences through six rounds of SELEX...... with recombinant his-tagged CD73 immobilised on anti-his plates. Enriched pools isolated from rounds one, three and six were subjected to next-generation sequencing and analysed for enrichment using custom bioinformatics software. The software identified aptamer sequences via the primers and then performed several...

  10. Radioiodine removal in nuclear facilities

    International Nuclear Information System (INIS)

    1980-01-01

    Technical means are reviewed available for the retention of radioiodine in nuclear power plants and fuel reprocessing plants, its immobilization, storage, and disposal. The removal of iodine species from gaseous effluents of nuclear power plants using impregnated activated charcoal is dealt with. Various scrubbing techniques for trapping iodine from the head-end and dissolver off-gases are discussed as well as solid adsorbents for iodine which may be used to clean up other gaseous streams. Current practices and activities for radioiodine treatment and management in Belgian, Dutch, Swedish, USSR and UK nuclear installations are presented

  11. Endocrine ophthalmopathy and radioiodine therapy

    International Nuclear Information System (INIS)

    Karlsson, F. Anders

    2006-01-01

    Endocrine ophthalmopathy is to some degree present in most patients with Graves' disease. In few cases, a severe form of the condition develops and in the majority of these cases, the course of the eye problems has been influenced by the treatment for thyrotoxicosis. In this regard, radioiodine therapy has been increasingly recognized as carrying a special risk. Here, the current understanding of endocrine ophthalmopathy and the risks associated with the development of severe eye disease are discussed. The results of a retrospective investigation of patients with severe eye disease in our hospital, and the experience with corticosteroid administration following radioiodine in order to reduce the risk of ophthalmopathy, are also presented

  12. Radioiodine therapy and thyrostatic drugs and iodine

    Energy Technology Data Exchange (ETDEWEB)

    Moka, D.; Dietlein, M.; Schicha, H. [Department of Nuclear Medicine, University of Cologne, Joseph Stelzmannstrasse 9, 50924 Koeln (Germany)

    2002-08-01

    Radioiodine therapy is now the most common definite treatment for persistent hyperthyroidism. The outcome of radioiodine therapy depends mainly on the absorbed energy dose in the diseased thyroid tissue. The administered activity and the resulting target dose in the thyroid depend on both the biokinetics of radioiodine and the actual therapeutic effect of radioiodine in the thyroid. Thyrostatic drugs have a major influence on the kinetics of radioiodine in the thyroid and may additionally have a radioprotective effect. Pre-treatment with thyrostatic medication lowers the effective half-life and uptake of radioiodine. This can reduce the target dose in the thyroid and have a negative influence on the outcome of the therapy. Discontinuation of medication shortly before radioiodine administration can increase the absorbed energy dose in the thyroid without increasing the whole-body exposure to radiation as much as would a higher or second radioiodine administration. Furthermore, administration of non-radioactive iodine-127 2-3 days after radioiodine administration can also increase the effective half-life of radioiodine in the thyroid. Thus, improving the biokinetics of radioiodine will allow lower activities to be administered with lower effective doses to the rest of the body, while achieving an equally effective target dose in the thyroid. (orig.)

  13. Quality control of radioiodinated gastrin for radioimmunoassay

    International Nuclear Information System (INIS)

    Ginabreda, M.G.P.; Borghi, V.C.; Bettarello, A.

    1988-07-01

    Radioiodinated human gastrin has been prepared at IPEN laboratory for radioimmunoassay use. This work developed the quality control of this tracer analyzing parameters of the labelling reaction, chromatographic purification and radioimmunoassay. The radioiodination yield obtained in five experiments was reproducible and similar when analyzed on 7% polyaraylamide gel eletrophoresis - PAGE - (mean + - SD of 51.70 + - 10.76%) and by1 25 I incorporation checked through thrichloroacetic acid precipitation - TCA - (57-36 + - 9.69%). Similary, after purification the labelled gastrin revaled high and reproducible purity degree when submitted to PAGE (96.57 + - 1.06%) and CA (94.82 + - 4.20%) analysis. The respective specific activities varied from 62 to 307 uCi/ug, being determined by the self-displacement method, which is based on the immunoactivity of the tracer. In this way, the antibody titers required to bind 50% of the tracer ranged from 1:32.000 to 1:180.000. Consequently, the respective doses producing 50% fall in the maximum response of the radioimmunoassays ranged from 155.0 to 24.0 pmol/1, but remained unchanged for each tracer even after three months of its preparations. The tracers presented very low non-specific binding values (1.78 + - 0.79%), stablespecific binding values (46.49 + - 5.65%) and a good between-assay precision, evaluated by an internal quality control sample (25.71 + - 4.30%) with coefficient of variation of 16.74%). The PAGE analysis of the unlabeled gastrin used in the first and last radioiodination revealed an unique and unaltered component, confirming the quality of the tracers. (author) [pt

  14. An integrative 'omics' solution to the detection of recombinant human erythropoietin and blood doping.

    Science.gov (United States)

    Pitsiladis, Yannis P; Durussel, Jérôme; Rabin, Olivier

    2014-05-01

    Administration of recombinant human erythropoietin (rHumanEPO) improves sporting performance and hence is frequently subject to abuse by athletes, although rHumanEPO is prohibited by the WADA. Approaches to detect rHumanEPO doping have improved significantly in recent years but remain imperfect. A new transcriptomic-based longitudinal screening approach is being developed that has the potential to improve the analytical performance of current detection methods. In particular, studies are being funded by WADA to identify a 'molecular signature' of rHumanEPO doping and preliminary results are promising. In the first systematic study to be conducted, the expression of hundreds of genes were found to be altered by rHumanEPO with numerous gene transcripts being differentially expressed after the first injection and further transcripts profoundly upregulated during and subsequently downregulated up to 4 weeks postadministration of the drug; with the same transcriptomic pattern observed in all participants. The identification of a blood 'molecular signature' of rHumanEPO administration is the strongest evidence to date that gene biomarkers have the potential to substantially improve the analytical performance of current antidoping methods such as the Athlete Biological Passport for rHumanEPO detection. Given the early promise of transcriptomics, research using an 'omics'-based approach involving genomics, transcriptomics, proteomics and metabolomics should be intensified in order to achieve improved detection of rHumanEPO and other doping substances and methods difficult to detect such a recombinant human growth hormone and blood transfusions.

  15. Recombinant human acetylcholine receptor alpha-subunit induces chronic experimental autoimmune myasthenia gravis.

    Science.gov (United States)

    Lennon, V A; Lambert, E H; Leiby, K R; Okarma, T B; Talib, S

    1991-04-01

    A synthetic gene encoding the 210 N-terminal residues of the alpha-subunit of the nicotinic acetylcholine receptor (AChR) of human skeletal muscle was cloned into an inducible expression plasmid to produce a fusion protein in high yield in Escherichia coli. Like native human AChR, the recombinant human alpha 1-210 protein induced AChR-binding, AChR-modulating, and AChR-blocking autoantibodies in rats when injected once intradermally as an emulsion in CFA, with Bordetella pertussis vaccine as supplementary adjuvant. The minimum dose of recombinant protein required to induce biochemical signs of experimental autoimmune myasthenia gravis (EAMG) with 100% incidence was 2.2 micrograms. With 6.6 to 22 micrograms, serum levels of autoantibodies were persistent, and clinically apparent EAMG lasted more than a month. Clinical, electrophysiological, and biochemical indices of EAMG induced by doses of 66 micrograms or more were more uniformly severe and persistent, with 33% fatality. Rats receiving a control extract of E. coli containing plasmid without the alpha 1-210 codon insert, with adjuvants, did not develop autoantibodies or signs of EAMG. This highly reproducible new model of EAMG induced by a recombinant human autoantigen should be valuable for testing Ag-specific immunotherapeutic strategies that might be applicable to treating acquired myasthenia gravis in humans.

  16. Radioiodine in the environment

    International Nuclear Information System (INIS)

    Grauby, A.; Saas, A.

    1979-01-01

    An experimental procedure was developed in order to follow the pathways of radioiodine, 129 I especially, through the various compartments of nuclear plant environment: atmosphere, soil, plants, precipitations, ... A partial evaluation of the results is made. Data are supplied on iodine transfer into soil, iodine distribution one year later, release of iodine at the level of soil and plants. The intake of various forms of iodine in a soil and the evaluation of availability to plants by chemical tests showed that radioactive iodine ( 125 I) reached equilibrium with stable iodine after 120 - 140 days of incubation. During this time, there was a large amount of iodine exchangeable against a KI solution N/10. Equilibrium was reached progressively by iodine transfer on soil organic molecules. According to the origin of contamination, the transfer was more or less fast and differences were especially noticed in volatile iodine release and on chemical mobility. During intake, potential hazard of soil, groundwater, plant and atmospheric contaminations was greater. Iodine-129 is presently one of the chief contributors to air pollution around nuclear installations because of its radioactive half-life (1.72 x 10 7 yr) rather than on account of the amounts released. Just as with iodine-131, it is released into the atmosphere under many chemical forms. However, 129 I reaches man through several food chains and not only through the air-grass-cow-milk food chain. The values of its deposition rate on soil and plants are fairly dispersed because of its changing reactivity due to its chemical instability. On account of this chemical complexity, the transfer of iodine along the air-soil-plant system was studied using 125 I. (author)

  17. Production of radioiodinated prosthetic group for indirect protein labeling

    International Nuclear Information System (INIS)

    Santos, Josefina da Silva

    2001-01-01

    Monoclonal antibodies and their fragments and, more recently, radiolabeled peptides have been extensively studied in order to develop radiopharmaceuticals for diagnostic and therapy in Nuclear Medicine. The radioiodination of proteins can be done by a direct method, with radioiodine being incorporated in to a tyrosine residue of the protein by electrophilic substitution. The main problem in the use of radioiodinated proteins, is that they are often dehalogenated in vivo by the action of specific enzymes, probably because of the structural similarity between iodophenyl groups and thyroid hormones. Several protein radioiodination methods have been developed in order to minimize this in vivo dehalogenation using prosthetic groups for indirect labeling. In this case, the radioiodine is first incorporated in to the prosthetic group that is subsequently attached to a terminal amino group or to a ε-amino group of lysine residue. The aim of this work is to obtain a radioiodinated prosthetic group for indirect labeling of proteins. The prosthetic group selected was the N-succinimidyl-4-radioiodine benzoate (SIB), obtained by the iodination of the p-bromobenzoic acid followed by the reaction with TSTU (0-(N-succinimidyl)-N,N,N',N'-tetramethyl uronium tetrafluoroborate) The results of these studies showed that the p-radio iodobenzoic acid was obtained with a radiochemical purity greater than 92% and a labeling yield of about 65%. Some reaction parameters were studied like temperature, time and Cu Cl mass (cataliser). The SIB was quantitatively obtained from p-radio iodobenzoic acid, using basic medium and after removing the water from the reaction using an nitrogen stream. The kinetic of this reaction is very fast with complete consumption of the p-radioiodebenzoic acid after 5 minutes. The coupling of the SIB prosthetic group to the protein was studied using Human Immunoglobulin (IgG) as a protein model. In a comparative way, the same protein was used on direct labeling

  18. Characterization of DNA binding, transcriptional activation, and regulated nuclear association of recombinant human NFATp

    Directory of Open Access Journals (Sweden)

    Seto Anita G

    2000-11-01

    Full Text Available Abstract Background NFATp is one member of a family of transcriptional activators whose nuclear accumulation and hence transcriptional activity is regulated in mammalian cells. Human NFATp exists as a phosphoprotein in the cytoplasm of naive T cells. Upon antigen stimulation, NFATp is dephosphorylated, accumulates in nuclei, and functions to regulate transcription of genes including those encoding cytokines. While the properties of the DNA binding domain of NFATp have been investigated in detail, biochemical studies of the transcriptional activation and regulated association with nuclei have remained unexplored because of a lack of full length, purified recombinant NFATp. Results We developed methods for expressing and purifying full length recombinant human NFATp that has all of the properties known to be associated with native NFATp. The recombinant NFATp binds DNA on its own and cooperatively with AP-1 proteins, activates transcription in vitro, is phosphorylated, can be dephosphorylated by calcineurin, and exhibits regulated association with nuclei in vitro. Importantly, activation by recombinant NFATp in a reconstituted transcription system required regions of the protein outside of the central DNA binding domain. Conclusions We conclude that NFATp is a bona fide transcriptional activator. Moreover, the reagents and methods that we developed will facilitate future studies on the mechanisms of transcriptional activation and nuclear accumulation by NFATp, a member of an important family of transcriptional regulatory proteins.

  19. Characterization of recombinant human erythropoietin produced in Chinese hamster ovary cells

    International Nuclear Information System (INIS)

    Davis, J.M.; Arakawa, T.; Strickland, T.W.; Yphantis, D.A.

    1987-01-01

    Physicochemical properties of recombinant human erythropoietin were examined. This protein, produced in Chinese hamster ovary cells, showed a conformation apparently identical with the natural product isolated from human urine when examined by circular dichroism, UV absorbance, and fluorescence spectroscopy. Sedimentation equilibrium experiments showed the recombinant erythropoietin preparation to be essentially a single macromolecular component with a molecular weight of 30,400 and a carbohydrate content of 39%. The Stokes radius of recombinant erythropoietin was estimated to be 32 A from gel filtration, much larger than the 20-A radius calculated for a sphere of the observed molecular weight. This difference may be ascribed to the extensive glycosylation. The fluorescence and phosphorescence spectra showed that the luminescent tryptophan(s) is (are) solvent-exposed and can be quenched by I - and acrylamide but not by Cs + . On acid titration, the recombinant erythropoietin showed a conformational transition with a midpoint of pH 4.1. This suggests that the net charges on the protein moiety rather than on the whole molecule play a role in protein structure stability

  20. A novel multi-epitope recombined protein for diagnosis of human brucellosis.

    Science.gov (United States)

    Yin, Dehui; Li, Li; Song, Xiuling; Li, Han; Wang, Juan; Ju, Wen; Qu, Xiaofeng; Song, Dandan; Liu, Yushen; Meng, Xiangjun; Cao, Hongqian; Song, Weiyi; Meng, Rizeng; Liu, Jinhua; Li, Juan; Xu, Kun

    2016-05-21

    In epidemic regions of the world, brucellosis is a reemerging zoonosis with minimal mortality but is a serious public hygiene problem. Currently, there are various methods for brucellosis diagnosis, however few of them are available to be used to diagnose, especially for serious cross-reaction with other bacteria. To overcome this disadvantage, we explored a novel multi-epitope recombinant protein as human brucellosis diagnostic antigen. We established an indirect enzyme-linked immunosorbent assay (ELISA) based on this recombinant protein. 248 sera obtained from three different groups including patients with brucellosis (146 samples), non-brucellosis patients (82 samples), and healthy individuals (20 samples) were tested by indirect ELISA. To evaluate the assay, a receiver-operating characteristic (ROC) analysis and immunoblotting were carried out using these characterized serum samples. For this test, the area under the ROC curve was 0.9409 (95 % confidence interval, 0.9108 to 0.9709), and a sensitivity of 88.89 % and a specificity of 85.54 % was given with a cutoff value of 0.3865 from this ROC analysis. The Western blot results indicate that it is feasible to differentiate human brucellosis and non-brucellosis with the newly established method based on this recombinant protein. Our results obtained high diagnostic accuracy of the ELISA assay which encourage the use of this novel recombinant protein as diagnostic antigen to implement serological diagnosis of brucellosis.

  1. Recombinant expression of homodimeric 660 kDa human thyroglobulin in soybean seeds: an alternative source of human thyroglobulin.

    Science.gov (United States)

    Powell, Rebecca; Hudson, Laura C; Lambirth, Kevin C; Luth, Diane; Wang, Kan; Bost, Kenneth L; Piller, Kenneth J

    2011-07-01

    Soybean seeds possess many qualities that make them ideal targets for the production of recombinant proteins. However, one quality often overlooked is their ability to stockpile large amounts of complex storage proteins. Because of this characteristic, we hypothesized that soybean seeds would support recombinant expression of large and complex proteins that are currently difficult or impossible to express using traditional plant and non-plant-based host systems. To test this hypothesis, we transformed soybeans with a synthetic gene encoding human thyroglobulin (hTG)-a 660 kDa homodimeric protein that is widely used in the diagnostic industry for screening and detection of thyroid disease. In the absence of a recombinant system that can produce recombinant hTG, research and diagnostic grade hTG continues to be purified from cadaver and surgically removed thyroid tissue. These less-than-ideal tissue sources lack uniform glycosylation and iodination and therefore introduce variability when purified hTG is used in sensitive ELISA screens. In this study, we report the successful expression of recombinant hTG in soybean seeds. Authenticity of the soy-derived protein was demonstrated using commercial ELISA kits developed specifically for the detection of hTG in patient sera. Western analyses and gel filtration chromatography demonstrated that recombinant hTG and thyroid-purified hTG are biologically similar with respect to size, mass, charge and subunit interaction. The recombinant protein was stable over three generations and accumulated to ~1.5% of total soluble seed protein. These results support our hypothesis that soybeans represent a practical alternative to traditional host systems for the expression of large and complex proteins.

  2. Analysis of human reticulocyte genes reveals altered erythropoiesis: potential use to detect recombinant human erythropoietin doping.

    Science.gov (United States)

    Varlet-Marie, Emmanuelle; Audran, Michel; Lejeune, Mireille; Bonafoux, Béatrice; Sicart, Marie-Therese; Marti, Jacques; Piquemal, David; Commes, Thérèse

    2004-08-01

    Enhancement of oxygen delivery to tissues is associated with improved sporting performance. One way of enhancing oxygen delivery is to take recombinant human erythropoietin (rHuEpo), which is an unethical and potentially dangerous practice. However, detection of the use of rHuEpo remains difficult in situations such as: i) several days after the end of treatment ii) when a treatment with low doses is conducted iii) if the rHuEpo effect is increased by other substances. In an attempt to detect rHuEpo abuse, we selected erythroid gene markers from a SAGE library and analyzed the effects of rHuEpo administration on expression of the HBB, FTL and OAZ genes. Ten athletes were assigned to the rHuEpo or placebo group. The rHuEpo group received subcutaneous injections of rHuEpo (50 UI/kg three times a week, 4 weeks; 20 UI/kg three times a week, 2 weeks). HBB, FTL and OAZ gene profiles were monitored by real time-polymerase chain reaction (PCR) quantification during and for 3 weeks after drug administration. The global analysis of these targeted genes detected in whole blood samples showed a characteristic profile of subjects misusing rHuEpo with a increase above the threshold levels. The individual analysis of OAZ mRNA seemed indicative of rHuEpo treatment. The performance-enhancing effect of rHuEpo treatment is greater than the duration of hematologic changes associated with rHuEpo misuse. Although direct electrophoretic methods to detect rHuEpo have been developed, recombinant isoforms of rHuEpo are not detectable some days after the last subcutaneous injection. To overcome these limitations indirect OFF models have been developed. Our data suggest that, in the near future, it will be possible to consolidate results achievable with the OFF models by analyzing selected erythroid gene markers as a supplement to indirect methods.

  3. Complete genome sequence analysis of novel human bocavirus reveals genetic recombination between human bocavirus 2 and human bocavirus 4.

    Science.gov (United States)

    Khamrin, Pattara; Okitsu, Shoko; Ushijima, Hiroshi; Maneekarn, Niwat

    2013-07-01

    Epidemiological surveillance of human bocavirus (HBoV) was conducted on fecal specimens collected from hospitalized children with diarrhea in Chiang Mai, Thailand in 2011. By partial sequence analysis of VP1 gene, an unusual strain of HBoV (CMH-S011-11), was initially identified as HBoV4. The complete genome sequence of CMH-S011-11 was performed and analyzed further to clarify whether it was a recombinant strain or a new HBoV variant. Analysis of complete genome sequence revealed that the coding sequence starting from NS1, NP1 to VP1/VP2 was 4795 nucleotides long. Interestingly, the nucleotide sequence of NS1 gene of CMH-S011-11 was most closely related to the HBoV2 reference strains detected in Pakistan, which contradicted to the initial genotyping result of the partial VP1 region in the previous study. In addition, comparison of NP1 nucleotide sequence of CMH-S011-11 with those of other HBoV1-4 reference strains also revealed a high level of sequence identity with HBoV2. On the other hand, nucleotide sequence of VP1/VP2 gene of CMH-S011-11 was most closely related to those of HBoV4 reference strains detected in Nigeria. The overall full-length sequence analysis revealed that this CMH-S011-11 was grouped within HBoV4 species, but located in a separate branch from other HBoV4 prototype strains. Recombination analysis revealed that CMH-S011-11 was the result of recombination between HBoV2 and HBoV4 strains with the break point located near the start codon of VP2. Copyright © 2013 Elsevier B.V. All rights reserved.

  4. Radioiodine therapy and subsequent pregnancy

    International Nuclear Information System (INIS)

    Brandao, Carmen Dolores G.; Miranda, Angelica E.; Corres, Nilson Duarte; Sieiro Neto, Lino; Corbo, Rossana; Vaisman, Mario

    2007-01-01

    Objectives: To evaluate abortion and fetal congenital anomaly rates in women previously submitted to radioiodine therapy for differentiated thyroid carcinoma. Study design: A case-control study of 108 pregnant women, 48 cases whose pregnancies were evaluated after they had undergone radioiodine therapy for differentiated thyroid carcinoma, and the control group consisted of 60 healthy pregnant women. Results: Of a total of 66 pregnancies, 14 conceived within the first year, 51 one or more years after the last administration of 131 I, the medical record of one patient was not available. The interval between the last radioiodine therapy administration and conception ranged from 1 month to 10 years. There were a total of 4 miscarriages, 2 of them for unknown reasons. There was one case of congenital anomaly and two preterms birth. Nine women presented the following pregnancy events: placental insufficiency, hypertensive crisis, placental detachment, risk of miscarriage, preterm labour and four miscarriages. No statistical difference was observed between the studied and control groups. Conclusion: Radioiodine was followed by no significant increase in untoward effects in neither the pregnancy nor the offspring. (author)

  5. Does recombinant human Epo increase exercise capacity by means other than augmenting oxygen transport?

    DEFF Research Database (Denmark)

    Lundby, C; Robach, P; Boushel, R

    2008-01-01

    This study was performed to test the hypothesis that administration of recombinant human erythropoietin (rHuEpo) in humans increases maximal oxygen consumption by augmenting the maximal oxygen carrying capacity of blood. Systemic and leg oxygen delivery and oxygen uptake were studied during...... before rHuEpo treatment). Blood buffer capacity remained unaffected by rHuEpo treatment and hemodilution. The augmented hematocrit did not compromise peak cardiac output. In summary, in healthy humans, rHuEpo increases maximal oxygen consumption due to augmented systemic and muscular peak oxygen delivery....

  6. Protection of Non-Human Primates against Rabies with an Adenovirus Recombinant Vaccine

    Science.gov (United States)

    Xiang, Z.Q.; Greenberg, L.; Ertl, H. C.; Rupprecht, C.E.

    2014-01-01

    Rabies remains a major neglected global zoonosis. New vaccine strategies are needed for human rabies prophylaxis. A single intramuscular immunization with a moderate dose of an experimental chimpanzee adenovirus (Ad) vector serotype SAd-V24, also termed AdC68, expressing the rabies virus glycoprotein, resulted in sustained titers of rabies virus neutralizing antibodies and protection against a lethal rabies virus challenge infection in a non-human primate model. Taken together, these data demonstrate the safety, immunogenicity, and efficacy of the recombinant Ad-rabies vector for further consideration in human clinical trials. PMID:24503087

  7. LDSplitDB: a database for studies of meiotic recombination hotspots in MHC using human genomic data.

    Science.gov (United States)

    Guo, Jing; Chen, Hao; Yang, Peng; Lee, Yew Ti; Wu, Min; Przytycka, Teresa M; Kwoh, Chee Keong; Zheng, Jie

    2018-04-20

    Meiotic recombination happens during the process of meiosis when chromosomes inherited from two parents exchange genetic materials to generate chromosomes in the gamete cells. The recombination events tend to occur in narrow genomic regions called recombination hotspots. Its dysregulation could lead to serious human diseases such as birth defects. Although the regulatory mechanism of recombination events is still unclear, DNA sequence polymorphisms have been found to play crucial roles in the regulation of recombination hotspots. To facilitate the studies of the underlying mechanism, we developed a database named LDSplitDB which provides an integrative and interactive data mining and visualization platform for the genome-wide association studies of recombination hotspots. It contains the pre-computed association maps of the major histocompatibility complex (MHC) region in the 1000 Genomes Project and the HapMap Phase III datasets, and a genome-scale study of the European population from the HapMap Phase II dataset. Besides the recombination profiles, related data of genes, SNPs and different types of epigenetic modifications, which could be associated with meiotic recombination, are provided for comprehensive analysis. To meet the computational requirement of the rapidly increasing population genomics data, we prepared a lookup table of 400 haplotypes for recombination rate estimation using the well-known LDhat algorithm which includes all possible two-locus haplotype configurations. To the best of our knowledge, LDSplitDB is the first large-scale database for the association analysis of human recombination hotspots with DNA sequence polymorphisms. It provides valuable resources for the discovery of the mechanism of meiotic recombination hotspots. The information about MHC in this database could help understand the roles of recombination in human immune system. DATABASE URL: http://histone.scse.ntu.edu.sg/LDSplitDB.

  8. Analysis of clinical factors for the determination of optimal serum level of thyrotropin after recombinant human thyroid-stimulating hormone administration

    International Nuclear Information System (INIS)

    Son, Seung Hyun; Lee, Sang Woo; Jung, Ji Hoon; Kim, Choon Young; Kim, Do Hoon; Jeong, Shin Young; Ahn, Byeong Cheol; Lee, Jae Tae

    2015-01-01

    To determine the optimal levels of thyroid-stimulating hormone (TSH) levels after administration of recombinant human TSH (rhTSH) to patients with differentiated thyroid cancer (DTC), we have analyzed the clinical parameters that affected the degree of the increase in serum levels of TSH. We retrospectively analyzed 276 patients with differentiated thyroid cancer (DTC), post-thyroidectomy and remnant ablation. Pearson’s correlation coefficient test was used to evaluate the correlation between serum levels of TSH after rhTSH stimulation and various clinical factors, including age, sex, height, weight, body mass index (BMI), body surface area (BSA), serum blood urea nitrogen, creatinine, and estimated glomerular filtration rate (GFR). Linear regression analysis was used to determine the predictors of the degree of increase in serum TSH level after rhTSH stimulation. After the rhTSH injections, all subjects achieved TSH levels of >30 μU/mL, with a mean of 203.8 ± 83.4 μU/mL. On univariate analysis, age (r = 0.255) and serum creatinine (r = 0.169) level were positive predictors for higher levels of serum TSH after rhTSH stimulation, while weight (r = –0.239), BMI (r = –0.223), BSA (r = –0.217), and estimated GFR (r = –0.199) were negative predictors. Multiple linear regression analysis revealed that serum creatinine was the most powerful independent predictor for serum levels of TSH, followed by age, BSA, and BMI. An increment in serum TSH after rhTSH stimulation was significantly affected by age, BSA, BMI, and creatinine, with creatinine being the most powerful predictor. By understanding the difference in the increased levels of TSH in various subjects, their dose of rhTSH can be adjusted during scheduling for radioiodine ablation, or during follow-up (recurrence surveillance) after surgery and ablation

  9. Analysis of clinical factors for the determination of optimal serum level of thyrotropin after recombinant human thyroid-stimulating hormone administration

    Energy Technology Data Exchange (ETDEWEB)

    Son, Seung Hyun; Lee, Sang Woo; Jung, Ji Hoon; Kim, Choon Young; Kim, Do Hoon; Jeong, Shin Young; Ahn, Byeong Cheol; Lee, Jae Tae [Dept. of Nuclear Medicine, Kyungpook National University Medical Center and School of Medicine, Daegu (Korea, Republic of)

    2015-12-15

    To determine the optimal levels of thyroid-stimulating hormone (TSH) levels after administration of recombinant human TSH (rhTSH) to patients with differentiated thyroid cancer (DTC), we have analyzed the clinical parameters that affected the degree of the increase in serum levels of TSH. We retrospectively analyzed 276 patients with differentiated thyroid cancer (DTC), post-thyroidectomy and remnant ablation. Pearson’s correlation coefficient test was used to evaluate the correlation between serum levels of TSH after rhTSH stimulation and various clinical factors, including age, sex, height, weight, body mass index (BMI), body surface area (BSA), serum blood urea nitrogen, creatinine, and estimated glomerular filtration rate (GFR). Linear regression analysis was used to determine the predictors of the degree of increase in serum TSH level after rhTSH stimulation. After the rhTSH injections, all subjects achieved TSH levels of >30 μU/mL, with a mean of 203.8 ± 83.4 μU/mL. On univariate analysis, age (r = 0.255) and serum creatinine (r = 0.169) level were positive predictors for higher levels of serum TSH after rhTSH stimulation, while weight (r = –0.239), BMI (r = –0.223), BSA (r = –0.217), and estimated GFR (r = –0.199) were negative predictors. Multiple linear regression analysis revealed that serum creatinine was the most powerful independent predictor for serum levels of TSH, followed by age, BSA, and BMI. An increment in serum TSH after rhTSH stimulation was significantly affected by age, BSA, BMI, and creatinine, with creatinine being the most powerful predictor. By understanding the difference in the increased levels of TSH in various subjects, their dose of rhTSH can be adjusted during scheduling for radioiodine ablation, or during follow-up (recurrence surveillance) after surgery and ablation.

  10. Endostar, a recombined humanized endostatin, enhances the radioresponse for human nasopharyngeal carcinoma and human lung adenocarcinoma xenografts in mice

    International Nuclear Information System (INIS)

    Wen Qinglian; Meng Maobin; Tu Lingli; Jia Li; Zhou Lin; Xu Yong; Lu You; Yang Bo

    2009-01-01

    The purpose of this paper is to determine the efficacy of combining radiation therapy with endostar, a recombined humanized endostatin, in human nasopharyngeal carcinoma and human lung adenocarcinoma xenografts. Tumor xenografts were established in the hind limb of male athymic nude mice (BALB/c-nu) by subcutaneous transplantation. The tumor-bearing mice were assigned into four treatment groups: sham therapy (control), endostar (20 mg/kg, once daily for 10 days), radiation therapy (6 Gray per day to 30 Gray, once a day for 1 week), and endostar plus radiation therapy (combination). The experiment was repeated and mice were killed at days 3, 6, and 10 after initiation therapy, and the tumor tissues and blood samples were collected to analyze the kinetics of antitumor, antiangiogenesis, and antivascularization responses of different therapies. In human nasopharyngeal carcinoma and human lung adenocarcinoma xenografts, endostar significantly enhanced the effects of tumor growth inhibition, endothelial cell and tumor cell apoptosis induction, and improved tumor cell hypoxia of radiation therapy. Histological analyses demonstrated that endostar plus radiation also induced a significant reduction in microvascular density, microvascular area, and vascular endothelial growth factor and matrix metalloproteinase-2 expression compared with radiation and endostar alone respectively. We concluded that endostar significantly sensitized the function of radiation in antitumor and antiangiogenesis in human nasopharyngeal carcinoma and human lung adenocarcinoma xenografts by increasing the apoptosis of the endothelial cell and tumor cell, improving the hypoxia of the tumor cell, and changing the proangiogenic factors. These data provided a rational basis for clinical practice of this multimodality therapy. (author)

  11. [Recombinant human gapM1 expressed in Pichia pastoris and its anti-diabetic effect].

    Science.gov (United States)

    Mei, Xiang; Du, Renqian; Li, Xi; Huang, Haiyan; Yu, Min; Tang, Qiqun

    2009-08-01

    Adiponectin is an adipokine predominantly synthesized and secreted by adipocytes in the white adipose tissue, and it can lower the blood glucose level and increase free fatty acid oxidation. In the current study, we developed the globular domain of adiponectin (gapM1) to treat type II diabetes. In both flask and fermentor, we cultivated Pichia pastoris expressing recombinant gapM1 and established the purification procedure by using gel filtration and anion exchange chromatography. To evaluate the biological activity of recombinant gapM1, we used rat type II diabetes model fed high-fat diet in combination with low-dose STZ (Streptozocin) induction. We purified 200 mg gapM1 with purity of 96% from 10 liters of supernatant. The recombinant gapM1 significantly lowered blood glucose (34.2%), serum triglyceride (79.6%) and total cholesterol (62.1%) in type II diabetes induced rat. Therefore, the recombinant human gapM1 is successfully expressed in Pichia pastoris and effectively treated type II diabetes in rat models.

  12. Expression of human DNA polymerase β in Escherichia coli and characterization of the recombinant enzyme

    International Nuclear Information System (INIS)

    Abbotts, J.; SenGupta, D.N.; Zmudzka, B.; Widen, S.G.; Notario, V.; Wilson, S.H.

    1988-01-01

    The coding region of a human β-polymerase cDNA, predicting a 335 amino acid protein, was subcloned in the Escherichia coli expression plasmid pRC23. After induction of transformed cells, the crude soluble extract was found to contain a new protein immunoreactive with β-polymerase antibody and corresponding in size to the protein deduced from the cDNA. This protein was purified in a yield of 1-2 mg/50 g of cells. The recombinant protein had about the same DNA polymerase specific activity as β-polymerase purified from mammalian tissues, and template-primer specificity and immunological properties of the recombinant polymerase were similar to those of natural β-polymerases. The purified enzyme was free of nuclease activity. The authors studied detailed catalytic properties of the recombinant β-polymerase using defined template-primer systems. The results indicate that this β-polymerase is essentially identical with natural β-polymerases. The recombinant enzyme is distributive in mode of synthesis and is capable of detecting changes in the integrity of the single-stranded template, such as methylated bases and a double-stranded region. The enzyme recognizes a template region four to seven bases downstream of the primer 3' end and utilizes alternative primers if this downstream template region is double stranded. The enzyme is unable to synthesize past methylated bases N 3 -methyl-dT or O 6 -methyl-dG

  13. Genetic recombination within the human T-cell receptor α-chain gene complex

    International Nuclear Information System (INIS)

    Robinson, M.A.; Kindt, T.J.

    1987-01-01

    Genetic analyses of the human T-cell receptor (TCR) α-chain genes indicate that recombination events may occur frequently within this gene complex. Examination of the inheritance of restriction fragment length polymorphisms (RFLP) detected by using probes for constant or variable region gene segments made it possible to assign TCRα haplotypes to the 16 parents and 43 offspring of eight families studied. A total of six RFLP, three for the constant region and three for variable region segments, were examined in the present studies. Most enzyme and probe combinations tested revealed no polymorphism and those finally selected for the study showed limited polymorphism in that only two or, in one case, three allelic forms of the gene were seen. In spite of limited variability at this level, extensive heterogeneity was observed for the combinations of markers present in haplotypes, suggesting that frequent recombination events have occurred. Most strikingly, multiple combinations of RFLP occurring in close proximity of the TCRα constant region gene were observed in this study. A high recombination frequency for the TCRα gene complex is further supported by the observation that two children, one in each of two families, inherited recombinant TCRα haplotypes

  14. Recombinant human parvovirus B19 vectors: erythroid cell-specific delivery and expression of transduced genes.

    Science.gov (United States)

    Ponnazhagan, S; Weigel, K A; Raikwar, S P; Mukherjee, P; Yoder, M C; Srivastava, A

    1998-06-01

    A novel packaging strategy combining the salient features of two human parvoviruses, namely the pathogenic parvovirus B19 and the nonpathogenic adeno-associated virus type 2 (AAV), was developed to achieve erythroid cell-specific delivery as well as expression of the transduced gene. The development of such a chimeric vector system was accomplished by packaging heterologous DNA sequences cloned within the inverted terminal repeats of AAV and subsequently packaging the DNA inside the capsid structure of B19 virus. Recombinant B19 virus particles were assembled, as evidenced by electron microscopy as well as DNA slot blot analyses. The hybrid vector failed to transduce nonerythroid human cells, such as 293 cells, as expected. However, MB-02 cells, a human megakaryocytic leukemia cell line which can be infected by B19 virus following erythroid differentiation with erythropoietin (N. C. Munshi, S. Z. Zhou, M. J. Woody, D. A. Morgan, and A. Srivastava, J. Virol. 67:562-566, 1993) but lacks the putative receptor for AAV (S. Ponnazhagan, X.-S. Wang, M. J. Woody, F. Luo, L. Y. Kang, M. L. Nallari, N. C. Munshi, S. Z. Zhou, and A. Srivastava, J. Gen. Virol. 77:1111-1122, 1996), were readily transduced by this vector. The hybrid vector was also found to specifically target the erythroid population in primary human bone marrow cells as well as more immature hematopoietic progenitor cells following erythroid differentiation, as evidenced by selective expression of the transduced gene in these target cells. Preincubation with anticapsid antibodies against B19 virus, but not anticapsid antibodies against AAV, inhibited transduction of primary human erythroid cells. The efficiency of transduction of primary human erythroid cells by the recombinant B19 virus vector was significantly higher than that by the recombinant AAV vector. Further development of the AAV-B19 virus hybrid vector system should prove beneficial in gene therapy protocols aimed at the correction of inherited and

  15. Recombinant Human Parvovirus B19 Vectors: Erythroid Cell-Specific Delivery and Expression of Transduced Genes

    Science.gov (United States)

    Ponnazhagan, Selvarangan; Weigel, Kirsten A.; Raikwar, Sudhanshu P.; Mukherjee, Pinku; Yoder, Mervin C.; Srivastava, Arun

    1998-01-01

    A novel packaging strategy combining the salient features of two human parvoviruses, namely the pathogenic parvovirus B19 and the nonpathogenic adeno-associated virus type 2 (AAV), was developed to achieve erythroid cell-specific delivery as well as expression of the transduced gene. The development of such a chimeric vector system was accomplished by packaging heterologous DNA sequences cloned within the inverted terminal repeats of AAV and subsequently packaging the DNA inside the capsid structure of B19 virus. Recombinant B19 virus particles were assembled, as evidenced by electron microscopy as well as DNA slot blot analyses. The hybrid vector failed to transduce nonerythroid human cells, such as 293 cells, as expected. However, MB-02 cells, a human megakaryocytic leukemia cell line which can be infected by B19 virus following erythroid differentiation with erythropoietin (N. C. Munshi, S. Z. Zhou, M. J. Woody, D. A. Morgan, and A. Srivastava, J. Virol. 67:562–566, 1993) but lacks the putative receptor for AAV (S. Ponnazhagan, X.-S. Wang, M. J. Woody, F. Luo, L. Y. Kang, M. L. Nallari, N. C. Munshi, S. Z. Zhou, and A. Srivastava, J. Gen. Virol. 77:1111–1122, 1996), were readily transduced by this vector. The hybrid vector was also found to specifically target the erythroid population in primary human bone marrow cells as well as more immature hematopoietic progenitor cells following erythroid differentiation, as evidenced by selective expression of the transduced gene in these target cells. Preincubation with anticapsid antibodies against B19 virus, but not anticapsid antibodies against AAV, inhibited transduction of primary human erythroid cells. The efficiency of transduction of primary human erythroid cells by the recombinant B19 virus vector was significantly higher than that by the recombinant AAV vector. Further development of the AAV-B19 virus hybrid vector system should prove beneficial in gene therapy protocols aimed at the correction of inherited

  16. Recombinant Human Acidic Fibroblast Growth Factor (aFGF) Expressed in Nicotiana benthamiana Potentially Inhibits Skin Photoaging.

    Science.gov (United States)

    Ha, Jang-Ho; Kim, Ha-Neul; Moon, Ki-Beom; Jeon, Jae-Heung; Jung, Dai-Hyun; Kim, Su-Jung; Mason, Hugh S; Shin, Seo-Yeon; Kim, Hyun-Soon; Park, Kyung-Mok

    2017-07-01

    Responding to the need for recombinant acidic fibroblast growth factor in the pharmaceutical and cosmetic industries, we established a scalable expression system for recombinant human aFGF using transient and a DNA replicon vector expression in Nicotiana benthamiana . Recombinant human-acidic fibroblast growth factor was recovered following Agrobacterium infiltration of N. benthamiana . The optimal time point at which to harvest recombinant human acidic fibroblast growth factor expressing leaves was found to be 4 days post-infiltration, before necrosis was evident. Commassie-stained SDS-PAGE gels of His-tag column eluates, concentrated using a 10 000 molecular weight cut-off column, showed an intense band at the expected molecular weight for recombinant human acidic fibroblast growth factor. An immunoblot confirmed that this band was recombinant human acidic fibroblast growth factor. Up to 10 µg recombinant human-acidic fibroblast growth factor/g of fresh leaves were achieved by a simple affinity purification protocol using protein extract from the leaves of agroinfiltrated N. benthamiana . The purified recombinant human acidic fibroblast growth factor improved the survival rate of UVB-irradiated HaCaT and CCD-986sk cells approximately 89 and 81 %, respectively. N. benthamiana -derived recombinant human acidic fibroblast growth factor showed similar effects on skin cell proliferation and UVB protection compared to those of Escherichia coli -derived recombinant human acidic fibroblast growth factor. Additionally, N. benthamiana- derived recombinant human acidic fibroblast growth factor increased type 1 procollagen synthesis up to 30 % as well as reduced UVB-induced intracellular reactive oxygen species generation in fibroblast (CCD-986sk) cells.UVB is a well-known factor that causes various types of skin damage and premature aging. Therefore, the present study demonstrated that N. benthamiana -derived recombinant human acidic fibroblast growth factor

  17. Turbulent transfer characteristics of radioiodine effluents from air to grass

    Energy Technology Data Exchange (ETDEWEB)

    Markee, E. H. [ARFRO, Environmental Science Services Administration, Idaho Falls, Idaho (United States)

    1967-07-01

    A total of 20 controlled field releases of radioiodine have been performed at the National Reactor Testing Station in Idaho as a portion of a program to study the transmission of gaseous radioiodine through the air-vegetation-cow-milk-human chain. Most of the releases were conducted over typical pasture grasses during different wind and stability conditions. Radioiodine adherence to grass and carbon plates was measured during most of the tests. Vertical air concentration profiles and turbulence parameters were measured to determine flux characteristics. Analysis of the data reveals the complex interdisciplinary nature of transfer of radioiodine from air to a natural surface. The data are in reasonable agreement with the deposition models of Sheppard and Chamberlain when corrections for the physical and biological receptiveness of the grass and grass density are made. The average ratios of momentum to mass flux were found to be 0.9 in stable conditions and 1.4 in unstable conditions. These ratios demonstrate the effect on mass flux in the lowest 4m by a surface that acts as a partial sink for gaseous effluents. This series of releases indicates the need for further research on the biological receptiveness of grass and turbulent transfer within a grass canopy. (author)

  18. A New Technology Developed to Remove Aqueous Radioiodine

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Seung Yeop; Baik, Min Hoon [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2015-10-15

    Radioiodine is considered as a critical radionuclide by the International Commission on Radiological Protection(ICRP) in terms of the radiation and radiotoxicity risks to nature and human health. In addition, there were reports about great increases of thyroid cancer in children in Belarus from 1990. In natural environments, there were several reports for the unusual increase of radioactive iodine ratios in most environmental samples (i.e. soils, vegetables, and seafoods). Iodine usually exists as iodide, iodate, and organic iodine in the environment. Iodide is considered as the most difficult form among the iodine species to treat by the conventional adsorption-based technology. This is why urgent new technology is needed to suppress such a progressive accumulation of radioiodine in natural environments. The conventional methods based on the anion-exchange and adsorption techniques have been difficult to manage the growing release of radioiodine for decades. Here we suggest a new solution that uses a biostimulant way to isolate radioiodine. We discovered that natural SRB can play an important intermediary role to get anionic iodide and cationic copper combined strongly as perfect counterions by enzymatic Cu-reduction.

  19. Synthesis, chemical and biological quality control of radioiodinated peptides

    International Nuclear Information System (INIS)

    Rafii, H.; Khalaj, A.; Beiki, D.; Motameidi, F.; Maloobi, M.; Karimian-dehghan, M.; Keshavarrzi, F.

    2002-01-01

    Iodinated compounds with I-131, 125 and 123 have been widely used for biochemical function studies. In conjunction with SPECT, [I-123] labelled proteins have various diagnostic and therapeutic applications in nuclear medicine. Preparation of some radioiodinated peptides with tyrosine and/or lysine groups on their main chain molecules can be carried out with both direct and indirect methods, but lack of these groups in molecule cause the molecule dose not lend itself for direct radioiodination. In this study, human IgG and Formyl-Methyl-Leucyl-Phenylalanine, FMLF, have been chosen as a model compounds for direct and indirect radioiodination respectively. Here, we will describe the labelling procedure of [I-125] IgG using chloramine-T as a suitable oxidant agent and [I-125 and I-131] FMLF by indirect method using ATE/SIB as a prosthetic group in multi-step reactions. The obtained results for chemical quality control of intermediate radioiodinated SIB by HPLC and two labelled IgG and FMLF will be also discussed. Biological results, biodistribution studies and SPECT scans on mice per-injected labelled FMLF show a low uptake of thyroid but a high at urine and bladder, perhaps because of low molecular weight of FMLF. In this case, it seems to be better to separate the reaction mixture of labelled FMLF by BPLC than Sephadex-G50 gel filtration. (Author)

  20. Radioiodination of vasoactive intestinal peptide (VIP)

    International Nuclear Information System (INIS)

    Wang, Y.; Wang, L.; Yin, D.

    2002-01-01

    In recent years, increasing biochemical and radiochemical research has been performed to develop radiolabelled peptides as specific ligands for tumour associated receptors. VIP, a 28-amino acid peptide containing two tyrosines and three lysines, has demonstrated that various tumour cells express significantly higher amounts of VIP-receptors and could be applied to the clinic diagnosis. For these purposes, radiohalogenation of VIP by direct and indirect method was studied. Direct labelling works well for radioiodine but is limited to dehalogenation of labelling products in vivo. Conjugate labelling methods including Boltonhunter and wood reagents were developed but introduction of such a molecule to peptides may lead to the decrease of biological activity in vivo. In order to resolve these problems, N-Succinimidyl-3-(tri-nbutylstannyl) benzoate (ATE) was elected for the radioiodination of VIP and already employed to radioiodination of IgG successfully. The in vitro stability and biological activity would be compared in these two methods. Vasoactive intestinal peptide (VIP) and human immunoglobulin (IgG) were radioiodinated by direct and indirect methods. Iodogen was employed in direct method and N-Succinimidyl-3-(tri-n-butylstannyl) benzoate (ATE) was applied as a prosthetic group in the conjugation labelling. The subject of our study was optimizing the radiohalogenation of IgG and VIP followed by separation and analysis of reaction products. The advantages and disadvantages were illustrated by comparing the in vitro stability and biological activity in these two methods. Na 123 I was prepared by nuclear reaction of 124 Te(p, 2n) 123 I using cyclone-30. More than 95% of radiochemical purity, more than 95% of radionuclide purity and about 100 mCi/mL of radioactivity concentration were obtained. ATE was supplied by Dr. Pozzi and radioiodinated with iodogen and 96% of labelling efficiency was obtained. The stability of radioactive S 125 IB kept well in dark at 4

  1. Metastatic Follicular Thyroid Carcinoma Secreting Thyroid Hormone and Radioiodine Avid without Stimulation: A Case Report and Literature Review

    Directory of Open Access Journals (Sweden)

    Syed A. Abid

    2014-01-01

    Full Text Available Introduction. This is an extremely rare case of a patient with metastatic follicular thyroid cancer who continued to produce thyroid hormone and was iodine scan positive without stimulation after thyroidectomy and radioiodine (I-131 therapy. Patient Findings. A 76-year-old Caucasian male was diagnosed with metastatic follicular thyroid carcinoma on lung nodule biopsy. Total thyroidectomy was performed and he was ablated with 160 mCi of I-131 after recombinant human thyrotropin (rhTSH stimulation. Whole body scan (WBS after treatment showed uptake in bilateral lungs, right sacrum, and pelvis. The thyroglobulin decreased from 2,063 to 965 four months after treatment but rapidly increased to 2,506 eleven months after I-131. Thyroid stimulating hormone (TSH remained suppressed and free T4 remained elevated after I-131 therapy without thyroid hormone supplementation. He was treated with an additional 209 mCi with WBS findings positive in lung and pelvis. Despite I-131, new metastatic lesions were noted in the left thyroid bed and large destructive lesion to the first cervical vertebrae four months after the second I-131 dose. Conclusions. This case is exceptional because of its rarity and also due to the dissociation between tumor differentiation and aggressiveness. The metastatic lesions continued to secrete thyroid hormone and remained radioiodine avid with rapid progression after I-131 therapy.

  2. Virulence evolution of the human pathogen Neisseria meningitidis by recombination in the core and accessory genome.

    Directory of Open Access Journals (Sweden)

    Biju Joseph

    Full Text Available BACKGROUND: Neisseria meningitidis is a naturally transformable, facultative pathogen colonizing the human nasopharynx. Here, we analyze on a genome-wide level the impact of recombination on gene-complement diversity and virulence evolution in N. meningitidis. We combined comparative genome hybridization using microarrays (mCGH and multilocus sequence typing (MLST of 29 meningococcal isolates with computational comparison of a subset of seven meningococcal genome sequences. PRINCIPAL FINDINGS: We found that lateral gene transfer of minimal mobile elements as well as prophages are major forces shaping meningococcal population structure. Extensive gene content comparison revealed novel associations of virulence with genetic elements besides the recently discovered meningococcal disease associated (MDA island. In particular, we identified an association of virulence with a recently described canonical genomic island termed IHT-E and a differential distribution of genes encoding RTX toxin- and two-partner secretion systems among hyperinvasive and non-hyperinvasive lineages. By computationally screening also the core genome for signs of recombination, we provided evidence that about 40% of the meningococcal core genes are affected by recombination primarily within metabolic genes as well as genes involved in DNA replication and repair. By comparison with the results of previous mCGH studies, our data indicated that genetic structuring as revealed by mCGH is stable over time and highly similar for isolates from different geographic origins. CONCLUSIONS: Recombination comprising lateral transfer of entire genes as well as homologous intragenic recombination has a profound impact on meningococcal population structure and genome composition. Our data support the hypothesis that meningococcal virulence is polygenic in nature and that differences in metabolism might contribute to virulence.

  3. Maxillary anterior ridge augmentation with recombinant human bone morphogenetic protein 2.

    Science.gov (United States)

    Edmunds, Ryan K; Mealey, Brian L; Mills, Michael P; Thoma, Daniel S; Schoolfield, John; Cochran, David L; Mellonig, Jim

    2014-01-01

    No human studies exist on the use of recombinant human bone morphogenetic protein 2 (rhBMP-2) on an absorbable collagen sponge (ACS) as a sole graft material for lateral ridge augmentation in large ridge defect sites. This series evaluates the treatment outcome of maxillary anterior lateral ridge augmentation with rhBMP-2/ACS. Twenty patients were treated with rhBMP-2/ACS and fixation screws for space maintenance. Cone beam volumetric tomography measurements were used to determine gain in ridge width, and a bone core biopsy was obtained. The mean horizontal ridge gain was 1.2 mm across sites, and every site gained width.

  4. INACTIVITY OF RECOMBINANT ELA2B PROVIDES A NEW EXAMPLE OF EVOLUTIONARY ELASTASE SILENCING IN HUMANS

    OpenAIRE

    Szepessy, Edit; Sahin-Tóth, Miklós

    2005-01-01

    BACKGROUND. The archetypal mammalian elastase (ELA1) is not expressed in the human pancreas, because evolutionary mutations suppressed transcription of the ELA1 gene. AIMS. In this study we tested the theory that the unique duplication of the ELA2 gene in humans might compensate for the loss of ELA1. METHODS. Recombinant ELA2A and ELA2B were expressed in Escherichia coli, and their activity was tested on Glt-Ala-Ala-Pro-Leu-p-nitroanilide, DQ elastin and bovine milk protein. RESULTS. Surprisi...

  5. Translational mixed-effects PKPD modelling of recombinant human growth hormone - from hypophysectomized rat to patients

    DEFF Research Database (Denmark)

    Thorsted, A; Thygesen, P; Agersø, H

    2016-01-01

    BACKGROUND AND PURPOSE: We aimed to develop a mechanistic mixed-effects pharmacokinetic (PK)-pharmacodynamic (PD) (PKPD) model for recombinant human growth hormone (rhGH) in hypophysectomized rats and to predict the human PKPD relationship. EXPERIMENTAL APPROACH: A non-linear mixed-effects model...... was developed from experimental PKPD studies of rhGH and effects of long-term treatment as measured by insulin-like growth factor 1 (IGF-1) and bodyweight gain in rats. Modelled parameter values were scaled to human values using the allometric approach with fixed exponents for PKs and unscaled for PDs...... s.c. administration was over predicted. After correction of the human s.c. absorption model, the induction model for IGF-1 well described the human PKPD data. CONCLUSIONS: A translational mechanistic PKPD model for rhGH was successfully developed from experimental rat data. The model links...

  6. New prospects in radioiodine production technologies and radioiodination

    Energy Technology Data Exchange (ETDEWEB)

    Miller, J [Magyar Tudomanyos Akademia, Budapest. Izotopintezete

    1984-01-01

    A new economical dry distillation procedure was developed to produce /sup 131/I or /sup 125/I from irradiated TeO/sub 2/ melt and xenon gas, respectively. The method can be applied to the thermodesorptive radioiodination of different types of water soluble or insoluble organic compounds: oleic acid, prostaglandin, insulin, ACTH, antipyrine. The labelling procedure is based on the thermolysis of the stable Pt(I) surface compound.

  7. Synthesis, in vitro binding, and tissue distribution of radioiodinated 2-[125I]N-(N-benzylpiperidin-4-yl)-2-iodo benzamide, 2-[125I]BP: a potential σ receptor marker for human prostate tumors

    International Nuclear Information System (INIS)

    John, Christy S.; Gulden, Mary E.; Li, Jinghua; Bowen, Wayne D.; McAfee, John G.; Thakur, Mathew L.

    1998-01-01

    The preclinical evaluation of a σ receptor-specific radiopharmaceutical that binds to human prostate tumor cells with a high affinity is described. We have synthesized and radioiodinated 2-[ 125 I]-N-(N-benzylpiperidin-4-yl)-2-iodobenzamide (2-[ 125 I]BP) that possesses high affinity for both σ-1 and σ-2 receptor subtypes that are expressed on a variety of tumor cells. 2-IBP was synthesized, purified and characterized by routine spectroscopic and analytical methods. Radioiodination was accomplished using an oxidative iododestannylation reaction in the presence of chloramine T in high yields (76%-93%) with a very high-specific activity (1700-1900 Ci/mmol). The in vitro competition binding studies of 2-[ 125 I]BP with various σ receptor ligands in LnCAP human prostate tumor cells showed a dose-dependent saturable binding. The inhibition constants (K i , nM) for binding of 2-[ 125 I]BP to human prostate tumor cells for 4-IBP, haloperidol and 2-IBP were 4.09, 6.34 and 1.6 nM, respectively. The clearance of 2-[ 125 I]BP, in Sprague-Dawley rats, was rapid from the blood pool, other normal tissues and the total body. Tissue distribution studies in nude mice bearing human prostate tumor (DU-145) also showed a fast clearance from normal organs. The tumor had the highest percentage of injected dose per gram (%ID/g) of all tissues at 4 h as well as 24 h (2.0 ± 0.05 and 0.147 ± 0.038 ID/g, respectively) postinjection. The in vivo receptor binding specificity was demonstrated using haloperidol (a known high-affinity σ receptor ligand). A significant decrease (>50%, p = 0.001) was observed in tumor concentration when haloperidol was used as a blocking agent. The high affinity of 2-[ 125 I]BP for σ receptor-binding sites, its fast in vivo clearance from normal organs and its high uptake and retention in tumor implies that 2-[ 123 I]BP or 2-[ 131 I]BP may be a promising tracer for noninvasive imaging of human prostate tumors

  8. Characterization of recombinant human lysosomal beta-hexosaminidases produced in the methylotrophic yeast Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Angela Johana Espejo Mojica

    2016-08-01

    Full Text Available β-hexosaminidases (Hex are dimeric enzymes involved in the lysosomal degradation of glycolipids and glycans. They are formed by α- and/or β-subunits encoded byHEXA and HEXB genes, respectively. Mutations in these genes lead to Tay Sachs or Sandhoff diseases, which are neurodegenerative disorders caused by the accumulation of non-degraded glycolipids. Although tissue-derived Hex have been widely characterized, limited information is available for recombinant β-hexosaminidases. In this study, human lysosomal recombinant Hex (rhHex-A, rhHex-B, and rhHex-S were produced in the methylotrophic yeast Pichia pastoris GS115. The highest specific enzyme activities were 13,124 for rhHexA; 12,779 for rhHex-B; and 14,606 U .mg-1 for rhHex-S. These results were 25- to 50-fold higher than those obtained from normal human leukocytes. Proteins were purified and characterized at different pH and temperature conditions. All proteins were stable at acidic pH, and at 4 °C and 37 °C. At 45 °C rhHex-S was completely inactivated, while rhHex-A and rhHex-B showed high stability. This study demonstrates P. pastoris GS115 potential for polymeric lysosomal enzyme production, and describes the characterization of recombinant β-hexosaminidases produced within the same host.

  9. Genome-scale metabolic model of Pichia pastoris with native and humanized glycosylation of recombinant proteins.

    Science.gov (United States)

    Irani, Zahra Azimzadeh; Kerkhoven, Eduard J; Shojaosadati, Seyed Abbas; Nielsen, Jens

    2016-05-01

    Pichia pastoris is used for commercial production of human therapeutic proteins, and genome-scale models of P. pastoris metabolism have been generated in the past to study the metabolism and associated protein production by this yeast. A major challenge with clinical usage of recombinant proteins produced by P. pastoris is the difference in N-glycosylation of proteins produced by humans and this yeast. However, through metabolic engineering, a P. pastoris strain capable of producing humanized N-glycosylated proteins was constructed. The current genome-scale models of P. pastoris do not address native nor humanized N-glycosylation, and we therefore developed ihGlycopastoris, an extension to the iLC915 model with both native and humanized N-glycosylation for recombinant protein production, but also an estimation of N-glycosylation of P. pastoris native proteins. This new model gives a better prediction of protein yield, demonstrates the effect of the different types of N-glycosylation of protein yield, and can be used to predict potential targets for strain improvement. The model represents a step towards a more complete description of protein production in P. pastoris, which is required for using these models to understand and optimize protein production processes. © 2015 Wiley Periodicals, Inc.

  10. Preparation of Tc-99m-macroaggregated albumin from recombinant human albumin for lung perfusion imaging.

    Science.gov (United States)

    Hunt, A P; Frier, M; Johnson, R A; Berezenko, S; Perkins, A C

    2006-01-01

    Human serum albumin (HSA) extracted from pooled blood taken from human donors is used in the production of (99m)Tc-labelled macroaggregated albumin (MAA) for lung perfusion imaging. However, concerns for the safety of blood-derived products due to potential contamination by infective agents (e.g. new variant CJD), make alternative production methods necessary. Recombinant DNA technology is a promising method of albumin production avoiding problems associated with human-derived HSA. This paper presents results comparing MAA prepared from recombinant human albumin (rHA, Recombumin) (rMAA) with in-house produced HSA MAA (hMAA) and commercially available MAA (cMAA). (99m)Tc-MAA was prepared using previously published production methods by heating a mixture of albumin and stannous chloride in acetate buffer (pH 5.4) at 70 degrees C for 20 min. Parameters investigated include aggregate size, radiolabelling efficiency, radiochemical and aggregate stability at 4 degrees C and in vitro (in whole human blood) at 37 degrees C and biodistribution studies. Results showed that rMAA could be produced with similar morphology, labelling efficiency and stability to hMAA and cMAA. Our findings confirm that rHA shows significant potential as a direct replacement for HSA in commercially available MAA.

  11. Vaccinia virus recombinants expressing chimeric proteins of human immunodeficiency virus and gamma interferon are attenuated for nude mice.

    OpenAIRE

    Giavedoni, L D; Jones, L; Gardner, M B; Gibson, H L; Ng, C T; Barr, P J; Yilma, T

    1992-01-01

    We have developed a method for attenuating vaccinia virus recombinants by expressing a fusion protein of a lymphokine and an immunogen. Chimeric genes were constructed that coded for gamma interferon (IFN-gamma) and structural proteins of the human immunodeficiency virus type 1 (HIV-1). In this study, we describe the biological and immunological properties of vaccinia virus recombinants expressing chimeric genes of murine or human IFN-gamma with glycoprotein gp120, gag, and a fragment of gp41...

  12. Recombinant expression in E. coli of human FGFR2 with its transmembrane and extracellular domains

    Directory of Open Access Journals (Sweden)

    Adam Bajinting

    2017-06-01

    Full Text Available Fibroblast growth factor receptors (FGFRs are a family of receptor tyrosine kinases containing three domains: an extracellular receptor domain, a single transmembrane helix, and an intracellular tyrosine kinase domain. FGFRs are activated by fibroblast growth factors (FGFs as part of complex signal transduction cascades regulating angiogenesis, skeletal formation, cell differentiation, proliferation, cell survival, and cancer. We have developed the first recombinant expression system in E. coli to produce a construct of human FGFR2 containing its transmembrane and extracellular receptor domains. We demonstrate that the expressed construct is functional in binding heparin and dimerizing. Size exclusion chromatography demonstrates that the purified FGFR2 does not form a complex with FGF1 or adopts an inactive dimer conformation. Progress towards the successful recombinant production of intact FGFRs will facilitate further biochemical experiments and structure determination that will provide insight into how extracellular FGF binding activates intracellular kinase activity.

  13. Recombinant Newcastle disease virus-vectored vaccines against human and animal infectious diseases.

    Science.gov (United States)

    Duan, Zhiqiang; Xu, Houqiang; Ji, Xinqin; Zhao, Jiafu

    2015-01-01

    Recent advances in recombinant genetic engineering techniques have brought forward a leap in designing new vaccines in modern medicine. One attractive strategy is the application of reverse genetics technology to make recombinant Newcastle disease virus (rNDV) deliver protective antigens of pathogens. In recent years, numerous studies have demonstrated that rNDV-vectored vaccines can induce quicker and better humoral and mucosal immune responses than conventional vaccines and are protective against pathogen challenges. With deeper understanding of NDV molecular biology, it is feasible to develop gene-modified rNDV vaccines accompanied by good safety, high efficacy, low toxicity and better immunogenicity. This review summarizes the development of reverse genetics technology in using NDV as a promising vaccine vector to design new vaccines for human and animal use.

  14. Studies of the cytosolic thymidine kinase in human cells and comparison to the recombinantly expressed enzyme

    DEFF Research Database (Denmark)

    Kock Jensen, Helle

    by recombinant technics to examine the relation between the TKl gene and the TKl protein. In the second part of this investigation a direct expression system for human TKl in E.coli was developed to produce a source of high amounts of TKl, to be able to examine the structure of TKl. The resulting recombinant TKl...... cells and that this modification can not be performed in E.coli....... infections. In the first part of the present investigation a sensitive test for quantitating TKl mRNA (competitive PCR) is developed and the results show that PHA stimulated lymphocytes reveal the same pattern concerning expression of TKl mRNA and TKl enzyme activity as serum-stimulated cells. This pattern...

  15. Enzymatic cross-linking of human recombinant elastin (HELP) as biomimetic approach in vascular tissue engineering.

    Science.gov (United States)

    Bozzini, Sabrina; Giuliano, Liliana; Altomare, Lina; Petrini, Paola; Bandiera, Antonella; Conconi, Maria Teresa; Farè, Silvia; Tanzi, Maria Cristina

    2011-12-01

    The use of polymers naturally occurring in the extracellular matrix (ECM) is a promising strategy in regenerative medicine. If compared to natural ECM proteins, proteins obtained by recombinant DNA technology have intrinsic advantages including reproducible macromolecular composition, sequence and molecular mass, and overcoming the potential pathogens transmission related to polymers of animal origin. Among ECM-mimicking materials, the family of recombinant elastin-like polymers is proposed for drug delivery applications and for the repair of damaged elastic tissues. This work aims to evaluate the potentiality of a recombinant human elastin-like polypeptide (HELP) as a base material of cross-linked matrices for regenerative medicine. The cross-linking of HELP was accomplished by the insertion of cross-linking sites, glutamine and lysine, in the recombinant polymer and generating ε-(γ-glutamyl) lysine links through the enzyme transglutaminase. The cross-linking efficacy was estimated by infrared spectroscopy. Freeze-dried cross-linked matrices showed swelling ratios in deionized water (≈2500%) with good structural stability up to 24 h. Mechanical compression tests, performed at 37°C in wet conditions, in a frequency sweep mode, indicated a storage modulus of 2/3 kPa, with no significant changes when increasing number of cycles or frequency. These results demonstrate the possibility to obtain mechanically resistant hydrogels via enzymatic crosslinking of HELP. Cytotoxicity tests of cross-linked HELP were performed with human umbilical vein endothelial cells, by use of transwell filter chambers for 1-7 days, or with its extracts in the opportune culture medium for 24 h. In both cases no cytotoxic effects were observed in comparison with the control cultures. On the whole, the results suggest the potentiality of this genetically engineered HELP for regenerative medicine applications, particularly for vascular tissue regeneration.

  16. Safety of recombinant human platelet-derived growth factor-BB in Augment® Bone Graft

    Directory of Open Access Journals (Sweden)

    Luis A Solchaga

    2012-12-01

    Full Text Available This article discusses nonclinical and clinical data regarding the safety of recombinant human platelet-derived growth factor-BB as a component of the Augment® Bone Graft (Augment. Augment is a bone graft substitute intended to be used as an alternative to autologous bone graft in the fusion of hindfoot and ankle joints. Nonclinical studies included assessment of the pharmacokinetic profile of intravenously administered recombinant human platelet-derived growth factor-BB in rat and dog, effects of intravenous administration of recombinant human platelet-derived growth factor-BB in a reproductive and development toxicity study in rats, and chronic toxicity and carcinogenicity of Augment in a 12-month implantation model. These studies showed that systemic exposure was brief and clearance was rapid. No signs of toxicity, carcinogenicity, or tumor promotion were observed even with doses far exceeding the maximum clinical dose. Results of clinical trials (605 participants and commercial use of recombinant human platelet-derived growth factor-BB containing products indicate that these products are not associated with increased incidence of adverse events or cancer. The safety data presented provide evidence that recombinant human platelet-derived growth factor-BB is a safe therapeutic when used in combination products as a single administration during surgical procedures for bone repair and fusion. There is no evidence associating use of recombinant human platelet-derived growth factor-BB in Augment with chronic toxicity, carcinogenicity, or tumor promotion.

  17. Radioiodine therapy of thyroid autonomy

    Energy Technology Data Exchange (ETDEWEB)

    Reiners, Christoph; Schneider, Peter [Clinic and Policlinic for Nuclear Medicine, University of Wuerzburg, Josef-Schneider-Strasse 2, 97080 Wuerzburg (Germany)

    2002-08-01

    Over half a century, treatment of thyroid autonomy with an oral dose of iodine-131 has proven to be effective. The optimum management strategy for the patient is, however, still a matter of debate. The article provides an overview of the pathogenesis of functional autonomy and its clinical relevance. According to the guidelines on both sides of the Atlantic, radioiodine treatment is considered the most comfortable and economical approach to the treatment of the toxic nodular goitre. Some differences in the preparation procedures in the guidelines of the American and the German Society of Nuclear Medicine are discussed with respect to therapy results and the subtypes of thyroid autonomy. The results of studies are summarised concerning changes in thyroid function and thyroid volume after a course of radioiodine treatment. Therapy-related risks, such as immunogenic hypothyroidism or thyroid cancer, are discussed. {sup 131}I treatment of functional autonomy and hyperthyroidism is considered an effective and safe procedure. (orig.)

  18. Radioiodine therapy of thyroid autonomy

    International Nuclear Information System (INIS)

    Reiners, Christoph; Schneider, Peter

    2002-01-01

    Over half a century, treatment of thyroid autonomy with an oral dose of iodine-131 has proven to be effective. The optimum management strategy for the patient is, however, still a matter of debate. The article provides an overview of the pathogenesis of functional autonomy and its clinical relevance. According to the guidelines on both sides of the Atlantic, radioiodine treatment is considered the most comfortable and economical approach to the treatment of the toxic nodular goitre. Some differences in the preparation procedures in the guidelines of the American and the German Society of Nuclear Medicine are discussed with respect to therapy results and the subtypes of thyroid autonomy. The results of studies are summarised concerning changes in thyroid function and thyroid volume after a course of radioiodine treatment. Therapy-related risks, such as immunogenic hypothyroidism or thyroid cancer, are discussed. 131 I treatment of functional autonomy and hyperthyroidism is considered an effective and safe procedure. (orig.)

  19. Radioiodine treatment of Grave's disease

    International Nuclear Information System (INIS)

    Heidenreich, P.; Vogt, H.; Dorn, R.; Graf, G.; Kopp, J.

    2001-01-01

    In Germany radioiodine therapy of Grave's disease is performed in patients older than 20 years, after at least one year of unsuccessful antithyroid therapy, intolerance against antithyroid medication, recurrences after surgical interventions and small goiters. Hyperthyroidism is eliminated with an ablative dose concept (300 Gy) in more than 90% associated with rate of hypothyroidism in the outcome of also more than 90%. Adverse prognostic factors are an insufficient dose to the thyroid and/or concomitant antithyreoid medication. Radioiodine therapy in Germany must be an inpatient single time approach due to quality assurance and radiation protection reasons. The mean hospitalization is only 3-4 days with the patient being discharged at an annual dose of less than 1 mSv at 2 m distance (dose rate at discharge [de

  20. Protection of non-human primates against rabies with an adenovirus recombinant vaccine

    International Nuclear Information System (INIS)

    Xiang, Z.Q.; Greenberg, L.; Ertl, H.C.; Rupprecht, C.E.

    2014-01-01

    Rabies remains a major neglected global zoonosis. New vaccine strategies are needed for human rabies prophylaxis. A single intramuscular immunization with a moderate dose of an experimental chimpanzee adenovirus (Ad) vector serotype SAd-V24, also termed AdC68, expressing the rabies virus glycoprotein, resulted in sustained titers of rabies virus neutralizing antibodies and protection against a lethal rabies virus challenge infection in a non-human primate model. Taken together, these data demonstrate the safety, immunogenicity, and efficacy of the recombinant Ad-rabies vector for further consideration in human clinical trials. - Highlights: • Pre-exposure vaccination with vaccine based on a chimpanzee derived adenovirus protects against rabies. • Protection is sustained. • Protection is achieved with single low-dose of vaccine given intramuscularly. • Protection is not affected by pre-existing antibodies to common human serotypes of adenovirus

  1. Protection of non-human primates against rabies with an adenovirus recombinant vaccine

    Energy Technology Data Exchange (ETDEWEB)

    Xiang, Z.Q. [The Wistar Institute of Anatomy and Biology, Philadelphia, PA (United States); Greenberg, L. [Centers for Disease Control and Prevention, Atlanta, GA (United States); Ertl, H.C., E-mail: ertl@wistar.upenn.edu [The Wistar Institute of Anatomy and Biology, Philadelphia, PA (United States); Rupprecht, C.E. [The Global Alliance for Rabies Control, Manhattan, KS (United States); Ross University School of Veterinary Medicine, Basseterre (Saint Kitts and Nevis)

    2014-02-15

    Rabies remains a major neglected global zoonosis. New vaccine strategies are needed for human rabies prophylaxis. A single intramuscular immunization with a moderate dose of an experimental chimpanzee adenovirus (Ad) vector serotype SAd-V24, also termed AdC68, expressing the rabies virus glycoprotein, resulted in sustained titers of rabies virus neutralizing antibodies and protection against a lethal rabies virus challenge infection in a non-human primate model. Taken together, these data demonstrate the safety, immunogenicity, and efficacy of the recombinant Ad-rabies vector for further consideration in human clinical trials. - Highlights: • Pre-exposure vaccination with vaccine based on a chimpanzee derived adenovirus protects against rabies. • Protection is sustained. • Protection is achieved with single low-dose of vaccine given intramuscularly. • Protection is not affected by pre-existing antibodies to common human serotypes of adenovirus.

  2. Potency of full-length MGF to induce maximal activation of the IGF-I R Is similar to recombinant human IGF-I at high equimolar concentrations

    NARCIS (Netherlands)

    J.A.M.J.L. Janssen (Joseph); L.J. Hofland (Leo); C.J. Strasburger; E.S.R.D. Van Dungen (Elisabeth S.R. Den); M. Thevis (Mario)

    2016-01-01

    textabstractAims To compare full-length mechano growth factor (full-length MGF) with human recombinant insulin-like growth factor-I (IGF-I) and human recombinant insulin (HI) in their ability to activate the human IGF-I receptor (IGF-IR), the human insulin receptor (IR-A) and the human insulin

  3. Differentiated thyroid cancer. New concept of radioiodine ablation; Differenziertes Schilddruesenkarzinom. Fortschritte bei der Radioiodablation

    Energy Technology Data Exchange (ETDEWEB)

    Dietlein, M.; Kobe, C. [Universitaetsklinikum Koeln (Germany). Klinik und Poliklinik fuer Nuklearmedizin; Universitaetsklinikum Koeln (Germany), Zentrum fuer Integrierte Onkologie Koeln-Bonn; Luster, M. [Universitaetsklinikum Ulm (Germany). Klinik und Poliklinik fuer Nuklearmedizin

    2010-12-15

    Ablative radioiodine therapy is the treatment of choice in patients with differentiated thyroid cancer, the only exception being the unifocal, very small papillary thyroid cancer. The TSH-stimulation can be achieved by a waiting period for 2-3 weeks after thyroidectomy without medication or by the use of recombinant human TSH (rhTSH). Both options lead to high success rates. 'Single dose cure' using activities between 1.85 and 3.7 GBq {sup 131}I is standard. Since 2010 rhTSH is approved by the EMA for the indications pT1-4, N0-1, cM0. Survey studies did not find any inferiority of ablation with rhTSH or iatrogenic hypothyroidism in the high-risk patient group. Renal clearance is not reduced after rhTSH administration, thus the {sup 131}I blood dose and the whole body doses are lower in patients under rhTSH. Comparing identical {sup 131}I activities after endogeneous or exogeneous stimulation, rhTSH will minimize the acute adverse effects of {sup 131}I. A short-term withdrawal of levothyroxine some days before rhTSH-injection lowers the iodine plasma level, which may be advantageous for the ablation success if lower 131I activities are used. A rhTSH-based diagnostic {sup 131}I whole-body scintigraphy 3-6 months after ablation is standard for therapy control. At this time, the rhTSH-stimulated thyroglobulin-level is essential for a personalized risk stratification. Tg-measurements by a second generation assay should be used for follow-up care. Metaanalyses have shown that radioiodine ablation lowers the mortality rate, the risk of locoregional recurrences and the risk of late metastasizing. Therefore, ablation has shown a clear benefit. (orig.)

  4. Radioiodine uptake of undifferentiated thyroid cancer cells by adenovirus-mediated Na+/ I- symporter gene transfer

    Energy Technology Data Exchange (ETDEWEB)

    So, Y.; Lee, Y. J.; Shin, J. H.; Oh, H. J.; Chung, J. K.; Lee, M. C.; Cho, B. Y. [College of Medicine, Univ. of Seoul National, Seoul (Korea, Republic of); Lee, K. H. [Samsung Medical Center, Seoul (Korea, Republic of)

    2003-07-01

    To increase radioiodine uptake on undifferentiated thyroid cancer cell (ARO cells) by adenovirus-mediated human Na+/I- symporter (hNIS) gene transfer. Recombinant adenovirus Ad-hNIS was manufactured successfully. After transfecting Ad-hNIS on ARO cells, in vitro I-125 uptake and efflux studies were performed. For in vivo studies, 1.510'8 p.f.u. (50 1) of Ad-hNIS was injected into xenograft ARO tumors on the R thigh of BALB/c nu/nu mice (n=12), and same amount of normal saline was injected into xenograft ARO tumors on the L thigh. Two, 3, 4 and 6 days after intratumoral injection of Ad-hNIS, I-131 images (3 mice per day) were taken and xenograft tumors on both thighs were all excised. Total RNA was extracted from each tumor tissue and RT-PCR was performed to confirm the hNIS expression of Ad-hNIS injected xenograft ARO tumors. I-125 uptake of Ad-hNIS transfected ARO cells was increased up to 233 folds at 120 minutes in vitro. I-125 efflux study revealed rapid washout of I-125 from Ad-hNIS transfected ARO cells. On dynamic image, I-131 uptake of Ad-hNIS injected ARO tumor was continuously increased until 60 minutes. Mean count ratios of xenograft ARO tumors (R/L) of 60 minutes I-131 images at 2, 3, 4 and 6 days after Ad-hNIS injection were 2.85, 2.54, 2.31, and 2.18, each. On RT-PCR, hNIS expression of Ad-hNIS transfected ARO xenograft tumors was confirmed. Radioiodine uptake was successfully increased in ARO cells by adenovirus-mediated hNIs gene transfer both in vitro and in vivo.

  5. [Genetic evidence for recombination and mutation in the emergence of human enterovirus 71].

    Science.gov (United States)

    Liu, Ai-Ping; Tan, Hui; Xie, Qun; Chen, Bai-Tang; Liu, Xiao-Feng; Zhang, Yong

    2014-09-01

    We wished to understand the genetic recombination and phylogenetic characteristics of human en- terovirus A71 (EV-A71) and to explore its potential virulence-related sites. Full-length genomes of three EV-A71 strains isolated from patients in Chenzhou City (China) were sequenced and analyzed. Possible re- combination events and crossover sites were analyzed with Recombination Detection Program v4. 1. 6 by comparison with the complete genome sequences of 231 strains of EV-A71. Similarly, plot and bootscanning analyses were undertaken with SimPlot v3. 5. 1. Phylogenetic trees based on the sequences of VP1 regions were constructed with MEGA v5. 2 using the Kimura two-parameter model and neighbor-joining method. Results suggested that recombination events were detected among the three EV-A71 isolates from Chenzhou City. The common main parent sequence was from JF799986 isolated from samples in Guang- zhou City (China) in 2009, and the minor parent sequence was TW/70516/08. Intertypic recombination e- vents were found in the C4b strain (strain SHZH98 isolated in 1998) and C4a strain (Fuyang strain isola- ted in 2008) with the prototype strains of CVA4 and CVA14 in the 3D region. The chi-square test was used to screen-out potential virulence-related sites with nucleotide substitutions of different types of hand, foot, and mouth disease (HFMD) cases using SPSS v19.0. Results suggested that there were no significant nucleotide substitutions between death cases and severe-HFMD cases. Eighteen significant nucleotide substitutions were found between death/severe-HFMD cases and mild-HFMD cases, and all these 18 substitutions were distributed only in P2 and P3 regions. Intertypic recombination among the predominant circulating EV-A71 strains in the Chinese mainland and other EV-A strains probably dates before 1998, and intratypic recombination might have occurred frequently in the HFMD outbreak from 2008 to 2012. Substitutions in the non-capsid region may be correlated with the

  6. A comparative study of recombinant and native frutalin binding to human prostate tissues

    Directory of Open Access Journals (Sweden)

    Domingues Lucília

    2009-09-01

    Full Text Available Abstract Background Numerous studies indicate that cancer cells present an aberrant glycosylation pattern that can be detected by lectin histochemistry. Lectins have shown the ability to recognise these modifications in several carcinomas, namely in the prostate carcinoma, one of the most lethal diseases in man. Thus, the aim of this work was to investigate if the α-D-galactose-binding plant lectin frutalin is able to detect such changes in the referred carcinoma. Frutalin was obtained from different sources namely, its natural source (plant origin and a recombinant source (Pichia expression system. Finally, the results obtained with the two lectins were compared and their potential use as prostate tumour biomarkers was discussed. Results The binding of recombinant and native frutalin to specific glycoconjugates expressed in human prostate tissues was assessed by using an immuhistochemical technique. A total of 20 cases of prostate carcinoma and 25 cases of benign prostate hyperplasia were studied. Lectins bound directly to the tissues and anti-frutalin polyclonal antibody was used as the bridge to react with the complex biotinilated anti-rabbit IgG plus streptavidin-conjugated peroxidase. DAB was used as visual indicator to specifically localise the binding of the lectins to the tissues. Both lectins bound to the cells cytoplasm of the prostate carcinoma glands. The binding intensity of native frutalin was stronger in the neoplasic cells than in hyperplasic cells; however no significant statistical correlation could be found (P = 0.051. On the other hand, recombinant frutalin bound exclusively to the neoplasic cells and a significant positive statistical correlation was obtained (P Conclusion Native and recombinant frutalin yielded different binding responses in the prostate tissues due to their differences in carbohydrate-binding affinities. Also, this study shows that both lectins may be used as histochemical biomarkers for the prostate

  7. Recombination gives a new insight in the effective population size and the history of the old world human populations.

    Science.gov (United States)

    Melé, Marta; Javed, Asif; Pybus, Marc; Zalloua, Pierre; Haber, Marc; Comas, David; Netea, Mihai G; Balanovsky, Oleg; Balanovska, Elena; Jin, Li; Yang, Yajun; Pitchappan, R M; Arunkumar, G; Parida, Laxmi; Calafell, Francesc; Bertranpetit, Jaume

    2012-01-01

    The information left by recombination in our genomes can be used to make inferences on our recent evolutionary history. Specifically, the number of past recombination events in a population sample is a function of its effective population size (Ne). We have applied a method, Identifying Recombination in Sequences (IRiS), to detect specific past recombination events in 30 Old World populations to infer their Ne. We have found that sub-Saharan African populations have an Ne that is approximately four times greater than those of non-African populations and that outside of Africa, South Asian populations had the largest Ne. We also observe that the patterns of recombinational diversity of these populations correlate with distance out of Africa if that distance is measured along a path crossing South Arabia. No such correlation is found through a Sinai route, suggesting that anatomically modern humans first left Africa through the Bab-el-Mandeb strait rather than through present Egypt.

  8. Radio-iodine in thyroid glands of swans, farm animals and humans, also in algae and river water from the Thames Valley, England

    Energy Technology Data Exchange (ETDEWEB)

    Howe, J.R.; Lloyd, M.K.

    1986-01-01

    A highly sensitive counting system has been used to measure radio-iodine in environmental samples from the Thames Valley. Iodine-125 and occasionally iodine-131 have been found in the thyroid glands of most of the swans that have died on the River Thames, the River Wey and the Grand Union Canal, and in algae and water samples from the Thames and many of its tributaries. The presence of this activity is ascribed to the waste discarded into the drainage system by hospitals and research laboratories, reaching the rivers via the effluent from sewage treatment works. The Thames is used as a source of drinking water, particularly in London and its western approaches. Weed and water samples collected from river water abstraction points, reservoirs, tap water supplies, and animal water troughs fed from this supply all contained low levels of iodine-125. The drinking water route can account for the iodine-125 found in the thyroids of farm animals from west Surrey and in a few people living in London. The amounts found constitute a trivial radiation dose to man and animals as they are far below the acceptable limit of exposure for man.

  9. Expression of Recombinant Human Coagulation Factor VII by the Lizard Leishmania Expression System

    Directory of Open Access Journals (Sweden)

    Sina Mirzaahmadi

    2011-01-01

    Full Text Available The variety of recombinant protein expression systems have been developed as a resource of FVII gene expression. In the current study, the authors used a novel protein expression system based on the Iranian Lizard Leishmania, a trypanosomatid protozoan as a host for expression of FVII. Plasmid containing cDNA encoding full-length human FVII was introduced into Lizard Leishmania and positive transfectants were analyzed by SDS-PAGE and Western blot analysis. Furthermore, biological activity of purified protein was detected by PT assay. The recombinant strain harboring a construct was analyzed for expression of FVII at the mRNA and protein level. Purified rFVII was obtained and in order to confirm the purified compound was in fact rFVII. Western blot analysis was carried out. Clotting time in PT assay was reduced about 30 seconds with the purified rFVII. In Conclusion, this study has demonstrated, for the first time, that Leishmania cells can be used as an expression system for producing recombinant FVII.

  10. Optimizing Production of Antigens and Fabs in the Context of Generating Recombinant Antibodies to Human Proteins.

    Directory of Open Access Journals (Sweden)

    Nan Zhong

    Full Text Available We developed and optimized a high-throughput project workflow to generate renewable recombinant antibodies to human proteins involved in epigenetic signalling. Three different strategies to produce phage display compatible protein antigens in bacterial systems were compared, and we found that in vivo biotinylation through the use of an Avi tag was the most productive method. Phage display selections were performed on 265 in vivo biotinylated antigen domains. High-affinity Fabs (<20nM were obtained for 196. We constructed and optimized a new expression vector to produce in vivo biotinylated Fabs in E. coli. This increased average yields up to 10-fold, with an average yield of 4 mg/L. For 118 antigens, we identified Fabs that could immunoprecipitate their full-length endogenous targets from mammalian cell lysates. One Fab for each antigen was converted to a recombinant IgG and produced in mammalian cells, with an average yield of 15 mg/L. In summary, we have optimized each step of the pipeline to produce recombinant antibodies, significantly increasing both efficiency and yield, and also showed that these Fabs and IgGs can be generally useful for chromatin immunoprecipitation (ChIP protocols.

  11. Anti-tumor effect of a recombinant plasmid expressing human interleukin-12: an initial research

    International Nuclear Information System (INIS)

    Zheng Chuansheng; Xia Xiangwen; Feng Gansheng; Li Xin; Liang Huimin; Liang Bin

    2010-01-01

    Objective: To study the anti-tumor effect of a recombinant plasmid expressing human interleukin-12 (pEGFP-CI I L- 12) in vivo and in vitro. Methods: We transduct the recombinant gene (pEGFP-CI I L-12) to liver cancer cell HepG 2 in vitro, and detect reproductive activity of the cell using MTT and the activity of expressing vascular endothelial growth factor(VEGF) using semiquantitative PCR. And then, we deliver the gene to rabbit liver tumor tissue intraarterial and combine with chemoembolization to observe the anti- tumor effect to VX 2 tumor in vivo. Results: There are no statistical difference compared With control group in activity of reproductive and expressing VEGF in vitro. In vivo, tumor growth rate significantly reduce in gene therapy combined with chemoembolization group. Conclusion: Recombinant gene (pEGFP-Cl I L-12) exhibit significant anti-tumor effect in vivo but not in vitro, perhaps the anti-tumor effect is associated with an indirect pathway instead of a direct pathway. (authors)

  12. Towards a more precise serological diagnosis of human tegumentary leishmaniasis using Leishmania recombinant proteins.

    Directory of Open Access Journals (Sweden)

    Ana Paula Souza

    Full Text Available BACKGROUND: Exposure to Leishmania induces a humoral immune response that can be used as a marker of parasite exposure. METHODOLOGY/PRINCIPAL FINDINGS: Herein, ELISA was used to screen sera from patients with Tegumentary Leishmaniasis (TL against different L. infantum-chagasi-derived recombinant proteins (rHSP70, rH2A, rH2B, rH3, rH4 and rKMP11. Among the recombinant proteins, rHSP70 and rH2A showed the best reactivity against human sera obtained from endemic areas of TL. Receiver-Operator Characteristics (ROC curve analysis was used to identify the effectiveness of these proteins for serodiagnosis of TL. ROC curves confirmed the superior performance of rHSP70 and rH2A, in comparison to the other tested recombinant proteins. Additionally, we evaluated the specificity of the response to rHSP70 and rH2A by testing sera obtained from patients with Chagas' disease, Tuberculosis, Leprosy or Systemic Lupus Erythematosus. In this case, rHSP70 displayed an increased ability to discriminate diseases, in comparison to SLA. CONCLUSION: Our results raise possibility of using rHSP70 for the serodiagnosis of TL.

  13. Hydroxylation of recombinant human collagen type I alpha 1 in transgenic maize co-expressed with a recombinant human prolyl 4-hydroxylase

    Directory of Open Access Journals (Sweden)

    Pappu Kameshwari M

    2011-06-01

    Full Text Available Abstract Background Collagens require the hydroxylation of proline (Pro residues in their triple-helical domain repeating sequence Xaa-Pro-Gly to function properly as a main structural component of the extracellular matrix in animals at physiologically relevant conditions. The regioselective proline hydroxylation is catalyzed by a specific prolyl 4-hydroxylase (P4H as a posttranslational processing step. Results A recombinant human collagen type I α-1 (rCIα1 with high percentage of hydroxylated prolines (Hyp was produced in transgenic maize seeds when co-expressed with both the α- and β- subunits of a recombinant human P4H (rP4H. Germ-specific expression of rCIα1 using maize globulin-1 gene promoter resulted in an average yield of 12 mg/kg seed for the full-length rCIα1 in seeds without co-expression of rP4H and 4 mg/kg seed for the rCIα1 (rCIα1-OH in seeds with co-expression of rP4H. High-resolution mass spectrometry (HRMS analysis revealed that nearly half of the collagenous repeating triplets in rCIα1 isolated from rP4H co-expressing maize line had the Pro residues changed to Hyp residues. The HRMS analysis determined the Hyp content of maize-derived rCIα1-OH as 18.11%, which is comparable to the Hyp level of yeast-derived rCIα1-OH (17.47% and the native human CIa1 (14.59%, respectively. The increased Hyp percentage was correlated with a markedly enhanced thermal stability of maize-derived rCIα1-OH when compared to the non-hydroxylated rCIα1. Conclusions This work shows that maize has potential to produce adequately modified exogenous proteins with mammalian-like post-translational modifications that may be require for their use as pharmaceutical and industrial products.

  14. Heterogeneity within populations of recombinant Chinese hamster ovary cells expressing human interferon-gamma.

    Science.gov (United States)

    Coppen, S R; Newsam, R; Bull, A T; Baines, A J

    1995-04-20

    The Chinese hamster ovary (CHO) cell line has great commercial importance in the production of recombinant human proteins, especially those for therapeutic use. Much attention has been paid to CHO cell population physiology in order to define factors affecting product fidelity and yield. Such studies have revealed that recombinant proteins, including human interferon-gamma (IFN-gamma), can be heterogeneous both in glycosylation and in proteolytic processing. The type of heterogeneity observed depends on the growth physiology of the cell population, although the relationship between them is complex. In this article we report results of a cytological study of the CHO320 line which expresses recombinant human IFN-gamma. When grown in suspension culture, this cell line exhibited three types of heterogeneity: (1) heterogeneity of the production of IFN-gamma within the cell population, (2) heterogeneity of the number of nuclei and mitotic spindles in dividing cells, and (3) heterogeneity of cellular environment. The last of these arises from cell aggregates which form in suspension culture: Some cells are exposed to the culture medium; others are fully enclosed within the mass with little or no direct access to the medium. Thus, live cells producing IFN-gamma are heterogeneous in their environment, with variable access to O(2) and nutrients. Within the aggregates, it appears that live cells proliferate on a dead cell mass. The layer of live cells can be several cells deep. Specific cell-cell attachments are observed between the living cells in these aggregates. Two proteins, known to be required for the formation of certain types of intercellular junctions, spectrin and vinculin, have been localized to the regions of cell-cell contact. The aggregation of the cells appears to be an active process requiring protein synthesis. (c) 1995 John Wiley & Sons, Inc.

  15. Characterization of the oligosaccharide structure of human glycosylated prolactin (G-hPRL) native and recombinant

    International Nuclear Information System (INIS)

    Marcos Vinicius Nucci Capone

    2013-01-01

    Human prolactin (hPRL) is a polypeptide hormone secreted by the anterior pituitary under the regulation of the hypothalamus, involved in a variety of biological processes such as mammary gland development and lactation. The recombinant product is important in medical diagnosis and treatment of failure of lactation. This hormone may occur in the form of non-glycosylated protein (NGhPRL) and glycosylated (G-hPRL) with molecular weights of approximately 23 and 25 kilodalton (kDa), respectively; has a single N-glycosylation site located at asparagine (Asn) position 31, which is partially occupied, thus being a particularly interesting model of glycosylation. The biological activity of G-hPRL is lower compared to NG-hPRL (~4 times) and its physiological function is not well defined: the portion of carbohydrate appears to have an important role in the hormone biosynthesis, secretion, biological activity, and plasma survival of the hormone. The main objective of this study was to compare the structures of N-glycans present in glycosylated pituitary prolactin (G-hPRL-NHPP) with those present in the recombinant. To obtain the recombinant G-hPRL the production was performed in laboratory scale from Chinese hamster ovary cells (CHO), genetically modified and adapted to growth in suspension. Cycloheximide (CHX), whose main effect was to increase the ratio G-hPRL/NG-hPRL from 5% to 38% was added to the culture medium, thereby facilitating the purification of G-hPRL. The G-hPRL was purified in two steps, a cation exchanger followed by a purification by reversed-phase high performance liquid chromatography (RP-HPLC) which demonstrated the efficient separation of the two isoforms of hPRL. Recombinant G-hPRL-IPEN was well characterized by several techniques confirming its purity and biological activity, including comparisons with other reference preparation of pituitary origin purchased from the N ational Hormone & Peptide Program (NHPPU. S.) . The composition of N-glycans present

  16. Generation of meiomaps of genome-wide recombination and chromosome segregation in human oocytes

    DEFF Research Database (Denmark)

    Ottolini, Christian S; Capalbo, Antonio; Newnham, Louise

    2016-01-01

    We have developed a protocol for the generation of genome-wide maps (meiomaps) of recombination and chromosome segregation for the three products of human female meiosis: the first and second polar bodies (PB1 and PB2) and the corresponding oocyte. PB1 is biopsied and the oocyte is artificially......-nucleotide polymorphisms (SNPs) genome-wide by microarray. Informative maternal heterozygous SNPs are phased using a haploid PB2 or oocyte as a reference. A simple algorithm is then used to identify the maternal haplotypes for each chromosome, in all of the products of meiosis for each oocyte. This allows mapping...

  17. Exogenous recombinant human growth hormone effects during suboptimal energy and zinc intake

    OpenAIRE

    Rising, Russell; Scaglia, Julio F; Cole, Conrad; Tverskaya, Rozalia; Duro, Debora; Lifshitz, Fima

    2005-01-01

    Abstract Background Energy and Zinc (Zn) deficiencies have been associated with nutritional related growth retardation as well as growth hormone (GH) resistance. In this study, the relationship between suboptimal energy and/or Zn intake and growth in rats and their response to immunoreactive exogenous recombinant human GH (GHi), was determined. Results Rats treated with GHi and fed ad-libitum energy and Zn (100/100) had increased IGFBP-3 (p < 0.05) as compared with NSS (215 ± 23 vs. 185 ± 17 ...

  18. Effects of low-dose recombinant human erythropoietin treatment on cognitive performance

    DEFF Research Database (Denmark)

    Viuff, Søren Lundgaard; Plenge, Ulla; Belhage, Bo

    2017-01-01

    , NUFI or self-reported results between the groups. CONCLUSIONS: In this small study, we found no significant effect of low-dose or micro-dose rhEpo on visual attention, cognitive performance in complex cognitive tasks or self-experienced cognitive performance compared with placebo. FUNDING: The Aase......INTRODUCTION: High-dose recombinant human erythropoietin (rhEpo) has been shown to improve cognitive performance in both healthy volunteers and in patients suffering from diseases affecting the brain. The aim of this study was to examine whether administration of low-dose and even micro-dose rh...

  19. Effects of low-dose recombinant human erythropoietin treatment on cognitive performance

    DEFF Research Database (Denmark)

    Viuff, Søren Lundgaard; Plenge, Ulla; Belhage, Bo

    2017-01-01

    -reported results between the groups. Conclusions: In this small study, we found no significant effect of low-dose or micro-dose rhEpo on visual attention, cognitive performance in complex cognitive tasks or self-experienced cognitive performance compared with placebo. Funding: The Aase and Ejnar Danielsen......Introduction: High-dose recombinant human erythropoietin (rhEpo) has been shown to improve cognitive performance in both healthy volunteers and in patients suffering from diseases affecting the brain. The aim of this study was to examine whether administration of low-dose and even micro-dose rh...

  20. Brief study about the distribution of recombinant human Epidermic Growth Factor (rh-EGF)

    International Nuclear Information System (INIS)

    Rodriguez Garcia, J.C.; De Dios D Espaux, R.; Bello Garciga, J.L.

    1997-01-01

    This report describes results of the study about biodistribution of I-125 recombinant human Epidermic Growth Factor (rhEGF). The radiolabelled product was administrated to Sprague Dawley rats in three different ways: intramuscular, subcutaneous and epidermic; the highest concentration of EGF in blood was found 4 hours after rhEGF administration, with a greater distribution in the plasma with regard to cellular pellet. The slowest plasma clearance corresponded to the intramuscular administration. The highest concentration of radiolabelled rhEGF was found in liver, kidney and intestine. It was found that radiolabelled EGF is excreted mainly throughout urine and faeces although other excretion pathways could exist

  1. New insights for identification of doping with recombinant human erythropoietin micro-doses after high hydration

    DEFF Research Database (Denmark)

    Martin, L.; Ashenden, M; Bejder, Jacob

    2016-01-01

    To minimize the chances of being caught after doping with recombinant human erythropoietins (rhEPO), athletes have turned to new practices using micro-doses and excess fluid ingestion to accelerate elimination and decrease the probability of detection. Our objective was to test the sensitivity...... subjects. After an injection in the evening, urine and plasma samples were collected the following morning. Half of the subjects then drank a bolus of water and new samples were collected 80 min later. Interestingly, rhEPO was detected in 100% of the samples even after water ingestion. A second similar...

  2. Enhancement of bone formation in rabbits by recombinant human growth hormone

    International Nuclear Information System (INIS)

    Ehrnberg, A.; Brosjoe, O.; Laaftman, P.; Nilsson, O.; Stroemberg, L.

    1993-01-01

    We studied the effect of human recombinant growth hormone on diaphyseal bone in 40 adult rabbits. The diaphyseal periosteum of one femur in each animal was mechanically stimulated by a nylon cerclage band. The bands induced an increase in bone formation, bone mineral content, and maximum torque capacity of the diaphyseal bone at 1 and 2 months. Growth hormone enhanced the anabolic effect of the cerclage bands on bone metabolism, evidenced by a further increase in torsional strength of the femurs. (au) (32 refs.)

  3. Frequency of intrachromosomal homologous recombination induced by UV radiation in normally repairing and excision repair-deficient human cells

    International Nuclear Information System (INIS)

    Tsujimura, T.; Maher, V.M.; McCormick, J.J.; Godwin, A.R.; Liskay, R.M.

    1990-01-01

    To investigate the role of DNA damage and nucleotide excision repair in intrachromosomal homologous recombination, a plasmid containing duplicated copies of the gene coding for hygromycin resistance was introduced into the genome of a repair-proficient human cell line, KMST-6, and two repair-deficient lines, XP2OS(SV) from xeroderma pigmentosum complementation group A and XP2YO(SV) from complementation group F. Neither hygromycin-resistance gene codes for a functional enzyme because each contains an insertion/deletion mutation at a unique site, but recombination between the two defective genes can yield hygromycin-resistant cells. The rates of spontaneous recombination in normal and xeroderma pigmentosum cell strains containing the recombination substrate were found to be similar. The frequency of UV-induced recombination was determined for three of these cell strains. At low doses, the group A cell strain and the group F cell strain showed a significant increase in frequency of recombinants. The repair-proficient cell strain required 10-to 20-fold higher doses of UV to exhibit comparable increases in frequency of recombinants. These results suggest that unexcised DNA damage, rather than the excision repair process per se, stimulates such recombination

  4. Residual endotoxin contaminations in recombinant proteins are sufficient to activate human CD1c+ dendritic cells.

    Directory of Open Access Journals (Sweden)

    Harald Schwarz

    Full Text Available Many commercially available recombinant proteins are produced in Escherichia coli, and most suppliers guarantee contamination levels of less than 1 endotoxin unit (EU. When we analysed commercially available proteins for their endotoxin content, we found contamination levels in the same range as generally stated in the data sheets, but also some that were higher. To analyse whether these low levels of contamination have an effect on immune cells, we stimulated the monocytic cell line THP-1, primary human monocytes, in vitro differentiated human monocyte-derived dendritic cells, and primary human CD1c+ dendritic cells (DCs with very low concentrations of lipopolysaccharide (LPS; ranging from 0.002-2 ng/ml. We show that CD1c+ DCs especially can be activated by minimal amounts of LPS, equivalent to the levels of endotoxin contamination we detected in some commercially available proteins. Notably, the enhanced endotoxin sensitivity of CD1c+ DCs was closely correlated with high CD14 expression levels observed in CD1c+ DCs that had been maintained in cell culture medium for 24 hours. When working with cells that are particularly sensitive to LPS, even low endotoxin contamination may generate erroneous data. We therefore recommend that recombinant proteins be thoroughly screened for endotoxin contamination using the limulus amebocyte lysate test, fluorescence-based assays, or a luciferase based NF-κB reporter assay involving highly LPS-sensitive cells overexpressing TLR4, MD-2 and CD14.

  5. Sensitive radioimmunoassay for detection of antibodies to recombinant human interferon-alpha A

    International Nuclear Information System (INIS)

    Palleroni, A.V.; Trown, P.W.

    1986-01-01

    A radioimmunoassay (RIA) for the detection of antibodies to recombinant human leukocyte interferon A (rHuIFN-alpha A) in human serum has been developed and validated against the standard antiviral neutralization bioassay (ANB). The assay measures the binding of 125 I-labeled rHuIFN-alpha A to immunoglobulins in serum. Aliquots of patients' sera are incubated with 125 I-rHuIFN-alpha A and the complexes formed between antibodies in the sera and the 125 I-rHuIFN-alpha A are precipitated with goat anti-human IgG serum. The radioactivity in the immune precipitate is a measure of the quantity of antibody (if present) in the serum. The sensitivity of this RIA is 5 ng of IgG/ml of serum

  6. Effect of recombinant adenovirus encoding human p53 tumor suppressor gene combined with radiation therapy on human lymphoma cells lines

    International Nuclear Information System (INIS)

    Yu Zeyang; Fan Wo; Li Dongqing; Zhu Ran; Wan Jianmei; Wang Yongqing; Wu Jinchang

    2008-01-01

    This paper analyzes the inhibitory effect and radiation sensitization of recombinant adenovirus encoding human p53 tumor suppressor gene (rAd-p53) on human lymphoma cell lines. Human lymphoma cell lines were treated with rAd-p53, radiation therapy and combined treatment, respectively. The cell growth inhibition was assessed by MTF. The cell cycle and apoptosis were detected by flow cytometry, and the p53 protein expression was detected by Western blotting. The results showed that extrinsic p53 gene have expressed to some degree, but not at high level. The role of inhibition and radiation sensitivity of rAd-p53 was not significant to human lymphoma cell lines. (authors)

  7. Finding trans-regulatory genes and protein complexes modulating meiotic recombination hotspots of human, mouse and yeast.

    Science.gov (United States)

    Wu, Min; Kwoh, Chee-Keong; Li, Xiaoli; Zheng, Jie

    2014-09-11

    The regulatory mechanism of recombination is one of the most fundamental problems in genomics, with wide applications in genome wide association studies (GWAS), birth-defect diseases, molecular evolution, cancer research, etc. Recombination events cluster into short genomic regions called "recombination hotspots". Recently, a zinc finger protein PRDM9 was reported to regulate recombination hotspots in human and mouse genomes. In addition, a 13-mer motif contained in the binding sites of PRDM9 is found to be enriched in human hotspots. However, this 13-mer motif only covers a fraction of hotspots, indicating that PRDM9 is not the only regulator of recombination hotspots. Therefore, the challenge of discovering other regulators of recombination hotspots becomes significant. Furthermore, recombination is a complex process. Hence, multiple proteins acting as machinery, rather than individual proteins, are more likely to carry out this process in a precise and stable manner. Therefore, the extension of the prediction of individual trans-regulators to protein complexes is also highly desired. In this paper, we introduce a pipeline to identify genes and protein complexes associated with recombination hotspots. First, we prioritize proteins associated with hotspots based on their preference of binding to hotspots and coldspots. Second, using the above identified genes as seeds, we apply the Random Walk with Restart algorithm (RWR) to propagate their influences to other proteins in protein-protein interaction (PPI) networks. Hence, many proteins without DNA-binding information will also be assigned a score to implicate their roles in recombination hotspots. Third, we construct sub-PPI networks induced by top genes ranked by RWR for various species (e.g., yeast, human and mouse) and detect protein complexes in those sub-PPI networks. The GO term analysis show that our prioritizing methods and the RWR algorithm are capable of identifying novel genes associated with

  8. Recombinant human thyrotropin prior to radioiodine therapy improves the size reduction of non-toxic nodular goiter: a prospective randomized double-blinded trial

    International Nuclear Information System (INIS)

    E Nielsen, Viveque; Bonnema, Steen; Hegedues, Laszlo; Grupe, Peter; Boel-Joergensen, Henrik

    2005-01-01

    Full text: Background: rh TSH increases the thyroid 131 I uptake (RAIU) and may have a role in the context of 131 I therapy of goiter. No placebo-controlled trial has yet been performed. Methods: In a double-blinded trial, 57 patients with nodular nontoxic goiter (51 F, 6 M) were randomized to receive either 0.3 mg rh TSH (n=28) or placebo (n=29) 24 h before 131 I. The thyroid dose was calculated based on thyroid size (measured by ultrasound) and RAUI at 24 h and 96 h. Thyroid size and function and patient satisfaction were monitored for 12 months. Results: At baseline the median goiter volume was 51 ml (range: 20-99 ml) in the placebo group and 59 ml (25-92 ml) in the rh TSH group (p=0.75). Three months after 131 I the goiter size was reduced to 38 ml (15-78 ml) and 43 ml (20-75 ml) in the two groups, respectively (p=0.001 within groups, p=0.96 between groups). At 12 months, the corresponding figures were 27 ml (15-82 ml) and 20 ml (6-59 ml); p=0.001 within groups compared with baseline, p=0.12 between groups. The relative goiter reduction at this time was 46 ± 22% in the placebo group, and 61 ± 15% in the rh TSH group (p=0.004). In addition to the influence of rh TSH, the magnitude of the goiter reduction correlated inversely with the initial goiter volume (p=0.019), whereas no significant correlation was found with the RAIU during therapy or with the absorbed thyroid dose. Discomfort during 131 I was reported by 10 patients in the placebo group and by 15 patients in the rh TSH group (p=0.12). Permanent hypothyroidism developed in 12% in the placebo group and in 52% in the rh TSH group (p=0.005). Patient satisfaction was generally very high without any major within group difference. Conclusion: In the first placebo-controlled double-blinded trial, we found that rh TSH prior to 131 I -therapy significantly improves thyroid size reduction by 33%, with a four-fold higher rate of hypothyroidism. These effects are, at least partially, mediated through other mechanisms than an increase in the RAIU. (author)

  9. Prestimulation with Recombinant Human Thyrotropin (rhTSH) Improves the Long-Term Outcome of Radioiodine Therapy for Multinodular Nontoxic Goiter

    DEFF Research Database (Denmark)

    Fast, Søren; Nielsen, Viveque Egsgaard; Grupe, Peter

    2012-01-01

    goiter volume reduction with rhTSH-augmented (131)I therapy improves the long-term reduction in goiter-related symptoms and reduces the need for additional therapy compared with plain (131)I therapy. Overall patient satisfaction is benefited, despite a higher rate of permanent hypothyroidism....

  10. Identification of the ligand-binding subunit of the human 5-hydroxytryptamine1A receptor with N-(p-azido-m-[125I] iodophenethyl)spiperone, a high affinity radioiodinated photoaffinity probe

    International Nuclear Information System (INIS)

    Raymond, J.R.; Fargin, A.; Lohse, M.J.; Regan, J.W.; Senogles, S.E.; Lefkowitz, R.J.; Caron, M.G.

    1989-01-01

    The ligand-binding subunit of the human 5-hydroxytryptamine1A (5-HT1A) receptor transiently expressed in COS-7 cells and of the native human 5-HT1A receptor derived from hippocampus and frontal cortex were identified by photoaffinity labeling with N-(p-azido-m-[125I]iodophenethyl)spiperone [( 125I]N3-NAPS), previously characterized as a high affinity radioiodinated D2-dopamine receptor probe. The identity of the ligand-binding subunit was confirmed by immunoprecipitation with an antipeptide rabbit antiserum, JWR21, raised against a synthetic peptide derived from the predicted amino acid sequence of the putative third intracellular loop of the human 5-HT1A receptor. In transiently transfected COS-7 cells expressing 14 +/- 3 pmol/mg of protein human 5-HT1A receptors, a single broad 75-kDa band was photoaffinity labeled by [125I]N3-NAPS. This band displayed the expected pharmacology of the 5-HT1A receptor, as evidenced by the ability of a series of competing ligands to block [125I]N3-NAPS photoincorporation. Moreover, antiserum JWR21 specifically and quantitatively immunoprecipitated the 75-kDa photoaffinity-labeled band from a soluble extract of the transfected COS-7 cell membranes, further confirming its identity. Finally, utilizing a combination of photoaffinity labeling and immunoprecipitation, the native ligand-binding subunit of 62-64 kDa was identified in human hippocampus and frontal cortex. The availability of the high specific activity, high affinity, photoaffinity ligand [125I]N3-NAPS and of a potent immunoprecipitating antiserum (JWR21) should greatly facilitate the biochemical characterization of the human 5-HT1A receptor

  11. Data set for mass spectrometric analysis of recombinant human serum albumin from various expression systems

    Directory of Open Access Journals (Sweden)

    Daryl G.S. Smith

    2015-09-01

    Full Text Available Human serum albumin (HSA is a versatile and important protein for the pharmaceutical industry (Fanali et al., Mol. Aspects Med. 33(3 (2012 209–290. Due to the potential transmission of pathogens from plasma sourced albumin, numerous expression systems have been developed to produce recombinant HSA (rHSA (Chen et al., Biochim. Biophys. Acta (BBA—Gen. Subj. 1830(12 (2013 5515–5525; Kobayashi, Biologicals 34(1 (2006 55–59. Based on our previous study showing increased glycation of rHSA expressed in Asian rice (Frahm et al., J. Phys. Chem. B 116(15 (2012 4661–4670, both supplier-to-supplier and lot-to-lot variability of rHSAs from a number of expression systems were evaluated using reversed phase liquid chromatography linked with MS and MS/MS analyses. The data are associated with the research article ‘Determination of Supplier-to-Supplier and Lot-to-Lot Variability in Glycation of Recombinant Human Serum Albumin Expressed in Oryza sativa’ where further analysis of rHSA samples with additional biophysical methods can be found (Frahm et al., PLoS ONE 10(9 (2014 e109893. We determined that all rHSA samples expressed in rice showed elevated levels of arginine and lysine hexose glycation compared to rHSA expressed in yeast, suggesting that the extensive glycation of the recombinant proteins is a by-product of either the expression system or purification process and not a random occurrence.

  12. Directed Selection of Recombinant Human Monoclonal Antibodies to Herpes Simplex Virus Glycoproteins from Phage Display Libraries

    Science.gov (United States)

    Sanna, Pietro Paolo; Williamson, R. Anthony; de Logu, Alessandro; Bloom, Floyd E.; Burton, Dennis R.

    1995-07-01

    Human monoclonal antibodies have considerable potential in the prophylaxis and treatment of viral disease. However, only a few such antibodies suitable for clinical use have been produced to date. We have previously shown that large panels of human recombinant monoclonal antibodies against a plethora of infectious agents, including herpes simplex virus types 1 and 2, can be established from phage display libraries. Here we demonstrate that facile cloning of recombinant Fab fragments against specific viral proteins in their native conformation can be accomplished by panning phage display libraries against viral glycoproteins "captured" from infected cell extracts by specific monoclonal antibodies immobilized on ELISA plates. We have tested this strategy by isolating six neutralizing recombinant antibodies specific for herpes simplex glycoprotein gD or gB, some of which are against conformationally sensitive epitopes. By using defined monoclonal antibodies for the antigen-capture step, this method can be used for the isolation of antibodies to specific regions and epitopes within the target viral protein. For instance, monoclonal antibodies to a nonneutralizing epitope can be used in the capture step to clone antibodies to neutralizing epitopes, or antibodies to a neutralizing epitope can be used to clone antibodies to a different neutralizing epitope. Furthermore, by using capturing antibodies to more immunodominant epitopes, one can direct the cloning to less immunogenic ones. This method should be of value in generating antibodies to be used both in the prophylaxis and treatment of viral infections and in the characterization of the mechanisms of antibody protective actions at the molecular level.

  13. New sensitive and specific assay for human immunodeficiency virus antibodies using labeled recombinant fusion protein and time-resolved fluoroimmunoassay.

    OpenAIRE

    Siitari, H; Turunen, P; Schrimsher, J; Nunn, M

    1990-01-01

    A new, rapid method for the detection of human immunodeficiency virus type 1 (HIV-1) antibody by time-resolved fluoroimmunoassay (TR-FIA) was developed. In this assay format, microtitration strips were coated with a recombinant fusion protein, and the same protein was labeled with europium and added into the wells simultaneously with the test specimens. The recombinant fusion protein contained the HIV-1 p24 gag protein sequence that carried an insertion, near the carboxyl terminus, of a 23-am...

  14. High level of expression of recombinant human granulocyte-macrophage colony stimulating factor in transgenic rice cell suspension culture

    DEFF Research Database (Denmark)

    Shin, Yun-Ji; Hong, Shin-Young; Kwon, Tae-Ho

    2003-01-01

    Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) has been previously produced in tobacco cell suspension cultures. However, the amount of hGM-CSF accumulated in the culture medium dropped quickly from its maximum of 150 microg/L at 5 d after incubation. To overcome...... of recombinant hGM-CSF in transgenic rice cell suspension culture and protease activity of this culture medium was low compared to that of tobacco culture system....

  15. Dose-dependent acute effects of recombinant human TSH (rhTSH) on thyroid size and function. Comparison of 0.1, 0.3 and 0.9 mg of rhTSH

    DEFF Research Database (Denmark)

    Fast, Søren; Nielsen, Viveque Egsgaard; Bonnema, Steen Joop

    2009-01-01

    Context: Recombinant human TSH (rhTSH) is used to augment the effect of radioiodine therapy for nontoxic multinodular goitre. Reports of acute thyroid swelling and hyperthyroidism warrant safety studies evaluating whether these side-effects are dose-dependent. Objective: To determine the effects...... on thyroid size and function of various doses of rhTSH. Design: In nine healthy male volunteers the effect of placebo, 0.1, 0.3 and 0.9 mg of rhTSH was examined in a paired design including four consecutive study rounds. Main outcome measures: Were evaluated at baseline, 24h, 48h, 96h, 7 days and 28 days...... after rhTSH and included: Thyroid volume (TV) estimation by planimetric ultrasound, and thyroid function by serum TSH, freeT3, freeT4 and Tg levels. Results: Following placebo or 0.1 mg rhTSH the TV did not change significantly from baseline at any time. At 24 and 48 hours after administration of 0.3 mg...

  16. Treatment of dwarfism with recombinant human insulin-like growth factor-1.

    Science.gov (United States)

    Ranke, Michael B; Wölfle, Joachim; Schnabel, Dirk; Bettendorf, Markus

    2009-10-01

    The growth hormone-IGF (insulin-like growth factor) system plays a central role in hormonal growth regulation. Recombinant human (rh) growth hormone (GH) has been available since the late 1980s for replacement therapy in GH-deficient patients and for the stimulation of growth in patients with short stature of various causes. Growth promotion by GH occurs in part indirectly through the induction of IGF-1 synthesis. In primary disturbances of IGF-1 production, short stature can only be treated with recombinant human IGF-1 (rhIGF-1). rhIGF-1 was recently approved for this indication but can also be used to treat other conditions. Selective review of the literature on IGF-1 therapy, based on a PubMed search. In children with severe primary IGF-1 deficiency (a rare condition whose prevalence is less than 1:10,000), the prognosis for final height is very poor (ca. 130 cm), and IGF-1 therapy is the appropriate form of pathophysiologically based treatment. There is no alternative treatment at present. The subcutaneous administration of IGF-1 twice daily in doses of 80 to 120 microg/kg accelerates growth and increases final height by 12 to 15 cm, according to current data. There is, however, a risk of hypoglycemia, as IGF-1 has an insulin-like effect. As treatment with IGF-1 is complex, this new medication should only be prescribed, for the time being, by experienced pediatric endocrinologists and diabetologists.

  17. Functional and structural characterization of recombinant dermcidin-1L, a human antimicrobial peptide

    International Nuclear Information System (INIS)

    Lai Yuping; Peng Yifei; Zuo Yi; Li Jun; Huang Jing; Wang Linfa; Wu Zirong

    2005-01-01

    Antimicrobial peptides from human skin are an important component of the innate immune response and play a key role as a first line of defense against infections. One such peptide is the recently discovered dermcidin-1L. To better understand its mechanism and to further investigate its antimicrobial spectrum, recombinant dermcidin-1L was expressed in Escherichia coli as a fusion protein and purified by affinity chromatography. The fusion protein was cleaved by factor Xa protease to produce recombinant dermcidin-1L. Antimicrobial and hemolytic assays demonstrated that dermcidin-1L displayed microbicidal activity against several opportunistic nosocomial pathogens, but no hemolytic activity against human erythrocytes even at concentrations up to 100 μM. Structural studies performed by circular dichroism spectroscopy indicated that the secondary structure of dermcidin-1L was very flexible, and both α-helix and β-sheet structures might be required for the antimicrobial activity. Our results confirmed previous findings indicating that dermcidin-1L could have promising therapeutic potentials and shed new light on the structure-function relationship of dermcidin-1L

  18. Process development and economic evaluation of recombinant human lactoferrin expressed in rice grain.

    Science.gov (United States)

    Nandi, Somen; Yalda, Dorice; Lu, Stephen; Nikolov, Zivko; Misaki, Ryo; Fujiyama, Kazuhito; Huang, Ning

    2005-06-01

    In this paper, we show that recombinant human lactoferrin (rhLF) has been stably expressed at 0.5% brown rice flour weight for nine generations. Process development indicates that rhLF can be efficiently extracted from rice flour in 20 mM phosphate buffer (pH 7.0) containing up to 0.5 M NaCl and at a ratio of 1 kg flour to 10 L buffer. After solid/liquid separation, the extract can then be loaded directly onto an ion-exchange column and rhLF can be eluted using 0.8 M NaCl. The resulting rhLF is about 95% pure. A range of biochemical and biophysical analyses were carried out and results indicated that the purified rhLF was identical to its native human counterpart other than its glycosylation. Economic analysis shows that at 600 kg/year scale, the cash cost to produce 1 g of rhLF of pharmaceutical grade is US$ 5.90. Analysis also indicates that the expression level has profound impact on costs related to planting, milling, extraction and purification, thus high level expression of recombinant protein in plants is one of the key parameters for the success of plant made pharmaceuticals.

  19. Purification and Characterization of Recombinant Human Lysozyme from Eggs of Transgenic Chickens.

    Directory of Open Access Journals (Sweden)

    Hanyu Wu

    Full Text Available Transgenic chickens as bioreactors have several advantages, such as the simple establishment procedure, correct glycosylation profile of expressed proteins, etc. Lysozyme is widely used in food industry, livestock farming, and medical field as a replacement of antibiotics because of its antibacterial and complement system-modulating activity. In this study, we used RT-PCR, Western blot, and immunofluorescence to detect the expression of recombinant human lysozyme (rhLY in the transgenic chicken. We demonstrated that the transgene of rhLY was genetically stable across different generations. We next optimized the purification procedure of rhLY from the transgenic eggs by utilizing two steps of cation-exchange chromatography and one gel-filtration chromatography. About 6 mg rhLY with the purity exceeding 90% was obtained from ten eggs, and the purification efficiency was about 75%. The purified rhLY had similar physicochemical and biological properties in molecular mass and antibacterial activity compared to the commercial human lysozyme. Additionally, both of them exhibited thermal stability at 60°C and tolerated an extensive pH range of 2 to 11. In conclusion, our study proved that the transgenic chickens we have previously generated were genetically stable and suitable for the production of active rhLY. We also provided a pipeline for purifying the recombinant proteins from transgenic eggs, which could be useful for other studies.

  20. Long-lived radioiodine in Japanese environment

    International Nuclear Information System (INIS)

    Seki, R.; Watanabe, M.; Kurihara, K.

    2000-01-01

    The amount of long-lived radioiodine, 129 I (half-life 1.57 x 10 7 y) in the Japanese environment has been studied by measuring thyroids of humans and animals. The collected samples thyroids of (1) humans in Ibaraki Prefecture, in Kanto district, the central part of Japan, (2) cattle in Aomori Prefecture, north part of Japan, and (3) wild deer in Chiba Prefecture, in Kanto district. The measured mean isotopic ratio 129 I/ 127 I for thyroids of cattle in Aomori Prefecture is 3.5 ± 1.8 x 10 -9 . A higher value of 14 ± 5 x 10 -9 has been obtained for thyroids of wild deer in Kanto district. On the other hand, the measured ratio for human thyroids in Kanto district is 1 ± 0.2 x 10 -9 . This value is significantly lower than that of cattle thyroids in Aomori and also those reported for human thyroids in Europe and USA. The higher mean ratio for cattle thyroid in Kanto district is possibly explained by the influence of nuclear reprocessing plant. Lower mean ratio for human thyroid might be due to higher dietary intake of algae. (author)

  1. Main Strategies of Plant Expression System Glycoengineering for Producing Humanized Recombinant Pharmaceutical Proteins.

    Science.gov (United States)

    Rozov, S M; Permyakova, N V; Deineko, E V

    2018-03-01

    Most the pharmaceutical proteins are derived not from their natural sources, rather their recombinant analogs are synthesized in various expression systems. Plant expression systems, unlike mammalian cell cultures, combine simplicity and low cost of procaryotic systems and the ability for posttranslational modifications inherent in eucaryotes. More than 50% of all human proteins and more than 40% of the currently used pharmaceutical proteins are glycosylated, that is, they are glycoproteins, and their biological activity, pharmacodynamics, and immunogenicity depend on the correct glycosylation pattern. This review examines in detail the similarities and differences between N- and O-glycosylation in plant and mammalian cells, as well as the effect of plant glycans on the activity, pharmacokinetics, immunity, and intensity of biosynthesis of pharmaceutical proteins. The main current strategies of glycoengineering of plant expression systems aimed at obtaining fully humanized proteins for pharmaceutical application are summarized.

  2. [A study of recombinant human sestrin 1 and sestrin 2 proteins produced in a prokaryotic system].

    Science.gov (United States)

    Rai, N; Kumar, R; Haque, Md A; Hassan, Md I; Dey, S

    2017-01-01

    Sestrins are highly conserved stress-inducible proteins capable of suppressing the production of ROS and signalling through mTORC1. Here we report a study of human sestrin1 (sesn1) and sestrin2 (sesn2) proteins produced in a pET28^(+) vector based prokaryotic system. Mass spectrometry analysis, western blot and surface plasmon resonance (SPR) of affinity purified sesn1 and sesn2 proteins confirmed their identity; biophysical characteristics were observed using circular dichroism (CD) showing that sesn1 and sesn2 have a predominant α-helical structure. Here we describe a simple, one step purification process to purify a large amount of sestrin proteins with significant yield. Further study of recombinant human sestrins may further facilitate the understanding of their roles in eukaryotic cells.

  3. Isolation of the functional human excision repair gene ERCC5 by intercosmid recombination

    International Nuclear Information System (INIS)

    Mudgett, J.S.; MacInnes, M.A.

    1990-01-01

    The complete human nucleotide exicision repair gene ERCC5 was isolated as a functional gene on overlapping cosmids. ERCC5 corrects the excision repair deficiency of Chinese hamster ovary cell line UV135, of complementation group 5. Cosmids that contained human sequences were obtained from a UV-resistant cell line derived from UV135 cells transformed with human genomic DNA. Individually, none of the cosmids complemented the UV135 repair defect; cosmid groups were formed to represent putative human genomic regions, and specific pairs of cosmids that effectively transformed UV135 cells to UV resistance were identified. Analysis of transformants derived from the active cosmid pairs showed that the functional 32-kbp ERCC5 gene was reconstructed by homologous intercosmid recombination. The cloned human sequences exhibited 100% concordance with the locus designated genetically as ERCC5 located on human chromosome 13q. Cosmid-transformed UV135 host cells repaired cytotoxic damage to levels about 70% of normal and repaired UV-irradiated shuttle vector DNA to levels about 82% of normal

  4. Characteristics of recombinantly expressed rat and human histamine H3 receptors.

    Science.gov (United States)

    Wulff, Birgitte S; Hastrup, Sven; Rimvall, Karin

    2002-10-18

    Human and rat histamine H(3) receptors were recombinantly expressed and characterized using receptor binding and a functional cAMP assay. Seven of nine agonists had similar affinities and potencies at the rat and human histamine H(3) receptor. S-alpha-methylhistamine had a significantly higher affinity and potency at the human than rat receptor, and for 4-[(1R*,2R*)-2-(5,5-dimethyl-1-hexynyl)cyclopropyl]-1H-imidazole (Perceptin) the preference was the reverse. Only two of six antagonists had similar affinities and potencies at the human and the rat histamine H(3) receptor. Ciproxifan, thioperamide and (1R*,2R*)-trans-2-imidazol-4 ylcyclopropyl) (cyclohexylmethoxy) carboxamide (GT2394) had significantly higher affinities and potencies at the rat than at the human histamine H(3) receptor, while for N-(4-chlorobenzyl)-N-(7-pyrrolodin-1-ylheptyl)guanidine (JB98064) the preference was the reverse. All antagonists also showed potent inverse agonism properties. Iodoproxyfan, Perceptin, proxyfan and GR175737, compounds previously described as histamine H(3) receptor antagonists, acted as full or partial agonists at both the rat and the human histamine H(3) receptor. Copyright 2002 Elsevier Science B.V.

  5. Radioiodine Therapy for Graves’ Disease

    International Nuclear Information System (INIS)

    Makhdomi, B.K.

    2015-01-01

    Treatment of grave diseases include Antithyroid medications, Iodine-131 therapy and Surgery. First patient treated with radioiodine (Iodine-130) in 1941, while Iodine-131 therapy was introduced in 1946. Factors affecting treatment include Iodine uptake; Size of goitre; Length of time I-131 is retained in the gland; Homogeneity of distribution within the gland; Radiosensitivity of thyroid cells and Drug interactions. Some side effects include Transient sore throat or mild dysphagia; Exacerbation of ophthalmopathy; Radiation thyroiditis and no increase in incidence of thyroid cancer and Leukemia or other malignancies. There no harmful effect upon Health of progeny; Fertility and Reproductive history

  6. Albinism-causing mutations in recombinant human tyrosinase alter intrinsic enzymatic activity.

    Science.gov (United States)

    Dolinska, Monika B; Kovaleva, Elena; Backlund, Peter; Wingfield, Paul T; Brooks, Brian P; Sergeev, Yuri V

    2014-01-01

    Tyrosinase (TYR) catalyzes the rate-limiting, first step in melanin production and its gene (TYR) is mutated in many cases of oculocutaneous albinism (OCA1), an autosomal recessive cause of childhood blindness. Patients with reduced TYR activity are classified as OCA1B; some OCA1B mutations are temperature-sensitive. Therapeutic research for OCA1 has been hampered, in part, by the absence of purified, active, recombinant wild-type and mutant human enzymes. The intra-melanosomal domain of human tyrosinase (residues 19-469) and two OCA1B related temperature-sensitive mutants, R422Q and R422W were expressed in insect cells and produced in T. ni larvae. The short trans-membrane fragment was deleted to avoid potential protein insolubility, while preserving all other functional features of the enzymes. Purified tyrosinase was obtained with a yield of >1 mg per 10 g of larval biomass. The protein was a monomeric glycoenzyme with maximum enzyme activity at 37°C and neutral pH. The two purified mutants when compared to the wild-type protein were less active and temperature sensitive. These differences are associated with conformational perturbations in secondary structure. The intramelanosomal domains of recombinant wild-type and mutant human tyrosinases are soluble monomeric glycoproteins with activities which mirror their in vivo function. This advance allows for the structure - function analyses of different mutant TYR proteins and correlation with their corresponding human phenotypes; it also provides an important tool to discover drugs that may improve tyrosinase activity and treat OCA1.

  7. Albinism-causing mutations in recombinant human tyrosinase alter intrinsic enzymatic activity.

    Directory of Open Access Journals (Sweden)

    Monika B Dolinska

    Full Text Available Tyrosinase (TYR catalyzes the rate-limiting, first step in melanin production and its gene (TYR is mutated in many cases of oculocutaneous albinism (OCA1, an autosomal recessive cause of childhood blindness. Patients with reduced TYR activity are classified as OCA1B; some OCA1B mutations are temperature-sensitive. Therapeutic research for OCA1 has been hampered, in part, by the absence of purified, active, recombinant wild-type and mutant human enzymes.The intra-melanosomal domain of human tyrosinase (residues 19-469 and two OCA1B related temperature-sensitive mutants, R422Q and R422W were expressed in insect cells and produced in T. ni larvae. The short trans-membrane fragment was deleted to avoid potential protein insolubility, while preserving all other functional features of the enzymes. Purified tyrosinase was obtained with a yield of >1 mg per 10 g of larval biomass. The protein was a monomeric glycoenzyme with maximum enzyme activity at 37°C and neutral pH. The two purified mutants when compared to the wild-type protein were less active and temperature sensitive. These differences are associated with conformational perturbations in secondary structure.The intramelanosomal domains of recombinant wild-type and mutant human tyrosinases are soluble monomeric glycoproteins with activities which mirror their in vivo function. This advance allows for the structure - function analyses of different mutant TYR proteins and correlation with their corresponding human phenotypes; it also provides an important tool to discover drugs that may improve tyrosinase activity and treat OCA1.

  8. Evidence for Within-Host Genetic Recombination among the Human Pegiviral Strains in HIV Infected Subjects.

    Science.gov (United States)

    Wu, Haoming; Padhi, Abinash; Xu, Junqiang; Gong, Xiaoyan; Tien, Po

    2016-01-01

    The non-pathogenic Human Pegivirus (HPgV, formerly GBV-C/HGV), the most prevalent RNA virus worldwide, is known to be associated with reduced morbidity and mortality in HIV-infected individuals. Although previous studies documented its ubiquity and important role in HIV-infected individuals, little is known about the underlying genetic mechanisms that maintain high genetic diversity of HPgV within the HIV-infected individuals. To assess the within-host genetic diversity of HPgV and forces that maintain such diversity within the co-infected hosts, we performed phylogenetic analyses taking into account 229 HPgV partial E1-E2 clonal sequences representing 15 male and 8 female co-infected HIV patients from Hubei province of central China. Our results revealed the presence of eleven strongly supported clades. While nine clades belonged to genotype 3, two clades belonged to genotype 2. Additionally, four clades that belonged to genotype 3 exhibited inter-clade recombination events. The presence of clonal sequences representing multiple clades within the HIV-infected individual provided the evidence of co-circulation of HPgV strains across the region. Of the 23 patients, six patients (i.e., five males and one female) were detected to have HPgV recombinant sequences. Our results also revealed that while male patients shared the viral strains with other patients, viral strains from the female patients had restricted dispersal. Taken together, the present study revealed that multiple infections with divergent HPgV viral strains may have caused within-host genetic recombination, predominantly in male patients, and therefore, could be the major driver in shaping genetic diversity of HPgV.

  9. Recombinant human proinsulin from transgenic corn endosperm: solvent screening and extraction studies

    Directory of Open Access Journals (Sweden)

    C. S. Farinas

    2007-09-01

    Full Text Available Recombinant pharmaceutical proteins are being produced in different systems such as bacteria and mammalian cell cultures. The use of transgenic plants as bioreactors has recently arisen as an alternative system offering many practical and economic advantages. However, finding an optimum strategy for the downstream processing (DSP of recombinant proteins from plants still remains a challenge. In this work, we studied the extraction of recombinant human proinsulin (rhProinsulin produced in the endosperm of transgenic corn seeds. An efficient extraction solvent was selected and the effects of temperature, solvent-to-solid ratio, time, and impeller rotational speed on the extraction were evaluated using an experimental design. After an extraction kinetics study, temperature was further evaluated to maximize rhProinsulin concentration in the extracts and to minimize the native corn components carbohydrates, phenolic compounds, and proteins. A high efficiency condition for extracting rhProinsulin with the selected solvent - 50 mM sodium bicarbonate buffer pH 10.0 and 5 mM DTT - was an extraction time of 2 h at a solvent-to-solid ratio of 10:1 and 25º C. The maximum rhProinsulin concentration in the extracts at that condition was 18.87 mg l-1 or 0.42% of the total soluble protein. These values are within the range in which the production of pharmaceutical proteins in plants can be competitive with other expression systems. The results presented provide information for the development of an additional production platform for the hormone insulin.

  10. Human recombinant RNASET2-induced inflammatory response and connective tissue remodeling in the medicinal leech.

    Science.gov (United States)

    Baranzini, Nicolò; Pedrini, Edoardo; Girardello, Rossana; Tettamanti, Gianluca; de Eguileor, Magda; Taramelli, Roberto; Acquati, Francesco; Grimaldi, Annalisa

    2017-05-01

    In recent years, several studies have demonstrated that the RNASET2 gene is involved in the control of tumorigenicity in ovarian cancer cells. Furthermore, a role in establishing a functional cross-talk between cancer cells and the surrounding tumor microenvironment has been unveiled for this gene, based on its ability to act as an inducer of the innate immune response. Although several studies have reported on the molecular features of RNASET2, the details on the mechanisms by which this evolutionarily conserved ribonuclease regulates the immune system are still poorly defined. In the effort to clarify this aspect, we report here the effect of recombinant human RNASET2 injection and its role in regulating the innate immune response after bacterial challenge in an invertebrate model, the medicinal leech. We found that recombinant RNASET2 injection induces fibroplasias, connective tissue remodeling and the recruitment of numerous infiltrating cells expressing the specific macrophage markers CD68 and HmAIF1. The RNASET2-mediated chemotactic activity for macrophages has been further confirmed by using a consolidated experimental approach based on injection of the Matrigel biomatrice (MG) supplemented with recombinant RNASET2 in the leech body wall. One week after injection, a large number of CD68 + and HmAIF-1 + macrophages massively infiltrated MG sponges. Finally, in leeches challenged with lipopolysaccharides (LPS) or with the environmental bacteria pathogen Micrococcus nishinomiyaensis, numerous macrophages migrating to the site of inoculation expressed high levels of endogenous RNASET2. Taken together, these results suggest that RNASET2 is likely involved in the initial phase of the inflammatory response in leeches.

  11. Radioiodine uptake in inactive pulmonary tuberculosis

    International Nuclear Information System (INIS)

    Bakheet, S.M.; Powe, J.; Al Suhaibani, H.; Hammami, M.M.; Bazarbashi, M.

    1999-01-01

    Radioiodine may accumulate at sites of inflammation or infection. We have seen such accumulation in six thyroid cancer patients with a history of previously treated pulmonary tuberculosis. We also review the causes of false-positive radioiodine uptake in lung infection/inflammation. Eight foci of radioiodine uptake were seen on six iodine-123 diagnostic scans. In three foci, the uptake was focal and indistinguishable from thyroid cancer pulmonary metastases from thyroid cancer. In the remaining foci, the uptake appeared nonsegmental, linear or lobar, suggesting a false-positive finding. The uptake was unchanged, variable in appearance or non-persistent on follow-up scans and less extensive than the fibrocystic changes seen on chest radiographs. In the two patients studied, thyroid hormone level did not affect the radioiodine lung uptake and there was congruent gallium-67 uptake. None of the patients had any evidence of thyroid cancer recurrence or of reactivation of tuberculosis and only two patients had chronic intermittent chest symptoms. Severe bronchiectasis, active tuberculosis, acute bronchitis, respiratory bronchiolitis, rheumatoid arthritis-associated lung disease and fungal infection such as Allescheria boydii and aspergillosis can lead to different patterns of radioiodine chest uptake mimicking pulmonary metastases. Pulmonary scarring secondary to tuberculosis may predispose to localized radioiodine accumulation even in the absence of clinically evident active infection. False-positive radioiodine uptake due to pulmonary infection/inflammation should be considered in thyroid cancer patients prior to the diagnosis of pulmonary metastases. (orig.)

  12. Distribution of recombination hotspots in the human genome--a comparison of computer simulations with real data.

    Directory of Open Access Journals (Sweden)

    Dorota Mackiewicz

    Full Text Available Recombination is the main cause of genetic diversity. Thus, errors in this process can lead to chromosomal abnormalities. Recombination events are confined to narrow chromosome regions called hotspots in which characteristic DNA motifs are found. Genomic analyses have shown that both recombination hotspots and DNA motifs are distributed unevenly along human chromosomes and are much more frequent in the subtelomeric regions of chromosomes than in their central parts. Clusters of motifs roughly follow the distribution of recombination hotspots whereas single motifs show a negative correlation with the hotspot distribution. To model the phenomena related to recombination, we carried out computer Monte Carlo simulations of genome evolution. Computer simulations generated uneven distribution of hotspots with their domination in the subtelomeric regions of chromosomes. They also revealed that purifying selection eliminating defective alleles is strong enough to cause such hotspot distribution. After sufficiently long time of simulations, the structure of chromosomes reached a dynamic equilibrium, in which number and global distribution of both hotspots and defective alleles remained statistically unchanged, while their precise positions were shifted. This resembles the dynamic structure of human and chimpanzee genomes, where hotspots change their exact locations but the global distributions of recombination events are very similar.

  13. Distribution of Recombination Hotspots in the Human Genome – A Comparison of Computer Simulations with Real Data

    Science.gov (United States)

    Mackiewicz, Dorota; de Oliveira, Paulo Murilo Castro; Moss de Oliveira, Suzana; Cebrat, Stanisław

    2013-01-01

    Recombination is the main cause of genetic diversity. Thus, errors in this process can lead to chromosomal abnormalities. Recombination events are confined to narrow chromosome regions called hotspots in which characteristic DNA motifs are found. Genomic analyses have shown that both recombination hotspots and DNA motifs are distributed unevenly along human chromosomes and are much more frequent in the subtelomeric regions of chromosomes than in their central parts. Clusters of motifs roughly follow the distribution of recombination hotspots whereas single motifs show a negative correlation with the hotspot distribution. To model the phenomena related to recombination, we carried out computer Monte Carlo simulations of genome evolution. Computer simulations generated uneven distribution of hotspots with their domination in the subtelomeric regions of chromosomes. They also revealed that purifying selection eliminating defective alleles is strong enough to cause such hotspot distribution. After sufficiently long time of simulations, the structure of chromosomes reached a dynamic equilibrium, in which number and global distribution of both hotspots and defective alleles remained statistically unchanged, while their precise positions were shifted. This resembles the dynamic structure of human and chimpanzee genomes, where hotspots change their exact locations but the global distributions of recombination events are very similar. PMID:23776462

  14. Production of functional human insulin-like growth factor binding proteins (IGFBPs) using recombinant expression in HEK293 cells

    DEFF Research Database (Denmark)

    Wanscher, Anne Sofie Molsted; Williamson, Michael; Ebersole, Tasja Wainani

    2015-01-01

    on human proteins with therapeutic relevance is needed to design and process the next generation of protein therapeutics. In order to conduct structural and functional investigations large quantities of recombinant proteins are needed. However, finding a suitable recombinant production system for proteins...... and the final protein yields were between 1 and 12mg protein per liter culture media. The recombinant IGFBPs contained PTMs and exhibited high-affinity interactions with their natural ligands IGF-1 and IGF-2.......Insulin-like growth factor binding proteins (IGFBPs) display many functions in humans including regulation of the insulin-like growth factor (IGF) signaling pathway. The various roles of human IGFBPs make them attractive protein candidates in drug discovery. Structural and functional knowledge...

  15. Reinvestigation of the radiopharmaceutical production of radioiodinated long-chain fatty acids. Vol. 3

    Energy Technology Data Exchange (ETDEWEB)

    El-Shaboury, G; El-Kolaly, M T; Raieh, M [Labelled Compounds Department, Hot Laboratories Center, Atomic Energy Authority, Cairo (Egypt)

    1996-03-01

    An improved and simple radiopharmaceutical production technique for radioiodination of {omega} - (Br) or (I) fatty acids ({omega}) - Br or I-(C H{sub 2}){sub n} Co{sub 2} H, where n = 16 to 17 with Na{sup 13I} for myocardial imaging study is described. The technique of production is reported here based on an isotope or non - isotopic exchange reaction between the inactive halogenated fatty acids and lyophilized ethanolic solution of Na{sup 131} I (previously dispensed in 0.02 M NaOH solution) at 80 Degree C within 30 min in absolute ethanol (dehydrated and redistill at 78.5 to 80 degree C over nitrobenzene b.p. 208 to 212 degree C). Although considerable radiochemical yields have been obtained using a wide variety of organic solvents as an isotropic or non-isotropic exchange media such as acetone, methyl ethyl ketone, benzene as well as di-n-propyl ketone; certain problems due to the evaporation of these solvents during the process of purification have been observed. These problems were completely avoided by the initial dissolution of the reactants in dehydrated ethyl alcohol which facilitate direct dispensing of the final radioiodinated acids in diluted human serum albumen without any decomposition. Basically, following the radioiodination process the radioiodinated fatty acids ethanolic solution is passed through out an AgCl-impregnated filter to remove the untreated inorganic radioiodine (2 to 5%). The final pure solution (containing over 99.o% radioiodinated pure fatty acids) is dispensed in human serum albumen (4%), and then sterilized by passing it through 0.22 mum milli pore bacterial filter. The technique is being applied for the radiopharmaceutical production of radioiodinated-phenyl long-chain fatty acids such as iodo phenoxy-, and iodobenzamido-heptadecanoic acids. 3 figs.

  16. Reinvestigation of the radiopharmaceutical production of radioiodinated long-chain fatty acids. Vol. 3

    International Nuclear Information System (INIS)

    El-Shaboury, G.; El-Kolaly, M.T.; Raieh, M.

    1996-01-01

    An improved and simple radiopharmaceutical production technique for radioiodination of ω - (Br) or (I) fatty acids (ω) - Br or I-(C H 2 ) n Co 2 H, where n = 16 to 17 with Na 13I for myocardial imaging study is described. The technique of production is reported here based on an isotope or non - isotopic exchange reaction between the inactive halogenated fatty acids and lyophilized ethanolic solution of Na 131 I (previously dispensed in 0.02 M NaOH solution) at 80 Degree C within 30 min in absolute ethanol (dehydrated and redistill at 78.5 to 80 degree C over nitrobenzene b.p. 208 to 212 degree C). Although considerable radiochemical yields have been obtained using a wide variety of organic solvents as an isotropic or non-isotropic exchange media such as acetone, methyl ethyl ketone, benzene as well as di-n-propyl ketone; certain problems due to the evaporation of these solvents during the process of purification have been observed. These problems were completely avoided by the initial dissolution of the reactants in dehydrated ethyl alcohol which facilitate direct dispensing of the final radioiodinated acids in diluted human serum albumen without any decomposition. Basically, following the radioiodination process the radioiodinated fatty acids ethanolic solution is passed through out an AgCl-impregnated filter to remove the untreated inorganic radioiodine (2 to 5%). The final pure solution (containing over 99.o% radioiodinated pure fatty acids) is dispensed in human serum albumen (4%), and then sterilized by passing it through 0.22 mum milli pore bacterial filter. The technique is being applied for the radiopharmaceutical production of radioiodinated-phenyl long-chain fatty acids such as iodo phenoxy-, and iodobenzamido-heptadecanoic acids. 3 figs

  17. 78 FR 78838 - Grant of Interim Extension of the Term of U.S. Patent No. 5,496,801; Recombinant Human...

    Science.gov (United States)

    2013-12-27

    ...] Grant of Interim Extension of the Term of U.S. Patent No. 5,496,801; Recombinant Human Parathyroid...,801. FOR FURTHER INFORMATION CONTACT: Mary C. Till by telephone at (571) 272-7755; by mail marked to... No. 5,496,801. The patent claims the human biological product recombinant human parathyroid hormone...

  18. Generation of a Chinese Hamster Ovary Cell Line Producing Recombinant Human Glucocerebrosidase

    Science.gov (United States)

    Novo, Juliana Branco; Morganti, Ligia; Moro, Ana Maria; Paes Leme, Adriana Franco; Serrano, Solange Maria de Toledo; Raw, Isaias; Ho, Paulo Lee

    2012-01-01

    Impaired activity of the lysosomal enzyme glucocerebrosidase (GCR) results in the inherited metabolic disorder known as Gaucher disease. Current treatment consists of enzyme replacement therapy by administration of exogenous GCR. Although effective, it is exceptionally expensive, and patients worldwide have a limited access to this medicine. In Brazil, the public healthcare system provides the drug free of charge for all Gaucher's patients, which reaches the order of $ 84 million per year. However, the production of GCR by public institutions in Brazil would reduce significantly the therapy costs. Here, we describe a robust protocol for the generation of a cell line producing recombinant human GCR. The protein was expressed in CHO-DXB11 (dhfr−) cells after stable transfection and gene amplification with methotrexate. As expected, glycosylated GCR was detected by immunoblotting assay both as cell-associated (~64 and 59 kDa) and secreted (63–69 kDa) form. Analysis of subclones allowed the selection of stable CHO cells producing a secreted functional enzyme, with a calculated productivity of 5.14 pg/cell/day for the highest producer. Although being laborious, traditional methods of screening high-producing recombinant cells may represent a valuable alternative to generate expensive biopharmaceuticals in countries with limited resources. PMID:23091360

  19. Generation of a Chinese Hamster Ovary Cell Line Producing Recombinant Human Glucocerebrosidase

    Directory of Open Access Journals (Sweden)

    Juliana Branco Novo

    2012-01-01

    Full Text Available Impaired activity of the lysosomal enzyme glucocerebrosidase (GCR results in the inherited metabolic disorder known as Gaucher disease. Current treatment consists of enzyme replacement therapy by administration of exogenous GCR. Although effective, it is exceptionally expensive, and patients worldwide have a limited access to this medicine. In Brazil, the public healthcare system provides the drug free of charge for all Gaucher’s patients, which reaches the order of $ 84 million per year. However, the production of GCR by public institutions in Brazil would reduce significantly the therapy costs. Here, we describe a robust protocol for the generation of a cell line producing recombinant human GCR. The protein was expressed in CHO-DXB11 (dhfr− cells after stable transfection and gene amplification with methotrexate. As expected, glycosylated GCR was detected by immunoblotting assay both as cell-associated (~64 and 59 kDa and secreted (63–69 kDa form. Analysis of subclones allowed the selection of stable CHO cells producing a secreted functional enzyme, with a calculated productivity of 5.14 pg/cell/day for the highest producer. Although being laborious, traditional methods of screening high-producing recombinant cells may represent a valuable alternative to generate expensive biopharmaceuticals in countries with limited resources.

  20. Pichia pastoris: a recombinant microfactory for antibodies and human membrane proteins.

    Science.gov (United States)

    Gonçalves, A M; Pedro, A Q; Maia, C; Sousa, F; Queiroz, J A; Passarinha, L A

    2013-05-01

    During the last few decades, it has become evident that the compatibility of the yeast biochemical environment with the ability to process and translate the RNA transcript, along with its capacity to modify a translated protein, are relevant requirements for selecting this host cell for protein expression in several pharmaceutical and clinical applications. In particular, Pichia pastoris is used as an industrial host for recombinant protein and metabolite production, showing a powerful capacity to meet required biomolecular target production levels in high-throughput assays for functional genomics and drug screening. In addition, there is a great advantage to using P. pastoris for protein secretion, even at high molecular weights, since the recovery and purification steps are simplified owing to relatively low levels of endogenous proteins in the extracellular medium. Clearly, no single microexpression system can provide all of the desired properties for human protein production. Moreover, chemical and physical bioprocess parameters, including culture medium formulation, temperature, pH, agitation, aeration rates, induction, and feeding strategies, can highly influence product yield and quality. In order to benefit from the currently available wide range of biosynthesis strategies using P. pastoris, this mini review focuses on the developments and technological fermentation achievements, providing both a comparative and an overall integration analysis. The main aim is to highlight the relevance and versatility of the P. pastoris biosystem to the design of more cost-effective microfactories to meet the increasing demands for recombinant membrane proteins and clinical antibodies for several therapeutic applications.

  1. Construction and characterization of human rotavirus recombinant VP8* subunit parenteral vaccine candidates.

    Science.gov (United States)

    Wen, Xiaobo; Cao, Dianjun; Jones, Ronald W; Li, Jianping; Szu, Shousun; Hoshino, Yasutaka

    2012-09-21

    Two currently licensed live oral rotavirus vaccines (Rotarix® and RotaTeq®) are highly efficacious against severe rotavirus diarrhea. However, the efficacy of such vaccines in selected low-income African and Asian countries is much lower than that in middle or high-income countries. Additionally, these two vaccines have recently been associated with rare case of intussusception in vaccinated infants. We developed a novel recombinant subunit parenteral rotavirus vaccine which may be more effective in low-income countries and also avert the potential problem of intussusception. Truncated recombinant VP8* (ΔVP8*) protein of human rotavirus strain Wa P[8], DS-1 P[4] or 1076 P[6] expressed in Escherichia coli was highly soluble and was generated in high yield. Guinea pigs hyperimmunized intramuscularly with each of the ΔVP8* proteins (i.e., P[8], P[4] or P[6]) developed high levels of homotypic as well as variable levels of heterotypic neutralizing antibodies. Moreover, the selected ΔVP8* proteins when administered to mice at a clinically relevant dosage, route and schedule, elicited high levels of serum anti-VP8* IgG and/or neutralizing antibodies. Our data indicated that the ΔVP8* proteins may be a plausible additional candidate as new parenteral rotavirus vaccines. Published by Elsevier Ltd.

  2. Radioiodine in the Savannah River Site environment

    Energy Technology Data Exchange (ETDEWEB)

    Kantelo, M.V.; Bauer, L.R.; Marter, W.L.; Murphy, C.E. Jr.; Zeigler, C.C.

    1993-01-15

    Radioiodine, which is the collective term for all radioactive isotopes of the element iodine, is formed at the Savannah River Site (SRS) principally as a by-product of nuclear reactor operations. Part of the radioiodine is released to the environment during reactor and reprocessing operations at the site. The purpose of this report is to provide an introduction to radioiodine production and disposition, its status in the environment, and the radiation dose and health risks as a consequence of its release to the environment around the Savannah River Plant. A rigorous dose reconstruction study is to be completed by thee Center for Disease Control during the 1990s.

  3. Radioiodine in the Savannah River Site environment

    International Nuclear Information System (INIS)

    Kantelo, M.V.; Bauer, L.R.; Marter, W.L.; Murphy, C.E. Jr.; Zeigler, C.C.

    1993-01-01

    Radioiodine, which is the collective term for all radioactive isotopes of the element iodine, is formed at the Savannah River Site (SRS) principally as a by-product of nuclear reactor operations. Part of the radioiodine is released to the environment during reactor and reprocessing operations at the site. The purpose of this report is to provide an introduction to radioiodine production and disposition, its status in the environment, and the radiation dose and health risks as a consequence of its release to the environment around the Savannah River Plant. A rigorous dose reconstruction study is to be completed by thee Center for Disease Control during the 1990s

  4. Radioiodine uptake by plants from soils

    International Nuclear Information System (INIS)

    Sabova, T.

    1976-01-01

    The uptake and accumulation of radioiodine by wheat, maize and peas from various types of soil have been studied. The uptake depends on the type of soil, on its content of organic matter and on the amount of fertilizer. Radioiodine is mainly accumulated in the roots. Accumulation in above-ground plant parts decreases in the following order: wheat, maize, peas. Uptake was highest from humus and clay soils and lowest from black and meadow soils. Application of chloride fertilizer or carrier iodine lead to an increase of radioiodine uptake in the whole plant. (author)

  5. Beyond radioiodine: novel therapies in advanced thyroid cancer

    International Nuclear Information System (INIS)

    Haugen, Bryan R.

    2004-01-01

    Full text: Thyroid cancer is a relatively common endocrine malignancy. Fortunately, many patients do well with standard therapy including surgery and radioiodine. A minority of patients have poorly differentiated thyroid carcinoma that is unresponsive to radioiodine therapy. Redifferentiation agents that 'reprogram ' these tumors to concentrate radioiodine would be of great value in treating patients with advanced thyroid cancer. The retinoid isotretinoin is the most extensively studied of these agents. It appears that 20-40% of patients respond to isotretinoin treatment by concentration of radioiodine in metastatic tumors, but the clinical utility of this redifferentiation is still unclear. In vitro studies suggest that the retinoid receptors RARβ and RXRγ are required for this effect. Abnormal DNA methylation may be an early event in thyroid tumorigenesis and methylation of the sodium iodide symporter (NIS) may play a role in loss of iodine concentration in these tumors. Inhibitors of methylation (5-azacytidine, phenylacetate and sodium butyrate) have been shown to increase NIS expression and iodine uptake in cell culture models, but published trials in humans are not yet available. Histone acetylation is required for efficient transcription of genes necessary for differentiated function. Proteins that cause histone deacetylation inhibit gene transcription and differentiated function. Inhibitors of histone deacetylation (depsipeptide, trichostatin A) have been shown to increase NIS expression and iodine uptake in poorly differentiated and undifferentiated cell lines. Finally, commonly used agents such as thiazolidine diones (diabetes) and HMG-CoA reductase inhibitors (hypercholesterolemia) have shown promise in preliminary in vitro studies in advanced thyroid cancer cell lines. Our own work has focused on receptor-selective retinoids and thiazolidine diones as potential therapy in patients with advanced thyroid cancer based on nuclear hormone receptor

  6. Review of the recombinant human interferon gamma as an immunotherapeutic: Impacts of production platforms and glycosylation.

    Science.gov (United States)

    Razaghi, Ali; Owens, Leigh; Heimann, Kirsten

    2016-12-20

    Human interferon gamma is a cytokine belonging to a diverse group of interferons which have a crucial immunological function against mycobacteria and a wide variety of viral infections. To date, it has been approved for treatment of chronic granulomatous disease and malignant osteopetrosis, and its application as an immunotherapeutic agent against cancer is an increasing prospect. Recombinant human interferon gamma, as a lucrative biopharmaceutical, has been engineered in different expression systems including prokaryotic, protozoan, fungal (yeasts), plant, insect and mammalian cells. Human interferon gamma is commonly expressed in Escherichia coli, marketed as ACTIMMUNE ® , however, the resulting product of the prokaryotic expression system is unglycosylated with a short half-life in the bloodstream; the purification process is tedious and makes the product costlier. Other expression systems also did not show satisfactory results in terms of yields, the biological activity of the protein or economic viability. Thus, the review aims to synthesise available information from previous studies on the production of human interferon gamma and its glycosylation patterns in different expression systems, to provide direction to future research in this field. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. PRDM9 variation strongly influences recombination hot-spot activity and meiotic instability in humans.

    Science.gov (United States)

    Berg, Ingrid L; Neumann, Rita; Lam, Kwan-Wood G; Sarbajna, Shriparna; Odenthal-Hesse, Linda; May, Celia A; Jeffreys, Alec J

    2010-10-01

    PRDM9 has recently been identified as a likely trans regulator of meiotic recombination hot spots in humans and mice. PRDM9 contains a zinc finger array that, in humans, can recognize a short sequence motif associated with hot spots, with binding to this motif possibly triggering hot-spot activity via chromatin remodeling. We now report that human genetic variation at the PRDM9 locus has a strong effect on sperm hot-spot activity, even at hot spots lacking the sequence motif. Subtle changes within the zinc finger array can create hot-spot nonactivating or enhancing variants and can even trigger the appearance of a new hot spot, suggesting that PRDM9 is a major global regulator of hot spots in humans. Variation at the PRDM9 locus also influences aspects of genome instability-specifically, a megabase-scale rearrangement underlying two genomic disorders as well as minisatellite instability-implicating PRDM9 as a risk factor for some pathological genome rearrangements.

  8. The effects of genetic polymorphism on treatment response of recombinant human growth hormone.

    Science.gov (United States)

    Chen, Shi; You, Hanxiao; Pan, Hui; Zhu, Huijuan; Yang, Hongbo; Gong, Fengying; Wang, Linjie; Jiang, Yu; Yan, Chengsheng

    2017-12-06

    Recombinant human growth hormone (rhGH) has been widely used in clinical treatment of growth hormone deficiency (GHD) or non GHD since 1985 and technology have achieved a great development in different long-acting formulations. Although the mathematical models for predicting the growth hormone response could help clinicians get to an individual personalized growth dose, many patients just can't reach the target height and the growth hormone responses differed.Genetic polymorphisms may play a role in the varies of individual responses in this treatment process.This article gives an overview of the genetic polymorphisms research of growth hormone in recent years, in order to give some potential suggestion and guide for the dose titration during treatment. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  9. The Use of Recombinant Human Platelet-Derived Growth Factor for Maxillary Sinus Augmentation.

    Science.gov (United States)

    Kubota, Atsushi; Sarmiento, Hector; Alqahtani, Mohammed Saad; Llobell, Arturo; Fiorellini, Joseph P

    The maxillary sinus augmentation procedure has become a predictable treatment to regenerate bone for implant placement. The purpose of this study was to evaluate the effect of recombinant human platelet-derived growth factor BB (rhPDGF-BB) combined with a deproteinized cancellous bovine bone graft for sinus augmentation. The lateral window approach was used for maxillary sinuses with minimal residual bone. After a healing period of 4 months, dental implants were placed and then restored following a 2-month osseointegration period. The result demonstrated increased bone height and ISQ values and a 100% survival rate. This study indicates that the addition of rhPDGF-BB to deproteinized cancellous bovine bone accelerated the healing period in maxillary sinuses with minimal native bone.

  10. Recombinant human bone morphogenetic protein 2 in augmentation procedures: case reports.

    Science.gov (United States)

    Luiz, Jaques; Padovan, Luis Eduardo Marques; Claudino, Marcela

    2014-01-01

    To successfully rehabilitate edentulous patients using endosseous implants, there must be enough available bone. Several techniques have been proposed for augmentation of sites with insufficient bone volume. Although autogenous bone has long been considered the gold standard for such procedures, the limited availability of graft material and a high morbidity rate are potential disadvantages of this type of graft. An alternative is to use recombinant human bone morphogenetic protein 2 (rhBMP-2), which is able to support bone regeneration in the oral environment. These cases demonstrate the applicability of rhBMP-2 in maxillary sinus elevation and augmentation procedures in the maxilla to enable dental implant placement. The use of rhBMP-2 in alveolar augmentation procedures had several clinical benefits for these patients.

  11. Preparation and in vitro evaluation of human recombinant annexin V using for apoptosis

    International Nuclear Information System (INIS)

    Chen Daming; Beijing Univ., Beijing; Qi Benzhong; Yang Hongwei; Luo Zhifu; Zhang Jinrong; Jin Xiaohai; Jia Bing; Xie Hong; Ma Dalong

    2004-01-01

    Human recombinant Annexin V was produced by expression in E coli with high efficiency through genetic engineering. The technique procedure concerned in the temperature and time of vector expression, the basic routine and purification of proteins was established in order to obtain a large quantity of Annexin V. the results of SDS-PAGE analysis and the apoptosis detection of single cell of thymocytes of Balb/c mice using FITC-Annexin V caused by dexamethasone with availableness show that the mature Annexin V with high purity and biologic activity is obtained by ion exchange chromatography. The results of cell binding assay show that its KD is 8.53 nmol/mL and RT is 8.79 nmol/mL. (authors)

  12. Exogenous recombinant human growth hormone effects during suboptimal energy and zinc intake

    Directory of Open Access Journals (Sweden)

    Duro Debora

    2005-04-01

    Full Text Available Abstract Background Energy and Zinc (Zn deficiencies have been associated with nutritional related growth retardation as well as growth hormone (GH resistance. In this study, the relationship between suboptimal energy and/or Zn intake and growth in rats and their response to immunoreactive exogenous recombinant human GH (GHi, was determined. Results Rats treated with GHi and fed ad-libitum energy and Zn (100/100 had increased IGFBP-3 (p Conclusion These results suggest that GHi enhances weight gain in rats with suboptimal energy and Zn intake but does not modify energy expenditure or physical activity index. Suboptimal Zn intake did not exacerbate the reduced growth or decrease in energy expenditure observed with energy restriction.

  13. Radioprotection of the intestinal crypts of mice by recombinant human interleukin-1 alpha

    International Nuclear Information System (INIS)

    Wu, S.G.; Miyamoto, T.

    1990-01-01

    Recombinant human interleukin-1 alpha (rHIL-1 alpha or IL-1) protected the intestinal crypt cells of mice against X-ray-induced damage. The survival of crypt cells measured in terms of their ability to form colonies of regenerating duodenal epithelium in situ was increased when IL-1 was given either before or after irradiation. The maximum degree of radioprotection was seen when the drug was given between 13 and 25 h before irradiation. The IL-1 dose producing maximum protection was about 6.3 micrograms/kg. This is the first report indicating that the cytokine IL-1 has a radioprotective effect in the intestine. The finding suggests that IL-1 may be of potential value in preventing radiation injury to the gut in the clinic

  14. Annulated heterocyclic bioisosteres of norarecoline. Synthesis and molecular pharmacology at five recombinant human muscarinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Bräuner-Osborne, Hans; Ebert, B; Brann, M R

    1995-01-01

    = 0.011 microM), and 4d (IC50 = 0.0008 microM). Pharmacological effects (EC50 or Ki values) and intrinsic activities (per cent of maximal carbachol responses) were determined using five recombinant human mAChRs (m1-m5) and the functional assay, receptor selection and amplification technology (R...... inhibitors of the binding of tritiated quinuclidinyl benzilate (QNB), pirenzepine (PZ), and oxotremorine-M (Oxo-M) to tissue membrane preparations. In the [3H]-Oxo-M binding assay, receptor affinities in the low nanomolar range were measured for 4a (IC50 = 0.010 microM), 4b (IC50 = 0.003 microM), 4c (IC50...

  15. The pharmacokinetics, distribution and degradation of human recombinant interleukin 1 beta in normal rats

    DEFF Research Database (Denmark)

    Wogensen, L D; Welinder, B; Hejnaes, K R

    1991-01-01

    -lives of distribution (T1/2 alpha) and elimination phases (T1/2 beta) of human recombinant interleukin 1 beta (rIL-1 beta), and its tissue distribution and cellular localization by means of mono-labelled, biologically active 125I-rIL-1 beta. After intravenous (i.v.) injection, 125I-rIL-1 beta was eliminated from...... the circulation with a T1/2 alpha of 2.9 min and a T1/2 beta of 41.1 min. The central and peripheral volume of distribution was 20.7 and 19.1 ml/rat, respectively, and the metabolic clearance rate was 16.9 ml/min/kg. The kidney and liver showed the highest accumulation of tracer, and autoradiography demonstrated...

  16. A Child with Local Lipohypertrophy following Recombinant Human Growth Hormone Administration

    Directory of Open Access Journals (Sweden)

    Ilan J. N. Koppen

    2016-01-01

    Full Text Available Local lipohypertrophy due to recombinant human growth hormone (rhGH administration is a rare phenomenon. Here, we report a case of an 11-year-old girl who presented with a paraumbilical swelling, approximately one year after the start of rhGH treatment for short stature due to the presumed diagnosis of partial growth hormone insensitivity. Ultrasound imaging revealed an asymmetric distribution of subcutaneous fat tissue at the rhGH administration site, indicating local lipohypertrophy. After sparing her routine injection site and alternating other sites, the swelling disappeared within 6 months. Although the precise cause of local lipohypertrophy resulting from rhGH administration is still unclear, it might be related to the presumed diagnosis of partial growth hormone insensitivity.

  17. Recombinant human DNase I reduces the viscosity of cystic fibrosis sputum.

    Science.gov (United States)

    Shak, S; Capon, D J; Hellmiss, R; Marsters, S A; Baker, C L

    1990-12-01

    Respiratory distress and progressive lung destruction in cystic fibrosis can be attributed to bacterial persistence and the accumulation of viscous purulent secretions in the airways. More than 30 yr ago it was suggested that the large amounts of DNA in purulent secretions contribute to its viscosity and that bovine pancreatic DNase I could reduce the viscosity. To evaluate the potential clinical utility of recombinant human DNase I (rhDNase) in the treatment of cystic fibrosis, we have cloned, sequenced, and expressed rhDNase. Catalytic amounts of rhDNase greatly reduce the viscosity of purulent cystic fibrosis sputum, transforming it within minutes from a nonflowing viscous gel to a flowing liquid. The reduction in viscosity is associated with a decrease in size of DNA in the sputum. Inhalation of a rhDNase aerosol may be a simple direct approach that will help individuals with cystic fibrosis and other patients with pneumonia or bronchitis to clear their airways of purulent secretions.

  18. Effects of prolonged recombinant human erythropoietin administration on muscle membrane transport systems and metabolic marker enzymes

    DEFF Research Database (Denmark)

    Juel, C; Thomsen, J J; Rentsch, R L

    2007-01-01

    on the expression of muscle membrane transport proteins. Likewise, improvements in performance may involve upregulation of metabolic enzymes. Since Epo is known to augment performance we tested the effect of rHuEpo on some marker enzymes that are related to aerobic capacity. For these purposes eight subjects...... performance by approximately 54%. Membrane transport systems and carbonic anhydrases involved in pH regulation remained unchanged. Of the Na(+), K(+)-pump isoforms only the density of the alpha2 subunit was decreased (by 22%) after treatment. The marker enzymes cytochrom c and hexokinase remained unchanged......Adaptations to chronic hypoxia involve changes in membrane transport proteins. The underlying mechanism of this response may be related to concomitant occurring changes in erythropoietin (Epo) levels. We therefore tested the direct effects of recombinant human erythropoietin (rHuEpo) treatment...

  19. Early postnatal treatment of hypogonadotropic hypogonadism with recombinant human FSH and LH

    DEFF Research Database (Denmark)

    Main, Katharina M; Schmidt, Ida M; Toppari, Jorma

    2002-01-01

    BACKGROUND: Patients with hypogonadotropic hypogonadism may be diagnosed shortly after birth because of micropenis and cryptorchidism, combined with subnormal LH and FSH concentrations during the postnatal period. OBJECTIVE: To investigate whether treating these patients with gonadotropins postna...... observed. CONCLUSIONS: Gonadotropin treatment in an infant with hypogonadotropic hypogonadism succeeded in inducing an increase in inhibin B and testicular growth.......BACKGROUND: Patients with hypogonadotropic hypogonadism may be diagnosed shortly after birth because of micropenis and cryptorchidism, combined with subnormal LH and FSH concentrations during the postnatal period. OBJECTIVE: To investigate whether treating these patients with gonadotropins...... the normal range (0.05-0.17 IU/l and 79-112 pg/ml respectively). METHODS: From 7.9 to 13.7 months of age, the patient was treated with recombinant human LH and FSH in doses of 20 and 21.3 IU s.c. twice weekly respectively. RESULTS: During treatment concentrations of LH, FSH, inhibin B and estradiol increased...

  20. Enhanced sialylation and in vivo efficacy of recombinant human α-galactosidase through in vitro glycosylation

    Directory of Open Access Journals (Sweden)

    Youngsoo Sohn

    2013-03-01

    Full Text Available Human α-galactosidase A (GLA has been used in enzymereplacement therapy for patients with Fabry disease. Weexpressed recombinant GLA from Chinese hamster ovary cellswith very high productivity. When compared to an approvedGLA (agalsidase beta, its size and charge were found to besmaller and more neutral. These differences resulted from thelack of terminal sialic acids playing essential roles in the serumhalf-life and proper tissue targeting. Because a simplesialylation reaction was not enough to increase the sialic acidcontent, a combined reaction using galactosyltransferase,sialyltransferase, and their sugar substrates at the same timewas developed and optimized to reduce the incubation time.The product generated by this reaction had nearly the samesize, isoelectric points, and sialic acid content as agalsidasebeta. Furthermore, it had better in vivo efficacy to degrade theaccumulated globotriaosylceramide in target organs of Fabrymice compared to an unmodified version. [BMB Reports 2013;46(3: 157-162

  1. DNA fragmentation and cytotoxicity by recombinant human tumor necrosis factor in L929 fibroblast cells

    International Nuclear Information System (INIS)

    Kosaka, T.; Kuwabara, M.; Koide, F.

    1992-01-01

    Induction of cell DNA fragmentation by treatment of recombinant human Tumor Necrosis Factor alpha (rhTNF alpha) was examined by using mouse L929 cells derived from mouse fibroblast cells. The amount of DNA fragments derived from rhTNF alpha-treated cells, detected by alkaline elution technique, was smaller than that derived from X-irradiated cells. The rhTNF alpha caused the DNA fragmentation depending on its incubation time and concentration. The DNA damage caused by rhTNF alpha treatment correlated with its cytotoxicity. This result suggested that the DNA fragmentation is one of causes of cell death. The treatment with proteinase K of DNA obtained from rhTNF alpha-treated cells did not increase the amount of DNA fragmentation, which indicates that rhTNF alpha causes DNA-fragmentation but not DNA-protein cross-linking

  2. Genetic Modification in Human Pluripotent Stem Cells by Homologous Recombination and CRISPR/Cas9 System.

    Science.gov (United States)

    Xue, Haipeng; Wu, Jianbo; Li, Shenglan; Rao, Mahendra S; Liu, Ying

    2016-01-01

    Genetic modification is an indispensable tool to study gene function in normal development and disease. The recent breakthrough of creating human induced pluripotent stem cells (iPSCs) by defined factors (Takahashi et al., Cell 131:861-872, 2007) provides a renewable source of patient autologous cells that not only retain identical genetic information but also give rise to many cell types of the body including neurons and glia. Meanwhile, the rapid advancement of genome modification tools such as gene targeting by homologous recombination (Capecchi, Nat Rev Genet 6:507-512, 2005) and genome editing tools such as CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas (CRISPR-associated) system, TALENs (Transcription activator-like effector nucleases), and ZFNs (Zinc finger nucleases) (Wang et al., Cell 153:910-918, 2013; Mali et al., Science 339:823-826, 2013; Hwang et al., Nat Biotechnol 31:227-229, 2013; Friedland et al., Nat Methods 10(8):741-743, 2013; DiCarlo et al., Nucleic Acids Res 41:4336-4343, 2013; Cong et al., Science 339:819-823, 2013) has greatly accelerated the development of human genome manipulation at the molecular level. This chapter describes the protocols for making neural lineage reporter lines using homologous recombination and the CRISPR/Cas system-mediated genome editing, including construction of targeting vectors, guide RNAs, transfection into hPSCs, and selection and verification of successfully targeted clones. This method can be applied to various needs of hPSC genetic engineering at high efficiency and high reliability.

  3. Expression and purification of recombinant truncated human keratinocyte growth factor-1

    International Nuclear Information System (INIS)

    Deng Lin; Ma Jisheng; Liu Xiaoju; Wang Xiaojie; Li Xiaokun; Gong Shouliang; Wang Huiyan; Tian Haishan

    2010-01-01

    Objective: To construct the genetic engineering bacteria highly expressing 23 amino acids human keratinocyte growth factor-1 (rhKGF1 dest23 ) missing N terminal, and provide experimental data for development of new drug for treatment of oral mucositis after radiotherapy and chemotherapy. Methods: PCR was used to synthese 23 amino acids rhKGF1 dest23 missing N terminal and sumo gene fragments, and construct four kinds of recombinant prokaryotic expression vectors: pET22b-rhKGF1 dest23 , pET22b-sumo-rhKGF1 dest23 , pET3c-rhKGF1 dest23 and pET3c-sumo-rhKGF1 dest23 , then they were transformed into prokaryotic expression host bacteria: Rosetta (DE3) plysS, BL21 (DE3), BL21 (DE3) Star plysS, origima(DE3) and BL21AI, the best expression combination of plasmid and host strain of rhKGF1 dest23 protein was screened and purified by CM ion-exchange and heparin affinity chromatography and identified with Western blotting. Results: pET22b-rhKGF1 dest23 plasmid and the BL21AI host bacteria was the best combination of expression, after induced by IPTG and arabinose, the majority of recombinant protein was expressed in soluble form, accounting for about 12% of the total bacterial proteins. Its purity reached to more than 95% of the protein after two steps chromatography, then conformed with Western blotting. Conclusion: Human genetic engineering bacteria of KGF1 dest23 is successfully constructed and induced by IPTG and arabinose, then after CM weak cation exchange and heparin affinity chromatography, the purified rhKGF1 dest23 protein is obtained. (authors)

  4. Detection of radioiodine-induced cytogenetic alterations in circulating lymphocytes of thyroid patients

    Energy Technology Data Exchange (ETDEWEB)

    Kasuba, V [Inst. for Medical Recearch and Occupational Health, Zagreb (Croatia). Laboratory for Mutagenesis; Konrady, A; Koeteles, G J [Frederic Joliot-Curie National Research Institute for Radiobiology and Radiohygiene, Budapest (Hungary); Kusic, Z [Clinical Hospital Sestre Milosrdnice, Zagreb (Croatia). Dept. of Oncology and Nuclear Medicine

    1994-10-01

    Radioiodines are often used for experimental purposes and for diagnosis and therapy in clinical practice. Human population might also be exposed to radioiodines in nuclear accidents. The ionizing energy of radioiodine affects not only the thyroid where it concentrates but also other tissues, especially the lymphocytes during their circulation through and around the gland containing the radioisotopes. Therefore, it seemed to be of interest to carry out investigations concerning the cytogenetic alterations in blood lymphocytes of patients treated with iodine-131. The method of choice was the relatively easily performable micronucleus assay in cytokinesis-blocked cultures of human peripheral lymphocytes. The test was performed on blood samples of 30 patients before the radioisotope treatment and one, two and four days after, one as well as 6 and - in a few cases - 12 weeks later. The amounts of iodine-131 injected were dependent on the clinical practices to reach the therapeutic radiation doses for hyperthyroidism and adenomas and were in the range of 220 and 5180 MBq. it was observed that the micronucleus frequency increased in the treated hyperthyroid patients while in patients with toxic adenomas the radioiodine did not result in an increase or even as compared to the pretreatment values in a few cases decreased values were seen. The results suggest individual differences in radiosensitivity as well as that the frequency of cytogenetic alterations depend on the physiological or pathological conditions of the thyroid. The significance of this observation will be discussed for dose assessments by cytogenetic techniques due to internal radioiodine. (author).

  5. Purification and crystallization of human Cu/Zn superoxide dismutase recombinantly produced in the protozoan Leishmania tarentolae

    International Nuclear Information System (INIS)

    Gazdag, Emerich Mihai; Cirstea, Ion Cristian; Breitling, Reinhard; Lukeš, Julius; Blankenfeldt, Wulf; Alexandrov, Kirill

    2010-01-01

    The structures of two new crystal forms of human Cu/Zn superoxide dismutase produced in the eukaryotic expression host L. tarentolae are reported. The rapid and inexpensive production of high-quality eukaryotic proteins in recombinant form still remains a challenge in structural biology. Here, a protein-expression system based on the protozoan Leishmania tarentolae was used to produce human Cu/Zn superoxide dismutase (SOD1) in recombinant form. Sequential integration of the SOD1 expression cassettes was demonstrated to lead to a linear increase in expression levels to up to 30 mg per litre. Chromatographic purification resulted in 90% pure recombinant protein, with a final yield of 6.5 mg per litre of culture. The protein was crystallized and the structures of two new crystal forms were determined. These results demonstrate the suitability of the L. tarentolae expression system for structural research

  6. Engineering of a Potent Recombinant Lectin-Toxin Fusion Protein to Eliminate Human Pluripotent Stem Cells.

    Science.gov (United States)

    Tateno, Hiroaki; Saito, Sayoko

    2017-07-10

    The use of human pluripotent stem cells (hPSCs) such as human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) in regenerative medicine is hindered by their tumorigenic potential. Previously, we developed a recombinant lectin-toxin fusion protein of the hPSC-specific lectin rBC2LCN, which has a 23 kDa catalytic domain (domain III) of Pseudomonas aeruginosa exotoxin A (rBC2LCN-PE23). This fusion protein could selectively eliminate hPSCs following its addition to the cell culture medium. Here we conjugated rBC2LCN lectin with a 38 kDa domain of exotoxin A containing domains Ib and II in addition to domain III (PE38). The developed rBC2LCN-PE38 fusion protein could eliminate 50% of 201B7 hPSCs at a concentration of 0.003 μg/mL (24 h incubation), representing an approximately 556-fold higher activity than rBC2LCN-PE23. Little or no effect on human fibroblasts, human mesenchymal stem cells, and hiPSC-derived hepatocytes was observed at concentrations lower than 1 μg/mL. Finally, we demonstrate that rBC2LCN-PE38 selectively eliminates hiPSCs from a mixed culture of hiPSCs and hiPSC-derived hepatocytes. Since rBC2LCN-PE38 can be prepared from soluble fractions of E. coli culture at a yield of 9 mg/L, rBC2LCN-PE38 represents a practical reagent to remove human pluripotent stem cells residing in cultured cells destined for transplantation.

  7. Tissue-engineered human bioartificial muscles expressing a foreign recombinant protein for gene therapy

    Science.gov (United States)

    Powell, C.; Shansky, J.; Del Tatto, M.; Forman, D. E.; Hennessey, J.; Sullivan, K.; Zielinski, B. A.; Vandenburgh, H. H.

    1999-01-01

    Murine skeletal muscle cells transduced with foreign genes and tissue engineered in vitro into bioartificial muscles (BAMs) are capable of long-term delivery of soluble growth factors when implanted into syngeneic mice (Vandenburgh et al., 1996b). With the goal of developing a therapeutic cell-based protein delivery system for humans, similar genetic tissue-engineering techniques were designed for human skeletal muscle stem cells. Stem cell myoblasts were isolated, cloned, and expanded in vitro from biopsied healthy adult (mean age, 42 +/- 2 years), and elderly congestive heart failure patient (mean age, 76 +/- 1 years) skeletal muscle. Total cell yield varied widely between biopsies (50 to 672 per 100 mg of tissue, N = 10), but was not significantly different between the two patient groups. Percent myoblasts per biopsy (73 +/- 6%), number of myoblast doublings prior to senescence in vitro (37 +/- 2), and myoblast doubling time (27 +/- 1 hr) were also not significantly different between the two patient groups. Fusion kinetics of the myoblasts were similar for the two groups after 20-22 doublings (74 +/- 2% myoblast fusion) when the biopsy samples had been expanded to 1 to 2 billion muscle cells, a number acceptable for human gene therapy use. The myoblasts from the two groups could be equally transduced ex vivo with replication-deficient retroviral expression vectors to secrete 0.5 to 2 microg of a foreign protein (recombinant human growth hormone, rhGH)/10(6) cells/day, and tissue engineered into human BAMs containing parallel arrays of differentiated, postmitotic myofibers. This work suggests that autologous human skeletal myoblasts from a potential patient population can be isolated, genetically modified to secrete foreign proteins, and tissue engineered into implantable living protein secretory devices for therapeutic use.

  8. Radioiodine: a boon and a bane emergency preparedness during accidental release of radioiodine

    International Nuclear Information System (INIS)

    Pahuja, D.N.

    1997-01-01

    Radioiodine, can be a double edged sword and can be dangerous and lethal. It will turn out to be a bane rather than a boon, exposing millions of individuals in and far away from the side of accident across geographical borders depending upon the weather conditions. Iodine is an indispensable element because of its being a constituent of the thyroid hormones, biosynthesized and released from the thyroid gland for the growth and over all metabolic functions. This gland weighing 20-30 g in a normal human adult, is comparatively very vascular organ with 5 lit. of blood flowing through it every hour. It contains 90% of the body iodine amounting to 5000-7000 μg, in the form of iodo aminoacids

  9. Identifying recombinants in human and primate immunodeficiency virus sequence alignments using quartet scanning

    Directory of Open Access Journals (Sweden)

    Martin Darren P

    2009-04-01

    Full Text Available Abstract Background Recombination has a profound impact on the evolution of viruses, but characterizing recombination patterns in molecular sequences remains a challenging endeavor. Despite its importance in molecular evolutionary studies, identifying the sequences that exhibit such patterns has received comparatively less attention in the recombination detection framework. Here, we extend a quartet-mapping based recombination detection method to enable identification of recombinant sequences without prior specifications of either query and reference sequences. Through simulations we evaluate different recombinant identification statistics and significance tests. We compare the quartet approach with triplet-based methods that employ additional heuristic tests to identify parental and recombinant sequences. Results Analysis of phylogenetic simulations reveal that identifying the descendents of relatively old recombination events is a challenging task for all methods available, and that quartet scanning performs relatively well compared to the triplet based methods. The use of quartet scanning is further demonstrated by analyzing both well-established and putative HIV-1 recombinant strains. In agreement with recent findings, we provide evidence that the presumed circulating recombinant CRF02_AG is a 'pure' lineage, whereas the presumed parental lineage subtype G has a recombinant origin. We also demonstrate HIV-1 intrasubtype recombination, confirm the hybrid origin of SIV in chimpanzees and further disentangle the recombinant history of SIV lineages in a primate immunodeficiency virus data set. Conclusion Quartet scanning makes a valuable addition to triplet-based methods for identifying recombinant sequences without prior specifications of either query and reference sequences. The new method is available in the VisRD v.3.0 package http://www.cmp.uea.ac.uk/~vlm/visrd.

  10. The pharmacokinetics, distribution and degradation of human recombinant interleukin 1 beta in normal rats

    DEFF Research Database (Denmark)

    Reimers, J; Wogensen, L D; Welinder, B

    1991-01-01

    Based upon in vivo rat experiments it was recently suggested that interleukin 1 in the circulation may be implicated in the initial events of beta-cell destruction leading to insulin-dependent diabetes mellitus (IDDM) in humans. The aim of the present study was to estimate half-lives of distribut......Based upon in vivo rat experiments it was recently suggested that interleukin 1 in the circulation may be implicated in the initial events of beta-cell destruction leading to insulin-dependent diabetes mellitus (IDDM) in humans. The aim of the present study was to estimate half......-lives of distribution (T1/2 alpha) and elimination phases (T1/2 beta) of human recombinant interleukin 1 beta (rIL-1 beta), and its tissue distribution and cellular localization by means of mono-labelled, biologically active 125I-rIL-1 beta. After intravenous (i.v.) injection, 125I-rIL-1 beta was eliminated from.......v., intraperitoneal (i.p.) and subcutaneous (s.c.) injections, as demonstrated by high performance size exclusion chromatography, trichloracetic acid precipitation and SDS-PAGE until 5 h after tracer injection. Pre-treatment with 'cold' rIL-1 beta enhanced degradation of a subsequent injection of tracer. The route...

  11. Formation of tissue factor activity following incubation of recombinant human tissue factor apoprotein with plasma lipoproteins

    International Nuclear Information System (INIS)

    Sakai, T.; Kisiel, W.

    1990-01-01

    Incubation of recombinant human tissue factor apoprotein (Apo-TF) with human plasma decreased the recalcified clotting time of this plasma in a time-and dose-dependent manner suggesting relipidation of the Apo-TF by plasma lipoproteins. Incubation of Apo-TF with purified preparations of human very low density, low density and high density lipoproteins resulted in tissue factor activity in a clotting assay. The order of effectiveness was VLDL greater than LDL much greater than HDL. Tissue factor activity generated by incubation of a fixed amount of Apo-TF with plasma lipoproteins was lipoprotein concentration-dependent and saturable. The association of Apo-TF with lipoprotein particles was supported by gel filtration studies in which 125 I-Apo-TF coeluted with the plasma lipoprotein in the void volume of a Superose 6 column in the presence and absence of calcium ions. In addition, void-volume Apo-TF-lipoprotein fractions exhibited tissue factor activity. These results suggest that the factor VIII-bypassing activity of bovine Apo-TF observed in a canine hemophilic model may be due, in part, to its association with plasma lipoproteins and expression of functional tissue factor activity

  12. Radioiodine treatment for pediatric hyperthyroid Grave's disease.

    Science.gov (United States)

    Chao, Ma; Jiawei, Xie; Guoming, Wang; Jianbin, Liu; Wanxia, Liu; Driedger, Al; Shuyao, Zuo; Qin, Zhang

    2009-10-01

    Grave's disease (GD) is an autoimmune disease in which excessive amounts of thyroid hormones circulate in the blood. Treatment for pediatric GD includes (1) antithyroid drugs (ATD), (2) radioiodine, and (3) thyroidectomy. Yet, the optimal therapy remains controversial. We collected studies from all electronically available sources as well as from conferences held in China. All studies using radioiodine and/or ATD and/or thyroidectomy were included. Information was found on 1,874 pediatric GD patients treated with radioiodine, 1,279 patients treated with ATD and 1,362 patients treated surgically. The cure rate for radioiodine was 49.8%; the incidence of hypothyroidism, 37.8%; of relapse, 6.3%; of adverse effects, 1.55%; and of drop outs, 0.6%. These data show that radioiodine treatment is safe and effective in pediatric GD with significant lower incidence of relapse and adverse effects but significantly higher incidence of hypothyroidism as compared with both ATD and thyroidectomy. For the time being, radioiodine treatment for pediatric GD remains an excellent first-line therapy and a good second-line therapy for patients with ATD failure, severe complications, or poor compliance.

  13. Preclinical and first-in-human evaluation of PRX-105, a PEGylated, plant-derived, recombinant human acetylcholinesterase-R

    International Nuclear Information System (INIS)

    Atsmon, Jacob; Brill-Almon, Einat; Nadri-Shay, Carmit; Chertkoff, Raul; Alon, Sari; Shaikevich, Dimitri; Volokhov, Inna; Haim, Kirsten Y.; Bartfeld, Daniel; Shulman, Avidor; Ruderfer, Ilya; Ben-Moshe, Tehila; Shilovitzky, Orit; Soreq, Hermona; Shaaltiel, Yoseph

    2015-01-01

    PRX-105 is a plant-derived recombinant version of the human ‘read-through’ acetylcholinesterase splice variant (AChE-R). Its active site structure is similar to that of the synaptic variant, and it displays the same affinity towards organophosphorus (OP) compounds. As such, PRX-105 may serve as a bio-scavenger for OP pesticides and chemical warfare agents. To assess its potential use in prophylaxis and treatment of OP poisoning we conducted several preliminary tests, reported in this paper. Intravenous (IV) PRX-105 was administered to mice either before or after exposure to an OP toxin. All mice who received an IV dose of 50 nmol/kg PRX-105, 2 min before being exposed to 1.33 × LD 50 and 1.5 × LD 50 of toxin and 10 min after exposure to 1.5 × LD 50 survived. The pharmacokinetic and toxicity profiles of PRX-105 were evaluated in mice and mini-pigs. Following single and multiple IV doses (50 to 200 mg/kg) no deaths occurred and no significant laboratory and histopathological changes were observed. The overall elimination half-life (t ½ ) in mice was 994 (± 173) min. Additionally, a first-in-human study, to assess the safety, tolerability and pharmacokinetics of the compound, was conducted in healthy volunteers. The t ½ in humans was substantially longer than in mice (average 26.7 h). Despite the small number of animals and human subjects who were assessed, the fact that PRX-105 exerts a protective and therapeutic effect following exposure to lethal doses of OP, its favorable safety profile and its relatively long half-life, renders it a promising candidate for treatment and prophylaxis against OP poisoning and warrants further investigation. - Highlights: • PRX-105 is a PEGylated plant-derived recombinant human acetylcholinesterase-R. • PRX-105 is a promising bio-scavenger for organophosphorous toxins at lethal doses. • PRX-105 was shown to protect animals both prophylactically and post-poisoning. • First-in-human study exhibited its safety

  14. Preclinical and first-in-human evaluation of PRX-105, a PEGylated, plant-derived, recombinant human acetylcholinesterase-R

    Energy Technology Data Exchange (ETDEWEB)

    Atsmon, Jacob [Clinical Research Center, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University (Israel); Brill-Almon, Einat; Nadri-Shay, Carmit; Chertkoff, Raul; Alon, Sari [Protalix Biotherapeutics, Science Park, Carmiel (Israel); Shaikevich, Dimitri; Volokhov, Inna; Haim, Kirsten Y. [Clinical Research Center, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University (Israel); Bartfeld, Daniel [Protalix Biotherapeutics, Science Park, Carmiel (Israel); Shulman, Avidor, E-mail: avidors@protalix.com [Protalix Biotherapeutics, Science Park, Carmiel (Israel); Ruderfer, Ilya; Ben-Moshe, Tehila; Shilovitzky, Orit [Protalix Biotherapeutics, Science Park, Carmiel (Israel); Soreq, Hermona [Department of Biological Chemistry, Institute of Life Sciences, The Hebrew University of Jerusalem (Israel); Shaaltiel, Yoseph [Protalix Biotherapeutics, Science Park, Carmiel (Israel)

    2015-09-15

    PRX-105 is a plant-derived recombinant version of the human ‘read-through’ acetylcholinesterase splice variant (AChE-R). Its active site structure is similar to that of the synaptic variant, and it displays the same affinity towards organophosphorus (OP) compounds. As such, PRX-105 may serve as a bio-scavenger for OP pesticides and chemical warfare agents. To assess its potential use in prophylaxis and treatment of OP poisoning we conducted several preliminary tests, reported in this paper. Intravenous (IV) PRX-105 was administered to mice either before or after exposure to an OP toxin. All mice who received an IV dose of 50 nmol/kg PRX-105, 2 min before being exposed to 1.33 × LD{sub 50} and 1.5 × LD{sub 50} of toxin and 10 min after exposure to 1.5 × LD{sub 50} survived. The pharmacokinetic and toxicity profiles of PRX-105 were evaluated in mice and mini-pigs. Following single and multiple IV doses (50 to 200 mg/kg) no deaths occurred and no significant laboratory and histopathological changes were observed. The overall elimination half-life (t{sub ½}) in mice was 994 (± 173) min. Additionally, a first-in-human study, to assess the safety, tolerability and pharmacokinetics of the compound, was conducted in healthy volunteers. The t{sub ½} in humans was substantially longer than in mice (average 26.7 h). Despite the small number of animals and human subjects who were assessed, the fact that PRX-105 exerts a protective and therapeutic effect following exposure to lethal doses of OP, its favorable safety profile and its relatively long half-life, renders it a promising candidate for treatment and prophylaxis against OP poisoning and warrants further investigation. - Highlights: • PRX-105 is a PEGylated plant-derived recombinant human acetylcholinesterase-R. • PRX-105 is a promising bio-scavenger for organophosphorous toxins at lethal doses. • PRX-105 was shown to protect animals both prophylactically and post-poisoning. • First-in-human study

  15. Effects of recombinant human epidermal growth factor on the proliferation and radiation survival of human fibroblast cell lines in vitro

    International Nuclear Information System (INIS)

    Kim, Hyun Sook; Kang, Ki Mun; Na, Jae Boem; Chai, Gyu Young; Lee, Sang Wook

    2006-01-01

    To explore the effect of recombinant human EGF on the proliferation and survival of human fibroblast cell lines following irradiation. Fibroblast was originated human skin and primary cultured. The trypan blue stain assay and MTT assay were used to study the proliferative effects of EGF on human fibroblast cell lines in vitro. An incubation of fibroblasts with rhEGF for 24 hours immediately after irradiation was counted everyday. Cell cycle distributions were analyzed by FACS analysis. Number of fibroblast was significant more increased rhEGF (1.0 nM, 10 nM, 100 nM, 1,000 nM) treated cell than control after 8 Gy irradiation. Most effective dose of rhEGF was at 160 nM. These survival differences were maintained at 1 week later. Proportion of S phase was significantly increased on rhEGF treated cells. rhEGF cause increased fibroblast proliferation following irradiation. We expect that rhEGF was effective for radiation induced wound healing

  16. Novel recombinant human lactoferrin: differential activation of oxidative stress related gene expression.

    Science.gov (United States)

    Kruzel, Marian L; Actor, Jeffrey K; Zimecki, Michał; Wise, Jasen; Płoszaj, Paulina; Mirza, Shaper; Kruzel, Mark; Hwang, Shen-An; Ba, Xueqing; Boldogh, Istvan

    2013-12-01

    Lactoferrin, an iron-binding protein found in high concentrations in mammalian exocrine secretions, is an important component of the host defense system. It is also a major protein of the secondary granules of neutrophils from which is released upon activation. Due to its potential clinical utility, recombinant human lactoferrin (rhLF) has been produced in various eukaryotic expression systems; however, none of these are fully compatible with humans. Most of the biopharmaceuticals approved by the FDA for use in humans are produced in mammalian expression systems. The Chinese hamster ovary cells (CHO) have become the system of choice for proteins that require post-translational modifications, such as glycoproteins. The aim of this study was to scale-up expression and purification of rhLF in a CHO expression system, verify its glycan primary structure, and assess its biological properties in cell culture models. A stable CHO cell line producing >200mg/L of rhLF was developed and established. rhLF was purified by a single-step cation-exchange chromatography procedure. The highly homogenous rhLF has a molecular weight of approximately 80 kDa. MALDI-TOF mass spectrometric analysis revealed N-linked, partially sialylated glycans at two glycosylation sites, typical for human milk LF. This novel rhLF showed a protective effect against oxidative stress in a similar manner to its natural counterpart. In addition, rhLF revealed a modulatory effect on cellular redox via upregulation of key antioxidant enzymes. These data imply that the CHO-derived rhLF is fully compatible with the native molecule, thus it has promise for human therapeutic applications. Copyright © 2013 Elsevier B.V. All rights reserved.

  17. Generation of insulin-producing human mesenchymal stem cells using recombinant adeno-associated virus.

    Science.gov (United States)

    Kim, Jeong Hwan; Park, Si-Nae; Suh, Hwal

    2007-02-28

    The purpose of current experiment is the generation of insulin-producing human mesenchymal stem cells as therapeutic source for the cure of type 1 diabetes. Type 1 diabetes is generally caused by insulin deficiency accompanied by the destruction of islet beta-cells. In various trials for the treatment of type 1 diabetes, cell-based gene therapy using stem cells is considered as one of the most useful candidate for the treatment. In this experiment, human mesenchymal stem cells were transduced with AAV which is containing furin-cleavable human preproinsulin gene to generate insulin-producing cells as surrogate beta-cells for the type 1 diabetes therapy. In the rAAV production procedure, rAAV was generated by transfection of AD293 cells. Human mesenchymal stems cells were transduced using rAAV with a various multiplicity of infection. Transduction of recombinant AAV was also tested using beta-galactosidse expression. Cell viability was determined by using MTT assay to evaluate the toxicity of the transduction procedure. Expression and production of Insulin were tested using reverse transcriptase-polymerase chain reaction and immunocytochemistry. Secretion of human insulin and C-peptide from the cells was assayed using enzyme-linked immunosorbent assay. Production of insulin and C-peptide from the test group represented a higher increase compared to the control group. In this study, we examined generation of insulin-producing cells from mesenchymal stem cells by genetic engineering for diabetes therapy. This work might be valuable to the field of tissue engineering for diabetes treatment.

  18. Repetitive in vivo treatment with human recombinant interleukin-1 beta modifies beta-cell function in normal rats

    DEFF Research Database (Denmark)

    Wogensen, L D; Reimers, J; Nerup, J

    1992-01-01

    It is unknown whether interleukin-1 exerts a bimodal effect on Beta-cell function in vivo, and whether interleukin-1 has a diabetogenic action in normal animals. We therefore studied: (a) acute effects 2 h after an intraperitoneal bolus injection of 4 micrograms of recombinant human interleukin-1...

  19. Activation of human T cells by a tumor vaccine infected with recombinant Newcastle disease virus producing IL-2

    NARCIS (Netherlands)

    Janke, M.; Peeters, B.; Zhao, H.; Leeuw, O.; Moormann, R.J.M.; Arnold, A.; Ziouta, Y.; Fournier, P.; Schirrmacher, V.

    2008-01-01

    A new recombinant (rec) Newcastle disease virus (NDV) with incorporated human interleukin 2 (IL-2) as foreign therapeutic gene [rec(IL-2)] will be described. The foreign gene in rec(IL-2) did not affect the main features of NDV replication nor its tumor selectivity. Biologically active IL-2 was

  20. Growth regulation on human acute myeloid leukemia effects of five recombinant hematopoietic factors in a serum-free culture system

    NARCIS (Netherlands)

    Delwel, E.; Salem, M.; Pellens, C.; Dorssers, L.; Wagemaker, G.; Clark, S.; Loewenberg, B

    1988-01-01

    The response of human acute myeloid leukemia (AML) cells to the distinct hematopoietic growth factors (HGFs), ie, recombinant interleukin-3 (IL-3), granulocyte-macrophage-CSF (GM-CSF), granulocyte-CSF (G-CSF), macrophage-CSF (M-CSF), and erythropoietin (Epo) was investigated under well-defined

  1. Recombinant human tissue factor pathway inhibitor exerts anticoagulant, anti-inflammatory and antimicrobial effects in murine pneumococcal pneumonia

    NARCIS (Netherlands)

    van den Boogaard, F. E.; Brands, X.; Schultz, M. J.; Levi, M. [=Marcel M.; Roelofs, J. J. T. H.; van 't Veer, C.; van der Poll, T.

    2011-01-01

    Background: Streptococcus (S.) pneumoniae is the most common causative pathogen in community-acquired pneumonia and a major cause of sepsis. Recombinant human tissue factor pathway inhibitor (rh-TFPI) attenuates sepsis-induced coagulation and has been evaluated in clinical trials involving patients

  2. Procedure guidelines for radioiodine therapy of differentiated thyroid cancer (version 3)

    International Nuclear Information System (INIS)

    Dietlein, M.; Schicha, H.; Eschner, W.; Luster, M.; Reiners, C.; Schober, O.; Muenster Univ.

    2007-01-01

    The procedure guideline for radioiodine therapy (RIT) of differentiated thyroid cancer (version 3) is the counterpart to the procedure guideline for 131 I whole-body scintigraphy (version 3) and specify the interdisciplinary guideline for thyroid cancer of the Deutsche Krebsgesellschaft concerning the nuclear medicine part. Recommendation for ablative 131 I therapy is given for all differentiated thyroid carcinoma (DTC) >1 cm. Regarding DTC ≤1 cm 131 I ablation may be helpful in an individual constellation. Preparation for 131 I ablation requires low iodine diet for two weeks and TSH stimulation by withdrawal of thyroid hormone medication or by use of recombinant human TSH (rhTSH). The advantages of rhTSH (no symptoms of hypothyroidism, lowerblood activity) and the advantages of endogenous TSH stimulation (necessary for 131 I-therapy in patients with metastases, higher sensitivity of 131 I whole-body scan) are discussed. In most centers standard activities are used for 131 I ablation. If pretherapeutic dosimetry is planned, the diagnostic administration of 131 I should not exceed 1-10MBq, alternative tracers are 123 I or 124 I. The recommendations for contraception and family planning are harmonized with the recommendation of ATA and ETA. Regarding the best possible protection of salivary glands the evidence is insufficient to recommend a specific setting. To minimize the risk of dental caries due to xerostomia patients should use preventive strategies for dental hygiene. (orig.)

  3. Differential gene expression in human granulosa cells from recombinant FSH versus human menopausal gonadotropin ovarian stimulation protocols

    Directory of Open Access Journals (Sweden)

    Bietz Mandi G

    2010-03-01

    Full Text Available Abstract Background The study was designed to test the hypothesis that granulosa cell (GC gene expression response differs between recombinant FSH and human menopausal gonadotropin (hMG stimulation regimens. Methods Females Results After exclusions, 1736 genes exhibited differential expression between groups. Over 400 were categorized as signal transduction genes, ~180 as transcriptional regulators, and ~175 as enzymes/metabolic genes. Expression of selected genes was confirmed by RT-PCR. Differentially expressed genes included A kinase anchor protein 11 (AKAP11, bone morphogenetic protein receptor II (BMPR2, epidermal growth factor (EGF, insulin-like growth factor binding protein (IGFBP-4, IGFBP-5, and hypoxia-inducible factor (HIF-1 alpha. Conclusions Results suggest that major differences exist in the mechanism by which pure FSH alone versus FSH/LH regulate gene expression in preovulatory GC that could impact oocyte maturity and developmental competence.

  4. Paclitaxel, ifosfamide and cisplatin with granulocyte colony-stimulating factor or recombinant human interleukin 3 and granulocyte colony-stimulating factor in ovarian cancer : A feasibility study

    NARCIS (Netherlands)

    Veldhuis, GJ; Willemse, PHB; Beijnen, JH; Piersma, H; vanderGraaf, WTA; deVries, EGE; Boonstra, J.

    1997-01-01

    The tolerability and efficacy of four courses of paclitaxel and ifosfamide plus cisplatin every 3 weeks was evaluated in patients with residual or refractory ovarian cancer. Additionally, supportive haematological effects of recombinant human interleukin 3 (rhIL-3) and recombinant human granulocyte

  5. Expression and kinetic properties of a recombinant 3 alpha-hydroxysteroid/dihydrodiol dehydrogenase isoenzyme of human liver.

    Science.gov (United States)

    Deyashiki, Y; Tamada, Y; Miyabe, Y; Nakanishi, M; Matsuura, K; Hara, A

    1995-08-01

    Human liver cytosol contains multiple forms of 3 alpha-hydroxysteroid dehydrogenase and dihydrodiol dehydrogenase with hydroxysteroid dehydrogenase activity, and multiple cDNAs for the enzymes have been cloned from human liver cDNA libraries. To understand the relationship of the multiple enzyme froms to the genes, a cDNA, which has been reported to code for an isoenzyme of human liver 3 alpha-hydroxysteroid/dihydrodiol dehydrogenase, was expressed in Escherichia coli. The recombinant enzyme showed structural and functional properties almost identical to those of the isoenzyme purified from human liver. In addition, the recombinant isoenzyme efficiently reduced 5 alpha-dihydrotestosterone and 5 beta-dihydrocortisone, the known substrates of human liver 3 alpha-hydroxysteroid dehydrogenase and chlordecone reductase previously purified, which suggests that these human liver enzymes are identical. Furthermore, the steady-state kinetic data for NADP(+)-linked (S)-1-indanol oxidation by the recombinant isoenzyme were consistent with a sequential ordered mechanism in which NADP+ binds first. Phenolphthalein inhibited this isoenzyme much more potently than it did the other human liver dihydrodiol dehydrogenases, and was a competitive inhibitor (Ki = 20 nM) that bound to the enzyme-NADP+ complex.

  6. Recombinant fragment of an antibody tailored for direct radioiodination

    Czech Academy of Sciences Publication Activity Database

    Sedláček, Juraj; Fábry, Milan; Sieglová, Irena; Král, Vlastimil; Uhnáková, Bronislava; Mudra, M.; Kronrád, L.; Sawicka, A.; Mikolajczak, R.; Řezáčová, Pavlína

    2012-01-01

    Roč. 55, č. 1 (2012), s. 52-56 ISSN 0362-4803 R&D Projects: GA MPO 2A-2TP1/076; GA MŠk 1M0505 Institutional research plan: CEZ:AV0Z50520514 Keywords : I125 labelling * single-chain antibody variable fragment * tyrosine-rich polypeptide segment * fusion protein Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.240, year: 2012

  7. Human Recombinant Peptide Sponge Enables Novel, Less Invasive Cell Therapy for Ischemic Stroke

    Directory of Open Access Journals (Sweden)

    Michiyuki Miyamoto

    2018-01-01

    Full Text Available Bone marrow stromal cell (BMSC transplantation has the therapeutic potential for ischemic stroke. However, it is unclear which delivery routes would yield both safety and maximal therapeutic benefits. We assessed whether a novel recombinant peptide (RCP sponge, that resembles human collagen, could act as a less invasive and beneficial scaffold in cell therapy for ischemic stroke. BMSCs from green fluorescent protein-transgenic rats were cultured and Sprague–Dawley rats were subjected to permanent middle cerebral artery occlusion (MCAo. A BMSC-RCP sponge construct was transplanted onto the ipsilateral intact neocortex 7 days after MCAo. A BMSC suspension or vehicle was transplanted into the ipsilateral striatum. Rat motor function was serially evaluated and histological analysis was performed 5 weeks after transplantation. The results showed that BMSCs could proliferate well in the RCP sponge and the BMSC-RCP sponge significantly promoted functional recovery, compared with the vehicle group. Histological analysis revealed that the RCP sponge provoked few inflammatory reactions in the host brain. Moreover, some BMSCs migrated to the peri-infarct area and differentiated into neurons in the BMSC-RCP sponge group. These findings suggest that the RCP sponge may be a promising candidate for animal protein-free scaffolds in cell therapy for ischemic stroke in humans.

  8. Over-expression in Escherichia coli and characterization of two recombinant isoforms of human FAD synthetase

    International Nuclear Information System (INIS)

    Brizio, Carmen; Galluccio, Michele; Wait, Robin; Torchetti, Enza Maria; Bafunno, Valeria; Accardi, Rosita; Gianazza, Elisabetta; Indiveri, Cesare; Barile, Maria

    2006-01-01

    FAD synthetase (FADS) (EC 2.7.7.2) is a key enzyme in the metabolic pathway that converts riboflavin into the redox cofactor FAD. Two hypothetical human FADSs, which are the products of FLAD1 gene, were over-expressed in Escherichia coli and identified by ESI-MS/MS. Isoform 1 was over-expressed as a T7-tagged protein which had a molecular mass of 63 kDa on SDS-PAGE. Isoform 2 was over-expressed as a 6-His-tagged fusion protein, carrying an extra 84 amino acids at the N-terminal with an apparent molecular mass of 60 kDa on SDS-PAGE. It was purified near to homogeneity from the soluble cell fraction by one-step affinity chromatography. Both isoforms possessed FADS activity and had a strict requirement for MgCl 2 , as demonstrated using both spectrophotometric and chromatographic methods. The purified recombinant isoform 2 showed a specific activity of 6.8 ± 1.3 nmol of FAD synthesized/min/mg protein and exhibited a K M value for FMN of 1.5 ± 0.3 μM. This is First report on characterization of human FADS, and First cloning and over-expression of FADS from an organism higher than yeast

  9. Structural consistency analysis of recombinant and wild-type human serum albumin

    Science.gov (United States)

    Cao, Hui-Ling; Sun, Li-Hua; Liu, Li; Li, Jian; Tang, Lin; Guo, Yun-Zhu; Mei, Qi-Bing; He, Jian-Hua; Yin, Da-Chuan

    2017-01-01

    Recombinant human serum albumin (rHSA) is potential alternatives for human serum albumin (HSA) which may ease severe shortage of HSA worldwide. In theory, rHSA and HSA are the same. Structure decides function. Therefore, the 3D structural consistency analysis of rHSA and HSA is outmost importance, which is the base of their function consistency. In this paper, the crystal structures of rHSA at resolution limit of 2.22 Å and HSA at 2.30 Å were determined by X-ray diffraction (XRD), which were deposited in the Protein Data Bank (PDB) with accession codes 4G03 (rHSA) and 4G04 (HSA). The differences between rHSA and HSA were systematically analyzed from the crystallization behavior, diffraction data and three-dimensional (3D) structure. The superimposed contrasted analysis indicated that rHSA and HSA achieved a structural similarity of 99% with an r.m.s. deviation of 0.397 Å for the corresponding overall Cα atoms. In addition, the number of α-helices in the rHSA or HSA molecule was verified to be 30. As a result, rHSA can potentially replace HSA. The study provides a theoretical and experimental basis for the clinical and additional applications of rHSA. Meanwhile, it is also a good example for applications of genetic engineering.

  10. Effect of intradermal human recombinant copper-zinc superoxide dismutase on random pattern flaps in rats.

    Science.gov (United States)

    Schein, Ophir; Westreich, Melvyn; Shalom, Avshalom

    2013-09-01

    Studies have focused on enhancing flap viability using superoxide dismutase (SOD), but only a few used SOD from human origin, and most gave the compound systemically. We evaluated the ability of SOD to improve random skin flap survival using human recombinant copper-zinc superoxide dismutase (Hr-CuZnSOD) in variable doses, injected intradermally into the flap. Seventy male Sprague Dawley rats were randomly assigned into 4 groups. Cephalic random pattern flaps were elevated on their backs and intradermal injections of different dosages of Hr-CuZnSOD were given 15 minutes before surgery. Flap survival was evaluated by fluorescein fluorescence. Analysis of variance (ANOVA) and t test statistical analyses were performed. Flap survival in all treated groups was significantly better than in the controls. The beneficial effect of HR-CuZnSOD on flap survival is attained when it is given intradermally into the flap tissue. Theoretically, Hr-CuZnSOD delivered with local anesthetics used in flap elevation may be a valuable clinical tool. Copyright © 2012 Wiley Periodicals, Inc.

  11. The red queen model of recombination hotspots evolution in the light of archaic and modern human genomes.

    Directory of Open Access Journals (Sweden)

    Yann Lesecque

    2014-11-01

    Full Text Available Recombination is an essential process in eukaryotes, which increases diversity by disrupting genetic linkage between loci and ensures the proper segregation of chromosomes during meiosis. In the human genome, recombination events are clustered in hotspots, whose location is determined by the PRDM9 protein. There is evidence that the location of hotspots evolves rapidly, as a consequence of changes in PRDM9 DNA-binding domain. However, the reasons for these changes and the rate at which they occur are not known. In this study, we investigated the evolution of human hotspot loci and of PRDM9 target motifs, both in modern and archaic human lineages (Denisovan to quantify the dynamic of hotspot turnover during the recent period of human evolution. We show that present-day human hotspots are young: they have been active only during the last 10% of the time since the divergence from chimpanzee, starting to be operating shortly before the split between Denisovans and modern humans. Surprisingly, however, our analyses indicate that Denisovan recombination hotspots did not overlap with modern human ones, despite sharing similar PRDM9 target motifs. We further show that high-affinity PRDM9 target motifs are subject to a strong self-destructive drive, known as biased gene conversion (BGC, which should lead to the loss of the majority of them in the next 3 MYR. This depletion of PRDM9 genomic targets is expected to decrease fitness, and thereby to favor new PRDM9 alleles binding different motifs. Our refined estimates of the age and life expectancy of human hotspots provide empirical evidence in support of the Red Queen hypothesis of recombination hotspots evolution.

  12. The red queen model of recombination hotspots evolution in the light of archaic and modern human genomes.

    Science.gov (United States)

    Lesecque, Yann; Glémin, Sylvain; Lartillot, Nicolas; Mouchiroud, Dominique; Duret, Laurent

    2014-11-01

    Recombination is an essential process in eukaryotes, which increases diversity by disrupting genetic linkage between loci and ensures the proper segregation of chromosomes during meiosis. In the human genome, recombination events are clustered in hotspots, whose location is determined by the PRDM9 protein. There is evidence that the location of hotspots evolves rapidly, as a consequence of changes in PRDM9 DNA-binding domain. However, the reasons for these changes and the rate at which they occur are not known. In this study, we investigated the evolution of human hotspot loci and of PRDM9 target motifs, both in modern and archaic human lineages (Denisovan) to quantify the dynamic of hotspot turnover during the recent period of human evolution. We show that present-day human hotspots are young: they have been active only during the last 10% of the time since the divergence from chimpanzee, starting to be operating shortly before the split between Denisovans and modern humans. Surprisingly, however, our analyses indicate that Denisovan recombination hotspots did not overlap with modern human ones, despite sharing similar PRDM9 target motifs. We further show that high-affinity PRDM9 target motifs are subject to a strong self-destructive drive, known as biased gene conversion (BGC), which should lead to the loss of the majority of them in the next 3 MYR. This depletion of PRDM9 genomic targets is expected to decrease fitness, and thereby to favor new PRDM9 alleles binding different motifs. Our refined estimates of the age and life expectancy of human hotspots provide empirical evidence in support of the Red Queen hypothesis of recombination hotspots evolution.

  13. Recombinant human parathyroid hormone related protein 1-34 and 1-84 and their roles in osteoporosis treatment.

    Directory of Open Access Journals (Sweden)

    Hua Wang

    Full Text Available Osteoporosis is a common disorder characterized by compromised bone strength that predisposes patients to increased fracture risk. Parathyroid hormone related protein (PTHrP is one of the candidates for clinical osteoporosis treatment. In this study, GST Gene Fusion System was used to express recombinant human PTHrP (hPTHrP 1-34 and 1-84. To determine whether the recombinant hPTHrP1-34 and 1-84 can enhance renal calcium reabsorption and promote bone formation, we examined effects of recombinant hPTHrP1-34 and 1-84 on osteogenic lineage commitment in a primary bone marrow cell culture system and on osteoporosis treatment. Results revealed that both of recombinant hPTHrP1-34 and 1-84 increased colony formation and osteogenic cell differentiation and mineralization in vitro; however, the effect of recombinant hPTHrP1-84 is a little stronger than that of hPTHrP1-34. Next, ovariectomy was used to construct osteoporosis animal model (OVX to test activities of these two recombinants in vivo. HPTHrP1-84 administration elevated serum calcium by up-regulating the expression of renal calcium transporters, which resulted in stimulation of osteoblastic bone formation. These factors contributed to augmented bone mass in hPTHrP1-84 treated OVX mice but did not affect bone resorption. There was no obvious bone mass alteration in hPTHrP1-34 treated OVX mice, which may be, at least partly, associated with shorter half-life of hPTHrP1-34 compared to hPTHrP1-84 in vivo. This study implies that recombinant hPTHrP1-84 is more effective than hPTHrP1-34 to enhance renal calcium reabsorption and to stimulate bone formation in vivo.

  14. Production of functional human insulin-like growth factor binding proteins (IGFBPs) using recombinant expression in HEK293 cells.

    Science.gov (United States)

    Wanscher, Anne Sofie Molsted; Williamson, Michael; Ebersole, Tasja Wainani; Streicher, Werner; Wikström, Mats; Cazzamali, Giuseppe

    2015-04-01

    Insulin-like growth factor binding proteins (IGFBPs) display many functions in humans including regulation of the insulin-like growth factor (IGF) signaling pathway. The various roles of human IGFBPs make them attractive protein candidates in drug discovery. Structural and functional knowledge on human proteins with therapeutic relevance is needed to design and process the next generation of protein therapeutics. In order to conduct structural and functional investigations large quantities of recombinant proteins are needed. However, finding a suitable recombinant production system for proteins such as full-length human IGFBPs, still remains a challenge. Here we present a mammalian HEK293 expression method suitable for over-expression of secretory full-length human IGFBP-1 to -7. Protein purification of full-length human IGFBP-1, -2, -3 and -5 was conducted using a two-step chromatography procedure and the final protein yields were between 1 and 12mg protein per liter culture media. The recombinant IGFBPs contained PTMs and exhibited high-affinity interactions with their natural ligands IGF-1 and IGF-2. Copyright © 2014 Elsevier Inc. All rights reserved.

  15. RPA mediates recombination repair during replication stress and is displaced from DNA by checkpoint signalling in human cells

    DEFF Research Database (Denmark)

    Sleeth, Kate M; Sørensen, Claus Storgaard; Issaeva, Natalia

    2007-01-01

    The replication protein A (RPA) is involved in most, if not all, nuclear metabolism involving single-stranded DNA. Here, we show that RPA is involved in genome maintenance at stalled replication forks by the homologous recombination repair system in humans. Depletion of the RPA protein inhibited...... the formation of RAD51 nuclear foci after hydroxyurea-induced replication stalling leading to persistent unrepaired DNA double-strand breaks (DSBs). We demonstrate a direct role of RPA in homology directed recombination repair. We find that RPA is dispensable for checkpoint kinase 1 (Chk1) activation...... and that RPA directly binds RAD52 upon replication stress, suggesting a direct role in recombination repair. In addition we show that inhibition of Chk1 with UCN-01 decreases dissociation of RPA from the chromatin and inhibits association of RAD51 and RAD52 with DNA. Altogether, our data suggest a direct role...

  16. Case study on human α1-antitrypsin: Recombinant protein titers obtained by commercial ELISA kits are inaccurate

    DEFF Research Database (Denmark)

    Hansen, Henning Gram; Kildegaard, Helene Faustrup; Min Lee, Gyun

    2016-01-01

    Accurate titer determination of recombinant proteins is crucial for evaluating protein production cell lines and processes. Even though enzyme-linked immunosorbent assay (ELISA) is the most widely used assay for determining protein titer, little is known about the accuracy of commercially available...... ELISA kits. We observed that estimations of recombinant human ø1-antitrypsin (rø1AT) titer by Coomassie-stained SDS-PAGE gels did not correspond to previously obtained titers obtained by a commercially available ELISA kit. This prompted us to develop two independent quantification assays based...... on biolayer interferometry and reversed-phase high-performance liquid chromatography. We compared the rø1AT titer obtained by these assays with three different off-the-shelf ELISA kits and found that the ELISA kits led to inconsistent results. The data presented here show that recombinant protein titers...

  17. Development of a stable radioiodinating reagent to label monoclonal antibodies for radiotherapy of cancer

    International Nuclear Information System (INIS)

    Wilbur, D.S.; Hadley, S.W.; Hylarides, M.D.; Abrams, P.G.; Beaumier, P.A.; Morgan, A.C.; Reno, J.M.; Fritzberg, A.R.

    1989-01-01

    A method of radioiodinating monoclonal antibodies such that the labeled antibodies do not undergo in vivo deiodination has been studied. The method utilizes conjugation of succinimidyl para-iodobenzoate to the antibody. The iodobenzoate was radiolabeled by using an organometallic intermediate to facilitate the reaction. Thus, succinimidyl para-tri-n-butylstannylbenzoate was radiolabeled in 60-90% radiochemical yield and subsequently conjugated to the antibody in 80-90% yield. Animal biodistribution studies were carried out with two separate anti-melanoma antibodies (9.2.27 and NR-M1-05) labeled by this method, and examined in nude mice bearing human melanoma tumor xenografts. Very large differences in the localization of radioactivity were observed in the thyroids and stomachs of mice when the iodobenzoyl-labeled antibodies were compared with the same antibodies labeled using the chloramine-T method of radioiodination. Few other significant differences in the tissue distribution of the radioiodinated antibodies were seen

  18. Comparative assessment of ELISAs using recombinant saposin-like protein 2 and recombinant cathepsin L-1 from Fasciola hepatica for the serodiagnosis of human Fasciolosis.

    Directory of Open Access Journals (Sweden)

    Bruno Gottstein

    2014-06-01

    Full Text Available Two recombinant Fasciola hepatica antigens, saposin-like protein-2 (recSAP2 and cathepsin L-1 (recCL1, were assessed individually and in combination in enzyme-linked immunosorbent assays (ELISA for the specific serodiagnosis of human fasciolosis in areas of low endemicity as encountered in Central Europe. Antibody detection was conducted using ProteinA/ProteinG (PAG conjugated to alkaline phosphatase. Test characteristics as well as agreement with results from an ELISA using excretory-secretory products (FhES from adult stage liver flukes was assessed by receiver operator characteristic (ROC analysis, specificity, sensitivity, Youdens J and overall accuracy. Cross-reactivity was assessed using three different groups of serum samples from healthy individuals (n=20, patients with other parasitic infections (n=87 and patients with malignancies (n=121. The best combined diagnostic results for recombinant antigens were obtained using the recSAP2-ELISA (87% sensitivity, 99% specificity and 97% overall accuracy employing the threshold (cut-off to discriminate between positive and negative reactions that maximized Youdens J. The findings showed that recSAP2-ELISA can be used for the routine serodiagnosis of chronic fasciolosis in clinical laboratories; the use of the PAG-conjugate offers the opportunity to employ, for example, rabbit hyperimmune serum for the standardization of positive controls.

  19. Combination therapy with sivelestat and recombinant human soluble thrombomodulin for ARDS and DIC patients

    Directory of Open Access Journals (Sweden)

    Miyoshi S

    2014-09-01

    Full Text Available Seigo Miyoshi,1 Ryoji Ito,1 Hitoshi Katayama,1 Kentaro Dote,2 Mayuki Aibiki,3 Hironobu Hamada,1,4 Takafumi Okura,1 Jitsuo Higaki1 1Department of Cardiology, Pulmonology, Hypertension and Nephrology, Ehime University Graduate School of Medicine, 2Intensive Care Division, Ehime University Hospital, 3Department of Emergency and Critical Care Medicine, School of Medicine, Ehime University, Shitsukawa, Toon, Ehime, 4Department of Physical Analysis and Therapeutic Sciences, Graduate School of Biomedical and Health Sciences, Hiroshima University, Kasumi, Minami-ku, Hiroshima, Japan Background: Neutrophil elastase, alveolar thrombin generation, and fibrin deposition play crucial roles in the development of acute respiratory distress syndrome (ARDS and disseminated intravascular coagulation (DIC. However, the usefulness of combination therapy with a selective neutrophil elastase inhibitor, sivelestat, and recombinant human soluble thrombomodulin (rhTM for patients with ARDS and DIC remains unknown. Methods: We conducted a retrospective data analysis of 142 ARDS patients with DIC to assess the effects of sivelestat combined with rhTM. Patients were divided into four groups: control (no sivelestat or rhTM treatment, sivelestat treatment alone, rhTM treatment alone, and combined treatment with sivelestat and rhTM. A Cox proportional hazard model was used to assess subject mortality rates. The efficacy of these drugs was evaluated based on survival rate, number of ventilator-free days, and change in PaO2/FIO2 (P/F ratios and DIC scores before and at 7 days after a diagnosis of ARDS with DIC. Results: Multivariate analysis showed that patient age, combination therapy, gas exchange, organ failure, cause, associated disease score, and serum C-reactive protein levels were predictors of mortality for patients with ARDS and DIC. As compared with untreated controls, combination therapy significantly improved the 60-day survival rate of patients with ARDS and DIC

  20. A review of available countermeasures to reduce radioiodine transfer to milk. With special regard to the use of stable iodine

    International Nuclear Information System (INIS)

    Howard, B.

    1993-03-01

    Contamination of milk by 131 I constitutes an important potential component of the collective dose to the population after a nuclear accident, and a relatively large dose saving can be gained by applying suitable countermeasures. Stable iodine prophylaxis is recommended for humans to prevent or reduce by the thyroid of inhaled radioiodine but controls on food are more appropriate to reduce radioiodine intakes from ingestion. Therefore it is important to have effective, practical countermeasures available to prevent radioiodine contamination of milk. This review evaluates currently available countermeasures which can be used to reduce radioiodine contamination of milk. In particular, it critically assesses available data on the effectiveness of stable iodine administration. The simplest and most effective countermeasure is to provide dairy animals with uncontaminated feed, this countermeasure has the added advantage that it will be effective for other radionuclides which may be present in the fallout. Alternatively, another effective response is to process the milk into storable dairy products for an appropriate length of time to allow for physical decay. The use of additives given to ruminants to reduce radioiodine in milk should be viewed as an alternative countermeasure which could be effective, but would not be the preferred option. However, it should be considered, particularly if difficulties are anticipated with the first two options. Stable iodine administration is a practical, and feasible option which has the potential to reduce radioiodine levels in milk, by at most a factor of three and the radiation dose to the animals thyroid. However, the stable iodine supplementation at low doses, and after the ingestion of radioiodine, may lead to increases in radioiodine levels in milk. Similarly, if ruminants are already receiving high iodine intakes, stable iodine is unlikely to be effective. Currently available data is inadequate to recommend a suitable dose

  1. Cytoprotective effect of recombinant human erythropoietin produced in transgenic tobacco plants.

    Directory of Open Access Journals (Sweden)

    Farooqahmed S Kittur

    Full Text Available Asialo-erythropoietin, a desialylated form of human erythropoietin (EPO lacking hematopoietic activity, is receiving increased attention because of its broader protective effects in preclinical models of tissue injury. However, attempts to translate its protective effects into clinical practice is hampered by unavailability of suitable expression system and its costly and limit production from expensive mammalian cell-made EPO (rhuEPO(M by enzymatic desialylation. In the current study, we took advantage of a plant-based expression system lacking sialylating capacity but possessing an ability to synthesize complex N-glycans to produce cytoprotective recombinant human asialo-rhuEPO. Transgenic tobacco plants expressing asialo-rhuEPO were generated by stably co-expressing human EPO and β1,4-galactosyltransferase (GalT genes under the control of double CaMV 35S and glyceraldehyde-3-phosphate gene (GapC promoters, respectively. Plant-produced asialo-rhuEPO (asialo-rhuEPO(P was purified by immunoaffinity chromatography. Detailed N-glycan analysis using NSI-FTMS and MS/MS revealed that asialo-rhuEPO(P bears paucimannosidic, high mannose-type and complex N-glycans. In vitro cytoprotection assays showed that the asialo-rhuEPO(P (20 U/ml provides 2-fold better cytoprotection (44% to neuronal-like mouse neuroblastoma cells from staurosporine-induced cell death than rhuEPO(M (21%. The cytoprotective effect of the asialo-rhuEPO(P was found to be mediated by receptor-initiated phosphorylation of Janus kinase 2 (JAK2 and suppression of caspase 3 activation. Altogether, these findings demonstrate that plants are a suitable host for producing cytoprotective rhuEPO derivative. In addition, the general advantages of plant-based expression system can be exploited to address the cost and scalability issues related to its production.

  2. Recombinant Production of Human Aquaporin-1 to an Exceptional High Membrane Density in Saccharomyces Cerevisiae

    DEFF Research Database (Denmark)

    Bomholt, Julie; Helix Nielsen, Claus; Scharff-Poulsen, Peter

    2014-01-01

    prevented Aquaporin1 mal-folding. Bioimaging of live yeast cells revealed that recombinant Aquaporin-1 accumulated in the yeast plasma membrane. A detergent screen for solubilization revealed that CYMAL-5 was superior in solubilizing recombinant Aquaporin-1 and generated a monodisperse protein preparation...

  3. Recombinant Production of Human Aquaporin-1 to an Exceptional High Membrane Density in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Bomholt, Julie; Helix Nielsen, Claus; Scharff-Poulsen, Peter

    2013-01-01

    of the expression temperature to 15°C almost completely prevented Aquaporin-1 mal-folding. Bioimaging of live yeast cells revealed that recombinant Aquaporin-1 accumulated in the yeast plasma membrane. A detergent screen for solubilization revealed that CYMAL-5 was superior in solubilizing recombinant Aquaporin-1...

  4. Construction and characterisation of a recombinant fowlpox virus that expresses the human papilloma virus L1 protein

    Directory of Open Access Journals (Sweden)

    Zanotto Carlo

    2011-11-01

    Full Text Available Abstract Background Human papilloma virus (HPV-16 is the most prevalent high-risk mucosal genotype. Virus-like-particle (VLP-based immunogens developed recently have proven to be successful as prophylactic HPV vaccines, but are still too expensive for developing countries. Although vaccinia viruses expressing the HPV-16 L1 protein (HPV-L1 have been studied, fowlpox-based recombinants represent efficient and safer vectors for immunocompromised hosts due to their ability to elicit a complete immune response and their natural host-range restriction to avian species. Methods A new fowlpox virus recombinant encoding HPV-L1 (FPL1 was engineered and evaluated for the correct expression of HPV-L1 in vitro, using RT-PCR, immunoprecipitation, Western blotting, electron microscopy, immunofluorescence, and real-time PCR assays. Results The FPL1 recombinant correctly expresses HPV-L1 in mammalian cells, which are non-permissive for the replication of this vector. Conclusion This FPL1 recombinant represents an appropriate immunogen for expression of HPV-L1 in human cells. The final aim is to develop a safe, immunogenic, and less expensive prophylactic vaccine against HPV.

  5. Increased biological activity of deglycosylated recombinant human granulocyte/macrophage colony-stimulating factor produced by yeast or animal cells

    International Nuclear Information System (INIS)

    Moonen, P.; Mermod, J.J.; Ernst, J.F.; Hirschi, M.; DeLamarter, J.F.

    1987-01-01

    Human granulocyte/macrophage colony-stimulating factor (hGM-CSF) produced by several recombinant sources including Escherichia coli, yeast, and animal cells was studied. Recombinant animal cells produced hGM-CSF in low quantities and in multiple forms of varying size. Mammalian hGM-CSF was purified 200,000-fold using immunoaffinity and lectin chromatography. Partially purified proteins produced in yeast and mammalian cells were assayed for the effects of deglycosylation. Following enzymatic deglycosylation, immunoreactivity was measured by radioimmunoassay and biological activity was measured in vitro on responsive human primary cells. Removal of N-linked oligosaccharides from both proteins increased their immunoreactivities by 4- to 8-fold. Removal of these oligosaccharides also increased their specific biological activities about 20-fold, to reach approximately the specific activity of recombinant hGM-CSF from E. coli. The E. coli produced-protein-lacking any carbohydrate- had by far the highest specific activity observed for the recombinant hGM-CSFs

  6. Closely related glycosylation patterns of recombinant human IL-2 expressed in a CHO cell line and natural IL-2

    International Nuclear Information System (INIS)

    Vita, N.; Magazin, M.; Marchese, E.; Lupker, J.; Ferrara, P.

    1990-01-01

    We report here the study of the glycosylation pattern of human recombinant (r) IL2 expressed in a Chinese hamster ovary (CHO) cell line. The human rIL2 secreted by this high-producing recombinant CHO cell line was metabolically radiolabelled with [35S]-methionine, or with [3H]-glucosamine and [3H]-galactose, purified to homogeneity, and then characterized. The electrophoretic analysis of the [35S]-methionine-labelled proteins present in the culture medium of the CHO cell line showed that the rIL2 represents approximately 12% of the total secreted proteins. Furthermore, pulse-chase experiments showed that the glycosylated rIL2 is synthesized and secreted within 30 min. The point of attachment and the structure of the carbohydrate moiety of the rIL2 was determined by: amino-terminal sequencing and fingerprint analysis of the 3H-labelled rIL2, mass spectroscopy of the amino-terminal tryptic octapeptide, and carbohydrate analysis after enzymatic (Vibrio cholerae neuraminidase and Aspergillus oryzae beta-galactosidase) or sulfuric acid hydrolysis. The results indicate that the recombinant protein possesses a sugar moiety O-linked to the threonine residue at position 3 of the polypeptide chain, and that sialic acid, galactose and N-acetyl galactosamine are components of this carbohydrate moiety. Taken together these results suggest that the recombinant molecule is identical to natural IL2

  7. The history of radioiodine therapy beginnings

    Directory of Open Access Journals (Sweden)

    Pavel Olegovich Rumiantsev

    2016-03-01

    Full Text Available This report contains historical review of radioiodine therapy invention and clinical implementation for the treatment of patients with diffuse toxic goiter and differentiated thyroid cancer. In 1923 Henry Plummer has strongly recommended to prescribe stable iodine in patients with Grave’s diseases after surgery in order to prevent thyrotoxic storm. Radioactive isotopes of iodine for the first time were obtained by Enrico Fermi in 1934 in laboratory. Clinical implementation of radioiodine was started in 40th years of past century. In 1940 experimentally have been shown that thyroid gland with Grave’s disease accumulated as much as 80% of radioiodine administered activity. Saul Hertz was the first physician who used radioactive iodine in January, 1941 with purpose to treat diffuse toxic goiter patients. Samuel Seidlin was first who used radioiodine in March, 1943 to treat patient with functioned differentiated thyroid cancer metastases. In Russia radioiodine treatment of patients started since 1982 in Medical Radiological Research Centre (former Institute of Medical Radiology, Obninsk.

  8. Cortical bone growth and maturational changes in dwarf rats induced by recombinant human growth hormone

    Science.gov (United States)

    Martinez, D. A.; Orth, M. W.; Carr, K. E.; Vanderby, R. Jr; Vailas, A. C.

    1996-01-01

    The growth hormone (GH)-deficient dwarf rat was used to investigate recombinant human (rh) GH-induced bone formation and to determine whether rhGH facilitates simultaneous increases in bone formation and bone maturation during rapid growth. Twenty dwarf rats, 37 days of age, were randomly assigned to dwarf plus rhGH (GH; n = 10) and dwarf plus vehicle (n = 10) groups. The GH group received 1.25 mg rhGH/kg body wt two times daily for 14 days. Biochemical, morphological, and X-ray diffraction measurements were performed on the femur middiaphysis. rhGH stimulated new bone growth in the GH group, as demonstrated by significant increases (P < 0.05) in longitudinal bone length (6%), middiaphyseal cross-sectional area (20%), and the amount of newly accreted bone collagen (28%) in the total pool of middiaphyseal bone collagen. Cortical bone density, mean hydroxyapatite crystal size, and the calcium and collagen contents (microgram/mm3) were significantly smaller in the GH group (P < 0.05). Our findings suggest that the processes regulating new collagen accretion, bone collagen maturation, and mean hydroxyapatite crystal size may be independently regulated during rapid growth.

  9. Inactivation of purified human recombinant monoamine oxidases A and B by rasagiline and its analogues.

    Science.gov (United States)

    Hubálek, Frantisek; Binda, Claudia; Li, Min; Herzig, Yaacov; Sterling, Jeffrey; Youdim, Moussa B H; Mattevi, Andrea; Edmondson, Dale E

    2004-03-25

    The inactivation of purified human recombinant monoamine oxidases (MAO) A and B by rasagiline [N-propargyl-1(R)-aminoindan] and four of its analogues [N-propargyl-1(S)-aminoindan (S-PAI), 6-hydroxy-N-propargyl-1(R)-aminoindan (R-HPAI), N-methyl-N-propargyl-1(R)-aminoindan (R-MPAI), and 6-(N-methyl-N-ethyl carbamoyloxy)-N-propargyl-1(R)-aminoindan (R-CPAI)] has been investigated. All compounds tested, with the exception of R-CPAI, form stoichiometric N(5) flavocyanine adducts with the FAD moiety of either enzyme. No H(2)O(2) is produced during either MAO A or MAO B inactivation, which demonstrates that covalent addition occurs in a single turnover. Rasagiline has the highest specificity for MAO B, as demonstrated by a 100-fold higher inhibition potency (k(inact)/K(i)) compared to MAO A, with the remaining compounds exhibiting lower isozyme specificities. MAO B and MAO A are more selective for the R-enantiomer (rasagiline) compared to the S-enantiomer (S-PAI) by 2500-fold and 17-fold, respectively. Differences in UV/vis and CD spectral data of the complexes of the studied compounds with both MAO A and MAO B are interpreted in light of crystallographic data of complexes of MAO B with rasagiline and its analogues (Binda, C.; et al. J. Med. Chem. 2004, 47, 1767-1774.

  10. A recombinant dromedary antibody fragment (VHH or nanobody) directed against human Duffy antigen receptor for chemokines.

    Science.gov (United States)

    Smolarek, Dorota; Hattab, Claude; Hassanzadeh-Ghassabeh, Gholamreza; Cochet, Sylvie; Gutiérrez, Carlos; de Brevern, Alexandre G; Udomsangpetch, Rachanee; Picot, Julien; Grodecka, Magdalena; Wasniowska, Kazimiera; Muyldermans, Serge; Colin, Yves; Le Van Kim, Caroline; Czerwinski, Marcin; Bertrand, Olivier

    2010-10-01

    Fy blood group antigens are carried by the Duffy antigen receptor for chemokines (DARC), a red cells receptor for Plasmodium vivax broadly implicated in human health and diseases. Recombinant VHHs, or nanobodies, the smallest intact antigen binding fragment derivative from the heavy chain-only antibodies present in camelids, were prepared from a dromedary immunized against DARC N-terminal extracellular domain and selected for DARC binding. A described VHH, CA52, does recognize native DARC on cells. It inhibits P. vivax invasion of erythrocytes and displaces interleukin-8 bound to DARC. The targeted epitope overlaps the well-defined DARC Fy6 epitope. K (D) of CA52-DARC equilibrium is sub-nanomolar, hence ideal to develop diagnostic or therapeutic compounds. Immunocapture by immobilized CA52 yielded highly purified DARC from engineered K562 cells. This first report on a VHH with specificity for a red blood cell protein exemplifies VHHs' potentialities to target, to purify, and to modulate the function of cellular markers.

  11. Whole-body irradiation transiently diminishes the adrenocorticotropin response to recombinant human interleukin-1{alpha}

    Energy Technology Data Exchange (ETDEWEB)

    Perlstein, R.S.; Mehta, N.R.; Neta, R.; Whitnall, M.H. [Armed Forces Radiobiology Research Institute, Bethesda, MD (United States); Mougey, E.H. [Walter Reed Army Institute of Research, Washington, DC (United States)

    1995-03-01

    Recombinant human interleukin-1{alpha} (rhIL-1{alpha}) has significant potential as a radioprotector and/or treatment for radiation-induced hematopoietic injury. Both IL-1 and whole-body ionizing irradiation acutely stimulate the hypothalamic-pituitary-adrenal axis. We therefore assessed the interaction of whole-body irradiation and rhIL-1{alpha} in altering the functioning of the axis in mice. Specifically, we determined the adrenocorticotropin (ACTH) and corticosterone responses to rhIL-1{alpha} administered just before and hours to days after whole-body or sham irradiation. Our results indicate that whole-body irradiation does not potentiate the rhIL-1{alpha}-induced increase in ACTH levels at the doses used. In fact, the rhIL-1{alpha}-induced increase in plasma ACTH is transiently impaired when the cytokine is administered 5 h after, but not 1 h before, exposure to whole-body irradiation. The ACTH response may be inhibited by elevated corticosterone levels after whole-body irradiation, or by other radiation-induced effects on the pituitary gland and hypothalamus. 36 refs., 3 figs.

  12. Height Outcome of Recombinant Human Growth Hormone Treatment in Achondroplasia Children: A Meta-Analysis.

    Science.gov (United States)

    Miccoli, Mario; Bertelloni, Silvano; Massart, Francesco

    2016-01-01

    Although recombinant human growth hormone (rhGH) is not approved to treat short stature of achondroplasia (ACH), some studies suggested growth improvement during short-term rhGH treatment. A meta-analysis of rhGH therapy efficacy in ACH children was performed. From 12 English-language studies, 558 (54.0% males) rhGH-treated ACH children were enrolled. Administration of rhGH (median dosage 0.21 mg/kg/ week; range 0.16-0.42 mg/kg/week) improved height (Ht) from baseline [-5.069 standard deviation score (SDS; 95% CI -5.109 to -5.029); p < 0.0001] to 12 [-4.325 SDS (95% CI -4.363 to -4.287); p < 0.0001] and 24 months [-4.073 SDS (95% CI -4.128 to -4.019); p < 0.0001]. Then, Ht remained approximately constant up to 5 years [-3.941 SDS (95% CI -4.671 to -3.212); p < 0.0001]. In ACH children, rhGH treatment increased Ht from -5.0 to -4.0 SDS during 5 years, but insufficient data are available on both the adult Ht and the changes of body proportions. © 2016 S. Karger AG, Basel.

  13. Recombinant human serum albumin hydrogel as a novel drug delivery vehicle

    International Nuclear Information System (INIS)

    Hirose, Masaaki; Tachibana, Akira; Tanabe, Toshizumi

    2010-01-01

    Serum albumin acts as a physiological carrier for various compounds including drugs. A hydrogel consisting of recombinant human serum albumin (rHSA) was prepared to take advantage of drug binding ability of albumin for a sustained drug release carrier. The hydrogel was prepared by mixing rHSA and dithiothreitol and casted to a polystyrene mold. Hydrogel formation was thought to occur through the intermolecular interaction of the hydrophobic groups by protein denaturation. The release of sodium benzoate and salicylic acid from the hydrogel completed in 2 h, while warfarin release continued for 24 h. The total amounts of the drugs released from 100 mg of 15 and 5% rHSA hydrogel were 2.3 and 1.4 μmol for warfarin, 1.4 and 1.1 μmol for salicylic acid and 0.9 and 0.9 μmol for sodium benzoate. These results reflected the order of the binding ability of drugs for intact albumin indicating that the drug binding ability of HSA still remained after the hydrogel formation. However, fibroblast cells attached and proliferated well on the hydrogel, indicating that denaturation of rHSA proceeded to the extent to allow the cell attachment. The present rHSA hydrogel might be suitable for a sustained release carrier of drugs having affinity for albumin.

  14. Inhibition of the recombinant human endostatin adenavirus on experimental choroidal neovascularization in rats

    Directory of Open Access Journals (Sweden)

    Li-Juan Chen

    2017-06-01

    Full Text Available AIM: To investigate the inhibition of the recombinant human endostatin adenavirus(Ad-Eson the experimental choroidal neovascularization(CNVmodels by intravitreous injection. METHODS: Experimental CNV models were induced by semiconductor laser in 30 male Brown Norway(BNrats and randomly divided into 3 groups with 10 rats in each group. At 21d after photocoagulation, the single administration group were given intravitreous injection with Ad-Es 0.01mL; the repeated administration group were given intravitreous injection with Ad-Es 0.01mL and a repeated injection 7d later; the saline control group were given intravitreous injection with saline 0.01mL. At 7d after final administration, the leakage of fundus fluorescein angiography(FFAwas observed. Various CNV areas were measured by using laser confocal microscopy of choroidal flatmount method. Pathology and ultrastructure were observed with light microscopy, the expressions of CD105 were measured by immunohistochemistry. RESULTS: The leakage of CNV of the administration group abviously decreased as compared with those in the saline group, the leakage of repeated administration group decreased compared with that of single administration group(PPCONCLUSION: Ad-Es can effectively inhibit semiconductor laser induced CNV in BN rats, and the inhibition effect of repeated administration group is better than that of single administration group. It may be a useful new method in the treatment of CNV.

  15. Quadrivalent human papillomavirus recombinant vaccine: The first vaccine for cervical cancers

    Directory of Open Access Journals (Sweden)

    Sharma Rashmi

    2007-01-01

    Full Text Available Gardasil ® is the first quadrivalent human papillomavirus (HPV- types 6, 11, 16, 18 recombinant vaccine approved by the FDA on June 8, 2006. It induces genotype-specific virus-neutralizing antibodies and prevents infection with HPV. Various clinical trials demonstrated a reduction in the incidence of vaccine-type-specific persistent infections and of associated moderate- and high-grade cervical dysplasias and carcinomas in situ after its use. Gardasil is currently approved by FDA for prevention of genital warts, cancers and precancerous conditions of cervix and vulva in 9-26 years old females. Three doses of 0.5 ml of gardasil each at 0, 2 and 6 months are given intramuscularly. It is contraindicated in individuals who are hypersensitive to the active substances or to any of the excipients of the vaccine, patients with bleeding abnormalities or patients on anticoagulant therapy and during pregnancy. However, the vaccine, at an estimated $300-500 per course, is too expensive for many women in developing countries. Moreover, question regarding the longevity of the protection by vaccine is still unsolved. Hence, longer studies are required to establish its real status in cancer prevention.

  16. How bio-questionable are the different recombinant human erythropoietin copy products in Thailand?

    Science.gov (United States)

    Halim, Liem Andhyk; Brinks, Vera; Jiskoot, Wim; Romeijn, Stefan; Praditpornsilpa, Kearkiat; Assawamakin, Anunchai; Schellekens, Huub

    2014-05-01

    The high prevalence of pure red cell aplasia in Thailand has been associated with the sharp increase in number of recombinant human erythropoietin (rhEPO) copy products, based on a classical generic regulatory pathway, which have entered the market. This study aims to assess the quality of rhEPO copy products being used in Thailand. Twelve rhEPO copy products were purchased from pharmacies in Thailand, shipped under controlled cold chain conditions to the Netherlands and characterized using (1) high performance size-exclusion chromatography, (2) asymmetrical flow field-flow fractionation, (3) sodium dodecyl sulfate polyacrylamide gel electrophoresis in combination with (4) Western blotting and additionally tested for (5) host cell protein impurities as well as (6) endotoxin contamination. Some of the tested rhEPO copy products showed high aggregate levels and contained a substantial amount of protein fragments. Also, one of rhEPO copy products had a high endotoxin level, exceeding the FDA limit. Our observations show that some of the tested copy products on the Thai market differ significantly from the originator rhEPO product, Epogen®. This comparison study supports a link between the quality attributes of copy rhEPO products and their immunogenicity.

  17. The preventive effect of recombinant human growth factor (rhEGF) on the recurrence of radiodermatitis

    International Nuclear Information System (INIS)

    Ryu, Seung-Hee; Kim, Yeun-Hwa; Lee, Sang-Wook; Hong, Joon-Pio

    2010-01-01

    The effects of topical application of recombinant human epidermal growth factor (rhEGF) on wound healing and the recurrence of radiodermatitis were assessed in the irradiated skin of BALB/c Nu/Nu mice. Mice irradiated with 45 Gy of radiation were divided into 5 groups and treated with 10, 50, and 100 μg/g rhEGF ointment, vehicle alone, or no treatment (control) for 6 months. Wounds were observed initially in all groups and complete healing time (HT 100 ) for initial wound repair did not differ significantly among groups. However, the rate of recurrence over 6 months was significantly lower in the EGF-treated groups than in the control group (p<0.05). Histological examination showed that treatment with the optimum dose of EGF (50 μg/g) accelerated normal wound healing when compared with the higher dose of EGF (100 μg/g), vehicle alone, or no treatment, with the latter group showing irregular epidermal thickness, poor definition of epidermis and dermis, and unstable dermal structure. Collagen distribution was also significantly increased in mice treated with 50 μg/g rhEGF (p<0.05) compared with the control or vehicle-treated group. Taken together, these results indicate that treatment with exogenous EGF (50 μg/g dose) can enhance radiation-induced wound repair while preserving structural tissue stability and preventing the recurrence of radiodermatitis. (author)

  18. Recombinant erythropoietin in humans has a prolonged effect on circulating erythropoietin isoform distribution

    DEFF Research Database (Denmark)

    Aachmann-Andersen, Niels Jacob; Just Christensen, Søren; Lisbjerg, Kristian

    2014-01-01

    The membrane-assisted isoform immunoassay (MAIIA) quantitates erythropoietin (EPO) isoforms as percentages of migrated isoforms (PMI). We evaluated the effect of recombinant human EPO (rhEPO) on the distribution of EPO isoforms in plasma in a randomized, placebo-controlled, double-blinded, cross......-over study. 16 healthy subjects received either low-dose Epoetin beta (5000 IU on days 1, 3, 5, 7, 9, 11 and 13); high-dose Epoetin beta (30.000 IU on days 1, 2 and 3 and placebo on days 5, 7, 9, 11 and 13); or placebo on all days. PMI on days 4, 11 and 25 was determined by interaction of N......-acetyl glucosamine with the glycosylation dependent desorption of EPO isoforms. At day 25, plasma-EPO in both rhEPO groups had returned to values not different from the placebo group. PMI with placebo, reflecting the endogenous EPO isoforms, averaged 82.5 (10.3) % (mean (SD)). High-dose Epoetin beta decreased PMI...

  19. A simple approach for human recombinant apolipoprotein E4 expression and purification.

    Science.gov (United States)

    Argyri, Letta; Skamnaki, Vassiliki; Stratikos, Efstratios; Chroni, Angeliki

    2011-10-01

    We report a simple expression and purification procedure for the production of recombinant apolipoprotein E4 (apoE4), an important protein for the lipid homeostasis in humans that plays critical roles in the pathogenesis of cardiovascular and neurodegenerative diseases. Our approach is based on the expression of a thioredoxin-apoE4 fusion construct in bacterial cells and subsequent removal of the fused thioredoxin using the highly specific 3C protease, avoiding costly and laborious lipidation-delipidation steps used before. Our approach results in rapid, high-yield production of structurally and functionally competent apoE4 as evidenced by secondary structure measurements, thermal and chemical melting profiles and the kinetic profile of solubilization of dimyristoyl-phosphatidylcholine (DMPC) vesicles. This protocol is appropriate for laboratories with little experience in apolipoprotein biochemistry and will facilitate future studies on the role of apoE4 in the pathogenesis of cardiovascular disease and neurodegenerative diseases, including Alzheimer's disease. Copyright © 2011 Elsevier Inc. All rights reserved.

  20. Allelic recombination between distinct genomic locations generates copy number diversity in human β-defensins

    Science.gov (United States)

    Bakar, Suhaili Abu; Hollox, Edward J.; Armour, John A. L.

    2009-01-01

    β-Defensins are small secreted antimicrobial and signaling peptides involved in the innate immune response of vertebrates. In humans, a cluster of at least 7 of these genes shows extensive copy number variation, with a diploid copy number commonly ranging between 2 and 7. Using a genetic mapping approach, we show that this cluster is at not 1 but 2 distinct genomic loci ≈5 Mb apart on chromosome band 8p23.1, contradicting the most recent genome assembly. We also demonstrate that the predominant mechanism of change in β-defensin copy number is simple allelic recombination occurring in the interval between the 2 distinct genomic loci for these genes. In 416 meiotic transmissions, we observe 3 events creating a haplotype copy number not found in the parent, equivalent to a germ-line rate of copy number change of ≈0.7% per gamete. This places it among the fastest-changing copy number variants currently known. PMID:19131514

  1. Recombinant human serum albumin hydrogel as a novel drug delivery vehicle

    Energy Technology Data Exchange (ETDEWEB)

    Hirose, Masaaki, E-mail: Hirose.Masaaki@mh.mt-pharma.co.jp [Advanced Medical Research Laboratory, Research Division, Mitsubishi Tanabe Pharma Corporation, 3-16-89 Kashima, Yodogawa-ku, Osaka 532-8505 (Japan); Department of Applied Chemistry and Bioengineering, Graduate School of Engineering, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); Tachibana, Akira; Tanabe, Toshizumi [Department of Applied Chemistry and Bioengineering, Graduate School of Engineering, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan)

    2010-06-15

    Serum albumin acts as a physiological carrier for various compounds including drugs. A hydrogel consisting of recombinant human serum albumin (rHSA) was prepared to take advantage of drug binding ability of albumin for a sustained drug release carrier. The hydrogel was prepared by mixing rHSA and dithiothreitol and casted to a polystyrene mold. Hydrogel formation was thought to occur through the intermolecular interaction of the hydrophobic groups by protein denaturation. The release of sodium benzoate and salicylic acid from the hydrogel completed in 2 h, while warfarin release continued for 24 h. The total amounts of the drugs released from 100 mg of 15 and 5% rHSA hydrogel were 2.3 and 1.4 {mu}mol for warfarin, 1.4 and 1.1 {mu}mol for salicylic acid and 0.9 and 0.9 {mu}mol for sodium benzoate. These results reflected the order of the binding ability of drugs for intact albumin indicating that the drug binding ability of HSA still remained after the hydrogel formation. However, fibroblast cells attached and proliferated well on the hydrogel, indicating that denaturation of rHSA proceeded to the extent to allow the cell attachment. The present rHSA hydrogel might be suitable for a sustained release carrier of drugs having affinity for albumin.

  2. Potency Evaluation of Recombinant Human Erythropoietin in Brazil: Assessment of Reproducibility Using a Practical Approach

    Directory of Open Access Journals (Sweden)

    Michele Cardoso do Nascimento

    2015-08-01

    Full Text Available In this study, we compared the results of potency determination of recombinant human erythropoietin (rhEPO obtained between 2010 and 2012 by the National Institute of Quality Control in Health (INCQS/Fiocruz, i.e., the National Control Laboratory (NCL, and by a manufacturer of rhEPO. In total, 47 different batches of commercially prepared rhEPO (alpha isoform were analyzed. All results, including those of the control and warning limits, remained within the limits recommended by European Pharmacopoeia (Ph. Eur.. All relative error (RE values were less than ± 30%, wh ereas most were approximately ± 20%. Applying the Bland-Altman plot, only two of 47 values remained outside the limits of agreement (LA. In addition, agreement of potency determination between INCQS and the manufacturer coefficient of variation of reproducibility (% CVR was considered satisfactory. Taken together, our results demonstrate (i. the potency assay of rhEPO performed at INCQS, is standardized and controlled, (ii. the comparison of our results with those of the manufacturer, revealed an adequate inter-laboratory variation, and (iii. the critical appraisal proposed here appears to be a feasible tool to assess the reproducibility of biological activity, providing additional information regarding monitoring and production consistency to manufacturers and NCLs.

  3. Cortical bone growth and maturational changes in dwarf rats induced by recombinant human growth hormone

    Science.gov (United States)

    Martinez, D. A.; Orth, M. W.; Carr, K. E.; Vanderby, R. Jr; Vailas, A. C.

    1996-01-01

    The growth hormone (GH)-deficient dwarf rat was used to investigate recombinant human (rh) GH-induced bone formation and to determine whether rhGH facilitates simultaneous increases in bone formation and bone maturation during rapid growth. Twenty dwarf rats, 37 days of age, were randomly assigned to dwarf plus rhGH (GH; n = 10) and dwarf plus vehicle (n = 10) groups. The GH group received 1.25 mg rhGH/kg body wt two times daily for 14 days. Biochemical, morphological, and X-ray diffraction measurements were performed on the femur middiaphysis. rhGH stimulated new bone growth in the GH group, as demonstrated by significant increases (P bone length (6%), middiaphyseal cross-sectional area (20%), and the amount of newly accreted bone collagen (28%) in the total pool of middiaphyseal bone collagen. Cortical bone density, mean hydroxyapatite crystal size, and the calcium and collagen contents (microgram/mm3) were significantly smaller in the GH group (P bone collagen maturation, and mean hydroxyapatite crystal size may be independently regulated during rapid growth.

  4. Aerosolized recombinant human DNase I for the treatment of cystic fibrosis.

    Science.gov (United States)

    Shak, S

    1995-02-01

    Respiratory symptoms, recurrent infectious exacerbations, and progressive lung destruction in cystic fibrosis can be attributed to bacterial persistence and the accumulation of viscous purulent secretions in the airways. Purulent secretions contain high concentrations of extracellular DNA, a viscous material released by leukocytes. To evaluate the potential clinical utility of recombinant human DNase I (rhDNase or Pulmozyme), the human enzyme was cloned, sequenced, and expressed. In in vitro studies, rhDNase has been shown to reduce the viscoelasticity, reduce the adhesiveness, and improve the mucociliary transportability of cystic fibrosis sputum. In short-term phase 1 and phase 2 clinical trials, rhDNase has been shown to be safely tolerated and to improve the FEV1, FVC, and symptoms of dyspnea. A long-term placebo-controlled phase 3 study was performed in 968 adults and children (> or = 5 years) with cystic fibrosis to determine the effect of rhDNase on the risk of respiratory exacerbations requiring parenteral antibiotics and on the FEV1. Compared with placebo-treated patients, patients treated with rhDNase once daily or twice daily experienced a reduced risk of respiratory exacerbations by 28% (p = 0.04) and 37% (p = 0.01), respectively, and had a mean improvement in FEV1 of 5.8% (p < 0.01) and 5.6% (p < 0.01), respectively. Compared with placebo-treated patients, patients treated with rhDNase spent 2.7 fewer days receiving parenteral antibiotics (p = 0.04) and spent 1.3 fewer days in the hospital (p = 0.06) over the 6-month treatment period. Inhalation of rhDNase did not cause anaphylaxis but was associated with upper airway symptoms (ie, voice alteration, hoarseness, pharyngitis) that were generally mild and transient. In conclusion, aerosol administration of rhDNase was safely tolerated, reduced the risk of infectious exacerbations requiring parenteral antibiotics, and improved pulmonary function and patient well-being.

  5. Phylogenetic evidence for intratypic recombinant events in a novel human adenovirus C that causes severe acute respiratory infection in children.

    Science.gov (United States)

    Wang, Yanqun; Li, Yamin; Lu, Roujian; Zhao, Yanjie; Xie, Zhengde; Shen, Jun; Tan, Wenjie

    2016-03-10

    Human adenoviruses (HAdVs) are prevalent in hospitalized children with severe acute respiratory infection (SARI). Here, we report a unique recombinant HAdV strain (CBJ113) isolated from a HAdV-positive child with SARI. The whole-genome sequence was determined using Sanger sequencing and high-throughput sequencing. A phylogenetic analysis of the complete genome indicated that the CBJ113 strain shares a common origin with HAdV-C2, HAdV-C6, HAdV-C1, HAdV-C5, and HAdV-C57 and formed a novel subclade on the same branch as other HAdV-C subtypes. BootScan and single nucleotide polymorphism analyses showed that the CBJ113 genome has an intra-subtype recombinant structure and comprises gene regions mainly originating from two circulating viral strains: HAdV-1 and HAdV-2. The parental penton base, pVI, and DBP genes of the recombinant strain clustered with the HAdV-1 prototype strain, and the E1B, hexon, fiber, and 100 K genes of the recombinant clustered within the HAdV-2 subtype, meanwhile the E4orf1 and DNA polymerase genes of the recombinant shared the greatest similarity with those of HAdV-5 and HAdV-6, respectively. All of these findings provide insight into our understanding of the dynamics of the complexity of the HAdV-C epidemic. More extensive studies should address the pathogenicity and clinical characteristics of the novel recombinant.

  6. Selectable high-yield recombinant protein production in human cells using a GFP/YFP nanobody affinity support.

    Science.gov (United States)

    Schellenberg, Matthew J; Petrovich, Robert M; Malone, Christine C; Williams, R Scott

    2018-03-25

    Recombinant protein expression systems that produce high yields of pure proteins and multi-protein complexes are essential to meet the needs of biologists, biochemists, and structural biologists using X-ray crystallography and cryo-electron microscopy. An ideal expression system for recombinant human proteins is cultured human cells where the correct translation and chaperone machinery are present. However, compared to bacterial expression systems, human cell cultures present several technical challenges to their use as an expression system. We developed a method that utilizes a YFP fusion-tag to generate recombinant proteins using suspension-cultured HEK293F cells. YFP is a dual-function tag that enables direct visualization and fluorescence-based selection of high expressing clones for and rapid purification using a high-stringency, high-affinity anti-GFP/YFP nanobody support. We demonstrate the utility of this system by expressing two large human proteins, TOP2α (340 KDa dimer) and a TOP2β catalytic core (260 KDa dimer). This robustly and reproducibly yields >10 mg/L liter of cell culture using transient expression or 2.5 mg/L using stable expression. Published 2018. This article is a US Government work and is in the public domain in the USA.

  7. Radiostatine and radioiodine uptake characterization in sodium iodine symporter-expressing cell lines

    International Nuclear Information System (INIS)

    Petrich, T.; Helmeke, H.J.; Meyer, G.J.; Knapp, W.H.; Poetter, E.

    2002-01-01

    Full text: The sodium iodide symporter (NIS) has been recognized as an attractive target for cancer gene therapy. Here we investigated NIS-mediated transport of the high LET α-emitter astatine, 211 At, in comparison to radioiodine. A constitutive expression vector harbouring the human NIS cDNA was used in combination with reporter gene vectors for transient transfection of 13 different human cancer cell lines. Radioiodine uptake was measured as well as transfection efficiencies. Six stable NIS-expressing cell lines (3 derived from thyroid carcinomas, 2 colon carcinoma, 1 glioblastoma) were generated by antibiotic selection. NIS expression was monitored by immunohistochemistry and RT-PCR. Subsequently the radioastatine and radioiodine uptake characteristics of genetically modified cells were studied in comparison to the respective control cells. After xenotransplantation in nude mice in vivo tumor imaging by scintigraphy and biodistribution studies following organ removal were performed. Transient transfection of NIS cDNA led to high specific sodium perchlorate-sensitive radioiodine uptake in NIS-expressing cells that roughly correlates to transfection efficiencies. Similarly, stable NIS-expressing cell lines were able to concentrate high levels of radioiodine and in addition showed comparable transport capacity for radioastatine. Accumulation of 211 At was inhibited by sodium perchlorate like iodide uptake and displayed dependency an extracellular Na + - and I - -ions as well. Compared to wash-out experiments in cell culture the effective half life of radioiodine and radioastatine in vivo was significantly prolonged. Preliminary dose calculations by MIRD concepts indicated higher tumor radiation doses for 211 At compared to 131 I. Tumor cells of different origins transfected with the NIS-expression vector specifically and significantly take-up radioiodine and radioastatine in vitro and in vivo. The data provide direct evidence that the NIS efficiently transports

  8. Recombinant human erythropoietin alpha improves the efficacy of radiotherapy of a human tumor xenograft, affecting tumor cells and microvessels

    International Nuclear Information System (INIS)

    Loevey, J.; Bereczky, B.; Gilly, R.; Kenessey, I.; Raso, E.; Simon, E.; Timar, J.; Dobos, J.; Vago, A.; Kasler, M.; Doeme, B.; Tovari, J.

    2008-01-01

    Background and purpose: tumor-induced anemia often occurs in cancer patients, and is corrected by recombinant human erythropoietins (rHuEPOs). Recent studies indicated that, besides erythroid progenitor cells, tumor and endothelial cells express erythropoietin receptor (EPOR) as well; therefore, rHuEPO may affect their functions. Here, the effect of rHuEPOα on irradiation in EPOR-positive human squamous cell carcinoma xenograft was tested. Material and methods: A431 tumor-bearing SCID mice were treated from the tumor implantation with rHuEPOα at human-equivalent dose. Xenografts were irradiated (5 Gy) on day 14, and the final tumor mass was measured on day 22. The systemic effects of rHuEPOα on the hemoglobin level, on tumor-associated blood vessels and on hypoxia-inducible factor-(HIF-)1α expression of the tumor xenografts were monitored. The proliferation, apoptosis and clonogenic capacity of A431 cancer cells treated with rHuEPOα and irradiation were also tested in vitro. Results: in vitro, rHuEPOα treatment alone did not modify the proliferation of EPOR-positive A431 tumor cells but enhanced the effect of irradiation on proliferation, apoptosis and clonogenic capacity. In vivo, rHuEPOα administration compensated the tumor-induced anemia in SCID mice and decreased tumoral HIF-1α expression but had no effect on tumor growth. At the same time rHuEPOα treatment significantly increased the efficacy of radiotherapy in vivo (tumor weight of 23.9 ± 4.7 mg and 34.9 ± 4.6 mg, respectively), mediated by increased tumoral blood vessel destruction. Conclusion: rHuEPOα treatment may modulate the efficacy of cancer radiotherapy not only by reducing systemic hypoxia and tumoral HIF-1α expression, but also by destroying tumoral vessels. (orig.)

  9. Procedure guideline for radioiodine test (version 3)

    International Nuclear Information System (INIS)

    Dietlein, M.; Schicha, H.; Eschner, W.; Deutsche Gesellschaft fuer Medizinische Physik; Koeln Univ.; Lassmann, M.; Deutsche Gesellschaft fuer Medizinische Physik; Wuerzburg Univ.; Leisner, B.; Allgemeines Krankenhaus St. Georg, Hamburg; Reiners, C.; Wuerzburg Univ.

    2007-01-01

    The version 3 of the procedure guideline for radioiodine test is an update of the guideline previously published in 2003. The procedure guideline discusses the pros and cons of a single measurement or of repeated measurements of the iodine-131 uptake and their optimal timing. Different formulas are described when one, two or three values of the radioiodine kinetic are available. The probe with a sodium-iodine crystal, alternatively or additionally the gamma camera using the ROI-technique are instrumentations for the measurement of iodine-131 uptake. A possible source of error is an inappropriate measurement (sonography) of the target volume. The patients' preparation includes the withdrawal of antithyroid drugs 2-3 days before radioiodine administration. The patient has to avoid iodine-containing medication and the possibility of additives of iodine in vitamin- and electrolyte-supplementation has to be considered. (orig.)

  10. International collaborative study for the calibration of proposed International Standards for thromboplastin, rabbit, plain and for thromboplastin, recombinant, human, plain

    DEFF Research Database (Denmark)

    van den Besselaar, A M H P; Chantarangkul, V; Angeloni, F

    2018-01-01

    BACKGROUND: The availability of International Standards for thromboplastin is essential for the calibration of routine reagents and hence the calculation of the International Normalized Ratio (INR). Stocks of the current 4(th) International Standards are running low. Candidate replacement materia......) international standard (rTF/09). The candidate materials have been accepted by WHO as the 5(th) International Standards for thromboplastin, rabbit plain, and thromboplastin, recombinant, human, plain. This article is protected by copyright. All rights reserved.......BACKGROUND: The availability of International Standards for thromboplastin is essential for the calibration of routine reagents and hence the calculation of the International Normalized Ratio (INR). Stocks of the current 4(th) International Standards are running low. Candidate replacement materials...... have been prepared. This report describes the calibration of the proposed 5(th) International Standards for thromboplastin, rabbit, plain (coded RBT/16) and for thromboplastin, recombinant, human, plain (coded rTF/16). METHODS: An international collaborative study was carried out for the assignment...

  11. Experimental biodistribution studies of 99mTc-recombinant human serum albumin (rHSA): a new generation of radiopharmaceutical

    International Nuclear Information System (INIS)

    Perkins, A.C.; Frier, M.

    1994-01-01

    Recombinant human serum albumin (rHSA) produced by cultured fermentation has been prepared in the form of microcapsules nominally 3-5 μm in diameter and radiolabelled with technetium-99m following reduction with stannous chloride. Radiochemical purity was assessed by chromatography on instant thin-layer chromatography and found to be greater than 90%. No evidence of aggregation was seen by microscopic examination. Imaging biodistribution studies in New Zealand white rabbits demonstrated targeting to the liver or lung, respectively, depending upon the size and surfactant properties of the microcapsules. This communication is the first to show scintigraphic studies using 99m Tc-labelled rHSA with the potential for lung, liver and cardiovascular imaging and demonstrates that recombinant DNA technology offers an important new source of materials suitable for use as radiopharmaceuticals without the need for pooled human blood products. (orig.)

  12. Isolation and structure determination of the intact sialylated N-linked carbohydrate chains of recombinant human follitropin expressed in Chinese hamster ovary cells

    NARCIS (Netherlands)

    Vliegenthart, J.F.G.; Hård, K.; Mekking, A.; Damm, J.B.L.; Kamerling, J.P.; Boer, W. de; Wijnands, R.A.

    1990-01-01

    Biologically active recombinant human follitropin has been expressed in Chinese hamster ovary cells. The carbohydrate chains of the recombinant glycoprotein hormone were enzymatically released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. The oligosaccharides were separated from

  13. Research on removal of radioiodine by charcoal

    International Nuclear Information System (INIS)

    Li Wangchang; Huang Yuying; Wu Yianwei; Jia Ming; Guo Liangtian

    1993-01-01

    The major R and D work carried out in the CIRP laboratory on removal of radioiodine is introduced, which involves the adsorption performances of various kinds of fruit shell base and coal base charcoal impregnated with chemicals, the influence of various parameters, the technique of non-destructive test for commercial scale iodine adsorber, and the iodine samplers for both gross iodine and iodine in different forms. The experimental results have been applied to the design and test of iodine adsorber and the monitoring of airborne radioiodine

  14. The Effect of Recombinant Human MG53 Protein on Tourniquet-induced Ischemia Reperfusion Injury in Rat Muscle

    Science.gov (United States)

    2014-06-01

    blind to the treatment , and the prevalence of damaged fibers was quantitated from 10 10x images from each muscle . Approximately 800 fibers were counted...therapeutic cell membrane repair in treatment of muscular dystrophy . Sci Transl Med. 2012; 4(139):139ra185. 11. Weisleder N, Lin P, Zhao X, Orange M, Zhu H...The effect of recombinant human MG53 protein on tourniquet- induced ischemia reperfusion injury in rat muscle Benjamin T. Corona, Ph.D.1, Koyal Garg

  15. Evidence of molecular evolution driven by recombination events influencing tropism in a novel human adenovirus that causes epidemic keratoconjunctivitis.

    Directory of Open Access Journals (Sweden)

    Michael P Walsh

    2009-06-01

    Full Text Available In 2005, a human adenovirus strain (formerly known as HAdV-D22/H8 but renamed here HAdV-D53 was isolated from an outbreak of epidemic keratoconjunctititis (EKC, a disease that is usually caused by HAdV-D8, -D19, or -D37, not HAdV-D22. To date, a complete change of tropism compared to the prototype has never been observed, although apparent recombinant strains of other viruses from species Human adenovirus D (HAdV-D have been described. The complete genome of HAdV-D53 was sequenced to elucidate recombination events that lead to the emergence of a viable and highly virulent virus with a modified tropism. Bioinformatic and phylogenetic analyses of this genome demonstrate that this adenovirus is a recombinant of HAdV-D8 (including the fiber gene encoding the primary cellular receptor binding site, HAdV-D22, (the epsilon determinant of the hexon gene, HAdV-D37 (including the penton base gene encoding the secondary cellular receptor binding site, and at least one unknown or unsequenced HAdV-D strain. Bootscanning analysis of the complete genomic sequence of this novel adenovirus, which we have re-named HAdV-D53, indicated at least five recombination events between the aforementioned adenoviruses. Intrahexon recombination sites perfectly framed the epsilon neutralization determinant that was almost identical to the HAdV-D22 prototype. Additional bootscan analysis of all HAdV-D hexon genes revealed recombinations in identical locations in several other adenoviruses. In addition, HAdV-D53 but not HAdV-D22 induced corneal inflammation in a mouse model. Serological analysis confirmed previous results and demonstrated that HAdV-D53 has a neutralization profile representative of the epsilon determinant of its hexon (HAdV-D22 and the fiber (HAdV-D8 proteins. Our recombinant hexon sequence is almost identical to the hexon sequences of the HAdV-D strain causing EKC outbreaks in Japan, suggesting that HAdV-D53 is pandemic as an emerging EKC agent. This documents

  16. Recombinant DNA derived monomeric insulin analogue: comparison with soluble human insulin in normal subjects.

    Science.gov (United States)

    Vora, J P; Owens, D R; Dolben, J; Atiea, J A; Dean, J D; Kang, S; Burch, A; Brange, J

    1988-11-12

    To compare the rate of absorption from subcutaneous tissue and the resulting hypoglycaemic effect of iodine-125 labelled soluble human insulin and a monomeric insulin analogue derived by recombinant DNA technology. Single blind randomised comparison of equimolar doses of 125I labelled soluble human insulin and insulin analogue. Study in normal people at a diabetes research unit and a university department of medical physics. Seven healthy male volunteers aged 20-39 not receiving any other drugs. After an overnight fast and a basal period of one hour two doses (0.05 and 0.1 U/kg) of 125I labelled soluble human insulin and insulin analogue were injected subcutaneously into the anterior abdominal wall on four separate days. To find a fast acting insulin for meal related requirements in insulin dependent diabetics. MEASUREMENTS and main results--Residual radioactivity at the injection site was measured continuously for the first two hours after injection of the 125I labelled preparations and thereafter for five minutes simultaneously with blood sampling. Frequent venous blood samples were obtained over six hours for determination of plasma immunoreactive insulin, insulin analogue, glucose, and glucagon values. Time to 50% of initial radioactivity at the injection site for the insulin analogue compared with soluble insulin was 61 v 135 minutes (p less than 0.05) with 0.05 U/kg and 67 v 145 minutes (p less than 0.001) with 0.1 U/kg. Concentrations in plasma increased faster after the insulin analogue compared with soluble insulin, resulting in higher plasma concentrations between 10 and 150 minutes (0.001 less than p less than 0.05) after 0.05 U/kg and between 40 and 360 minutes (0.001 less than p less than 0.05) after 0.1 U/kg. The hypoglycaemic response to insulin analogue was a plasma glucose nadir at 60 minutes with both doses compared with 90 and 120 minutes with soluble insulin at 0.5 and 0.1 U/kg respectively. The response of glucagon substantiated the earlier and

  17. An outbreak of severe infections among Australian infants caused by a novel recombinant strain of human parechovirus type 3.

    Science.gov (United States)

    Nelson, Tiffanie M; Vuillermin, Peter; Hodge, Jason; Druce, Julian; Williams, David T; Jasrotia, Rekha; Alexandersen, Soren

    2017-03-14

    Human parechovirus types 1-16 (HPeV1-16) are positive strand RNA viruses in the family Picornaviridae. We investigated a 2015 outbreak of HPeV3 causing illness in infants in Victoria, Australia. Virus genome was extracted from clinical material and isolates and sequenced using a combination of next generation and Sanger sequencing. The HPeV3 outbreak genome was 98.7% similar to the HPeV3 Yamagata 2011 lineage for the region encoding the structural proteins up to nucleotide position 3115, but downstream of that the genome varied from known HPeV sequences with a similarity of 85% or less. Analysis indicated that recombination had occurred, may have involved multiple types of HPeV and that the recombination event/s occurred between March 2012 and November 2013. However the origin of the genome downstream of the recombination site is unknown. Overall, the capsid of this virus is highly conserved, but recombination provided a different non-structural protein coding region that may convey an evolutionary advantage. The indication that the capsid encoding region is highly conserved at the amino acid level may be helpful in directing energy towards the development of a preventive vaccine for expecting mothers or antibody treatment of young infants with severe disease.

  18. Distribution of 131I-labeled recombinant human erythropoietin in maternal and fetal organs following intravenous administration in pregnant rats

    International Nuclear Information System (INIS)

    Yilmaz, O.; Lambrecht, F.Y.; Durkan, K.; Gokmen, N.; Erbayraktar, S.

    2007-01-01

    The aim of the present study was to demonstrate the possible transplacental transmission of 131 I labeled recombinant human erythropoietin ( 131 I-rh-EPO) in pregnant rats and its distribution through maternal and fetal organs. Six Wistar Albino Rats in their pregnancy of 18 days were used 131 I labeled recombinant human erythropoietin (specific activity = 2.4 μCi/IU) was injected into the tail vein of rats. After 30 minutes labeled erythropoietin infusion maternal stomach, kidney, lung, liver, brain and heart as well as fetus were removed. Then, the same organs were removed from each fetus. Measuring weight of maternal and fetal organs as well as placenta were followed by radioactivity count via Cd(Te) detector. 131 I labeled recombinant human erythropoietin was found to be able to pass rat placenta and its distribution order in fetal organs was similar to those of maternal organs. Besides, as measurements were performed closer to cornu uteri, uptakes were decreasing in every fetus and its corresponding placenta. (author)

  19. Increased preoperative collection of autologous blood with recombinant human erythropoietin therapy in tertiary care hospitals of Jammu

    Directory of Open Access Journals (Sweden)

    Kumkum Sharma

    2013-01-01

    Full Text Available Introduction: To study whether the administration of recombinant human erythropoietin increases the amount of autologous blood that can be collected before orthopaedic surgery. Materials and Methods: We conducted a randomized controlled trial of recombinant human erythropoietin in 68 adults scheduled for elective orthopedic procedures. The patients received either erythropoietin 600 units/kg of body weight or placebo intravenously every 5 th day prior to each phlebotomy for 21 days during which time up to 5 units of blood was collected. Patients were excluded from donation when their hematocrit values were less than 33%. All patients received iron sulphate 325mg orally 3 times daily. The mean number of units collected per patient was 4.33 ± 0.4 for erythropoietin group and 3.05± 0.71 for the placebo group. Results: The mean packed red cell volume donated by patients who received erythropoietin was 32% greater than that donated by patients who received placebo (196.3 vs. 169.4 ml, p<0.05. 68% in the placebo group and 9% of patients treated with erythropoietin were unable to donate ≥4 units. No adverse effects were attributed to erythropoietin. While participating in the study, complications developed in 2 patients one in each group necessitating their removal from the study. Conclusion: We conclude that recombinant human erythropoietin increases the ability of the patients about to undergo elective surgery to donate autologous blood units.

  20. Brucella melitensis VirB12 recombinant protein is a potential marker for serodiagnosis of human brucellosis.

    Science.gov (United States)

    Mirkalantari, Shiva; Zarnani, Amir-Hassan; Nazari, Mahboobeh; Irajian, Gholam Reza; Amirmozafari, Nour

    2017-03-03

    The numerous drawbacks of current serological tests for diagnosis of brucellosis which mainly results from cross reactivity with LPS from other gram-negative bacteria have generated an increasing interest to find more specific non-LPS antigens. Previous studies had indicated that Brucella VirB12 protein, a cell surface protein and component of type IV secretion system, induces antibody response during animal infection. However, this protein has not yet been tested as a serological diagnostic marker in human brucellosis. Recombinant VirB12 protein was prepared and evaluated the efficacy of it in an indirect enzyme-linked immunosorbent assay (ELISA) for brucellosis with sera collected from different region of Iran and the results were compared with a commercial ELISA kit. Sera from human brucellosis patients strongly reacted to the purified recombinant VirB12. The sensitivity, specificity, accuracy, negative predictive value and positive predictive value of recombinant VirB12-based ELISA related to the commercial-ELISA method were 87.8, 94, 90, 80 and 96.6% respectively. We concluded that antigenic VirB12 have a property value that can be considered as a candidate for using in serodiagnostic tests for human brucellosis.

  1. Evaluation of recombinant porin (rOmp2a) protein as a potential antigen candidate for serodiagnosis of Human Brucellosis.

    Science.gov (United States)

    Pathak, Prachi; Kumar, Ashu; Thavaselvam, Duraipandian

    2017-07-11

    Brucellosis is an important zoonotic disease caused by different Brucella species and human brucellosis is commonly prevalent in different states of India. Among various Brucella species, B. melitensis is most pathogenic to human and included as category B biothreat which can cause infection through aerosol, cut, wounds in skin and contact with infected animals. The diagnosis of human brucellosis is very important for proper treatment and management of disease as there is no vaccine available for human use. The present study was designed to clone, express and purify immunodominant recombinant omp2a (rOmp2a) porin protein of B. melitensis and to evaluate this new antigen candidate for specific serodiagnosis of human brucellosis by highly sensitive iELISA (indirect enzyme linked immunosorbent assay). Omp2a gene of B. melitensis 16 M strain was cloned and expressed in pET-SUMO expression system. The recombinant protein was purified under denaturing conditions using 8 M urea. The purified recombinant protein was confirmed by western blotting by reacting with anti-HIS antibody. The sero-reactivity of the recombinant protein was also checked by reacting with antisera of experimentally infected mice with B. melitensis 16 M at different time points. Serodiagnostic potential of recombinant porin antigen was tested against 185 clinical serum samples collected from regions endemic to brucellosis in southern part of India by iELISA. The samples were grouped into five groups. Group 1 contained cultured confirmed positive serum samples of brucellosis (n = 15), group 2 contained sera samples from positive cases of brucellosis previously tested by conventional methods of RBPT (n = 28) and STAT (n = 26), group 3 contained sera samples negative by RBPT(n = 36) and STAT (n = 32), group 4 contained sera samples of other febrile illness and PUO case (n = 35) and group 5 contained confirmed negative sera samples from healthy donors (n = 23). The rOmp2a was found to be

  2. Characterization of a recombinant humanized anti-cocaine monoclonal antibody and its Fab fragment.

    Science.gov (United States)

    Kirley, Terence L; Norman, Andrew B

    2015-01-01

    Variations of post-translational modifications are important for stability and in vivo behavior of therapeutic antibodies. A recombinant humanized anti-cocaine monoclonal antibody (h2E2) was characterized for heterogeneity of N-linked glycosylation and disulfide bonds. In addition, charge heterogeneity, which is partially due to the presence or absence of C-terminal lysine on the heavy chains, was examined. For cocaine overdose therapy, Fab fragments may be therapeutic, and thus, a simplified method of generation, purification, and characterization of the Fab fragment generated by Endoproteinase Lys-C digestion was devised. Both the intact h2E2 antibody and purified Fab fragments were analyzed for their affinities for cocaine and 2 of its metabolites, benzoylecgonine and cocaethylene, by fluorescence quenching of intrinsic antibody tyrosine and tryptophan fluorescence resulting from binding of these drugs. Binding constants obtained from fluorescence quenching measurements are in agreement with recently published radioligand and ELISA binding assays. The dissociation constants determined for the h2E2 monoclonal and its Fab fragment are approximately 1, 5, and 20 nM for cocaethylene, cocaine, and benzoylecgonine, respectively. Tryptophan fluorescence quenching (emission at 330 nm) was measured after either excitation of tyrosine and tryptophan (280 nm) or selective excitation of tryptophan alone (295 nm). More accurate binding constants are obtained using tryptophan selective excitation at 295 nm, likely due to interfering absorption of cocaine and metabolites at 280 nm. These quenching results are consistent with multiple tryptophan and tyrosine residues in or near the predicted binding location of cocaine in a previously published 3-D model of this antibody's variable region.

  3. A Biallelic Mutation in the Homologous Recombination Repair Gene SPIDR Is Associated With Human Gonadal Dysgenesis.

    Science.gov (United States)

    Smirin-Yosef, Pola; Zuckerman-Levin, Nehama; Tzur, Shay; Granot, Yaron; Cohen, Lior; Sachsenweger, Juliane; Borck, Guntram; Lagovsky, Irina; Salmon-Divon, Mali; Wiesmüller, Lisa; Basel-Vanagaite, Lina

    2017-02-01

    Primary ovarian insufficiency (POI) is caused by ovarian follicle depletion or follicle dysfunction, characterized by amenorrhea with elevated gonadotropin levels. The disorder presents as absence of normal progression of puberty. To elucidate the cause of ovarian dysfunction in a family with POI. We performed whole-exome sequencing in 2 affected individuals. To evaluate whether DNA double-strand break (DSB) repair activities are altered in biallelic mutation carriers, we applied an enhanced green fluorescent protein-based assay for the detection of specific DSB repair pathways in blood-derived cells. Diagnoses were made at the Pediatric Endocrine Clinic, Clalit Health Services, Sharon-Shomron District, Israel. Genetic counseling and sample collection were performed at the Pediatric Genetics Unit, Schneider Children's Medical Center Israel, Petah Tikva, Israel. Two sisters born to consanguineous parents of Israeli Muslim Arab ancestry presented with a lack of normal progression of puberty, high gonadotropin levels, and hypoplastic or absent ovaries on ultrasound. Blood samples for DNA extraction were obtained from all family members. Exome analysis to elucidate the cause of POI in 2 affected sisters. Analysis revealed a stop-gain homozygous mutation in the SPIDR gene (KIAA0146) c.839G>A, p.W280*. This mutation altered SPIDR activity in homologous recombination, resulting in the accumulation of 53BP1-labeled DSBs postionizing radiation and γH2AX-labeled damage during unperturbed growth. SPIDR is important for ovarian function in humans. A biallelic mutation in this gene may be associated with ovarian dysgenesis in cases of autosomal recessive inheritance. Copyright © 2017 by the Endocrine Society

  4. Effects of Recombinant Human Bone Morphogenetic Protein-2 on Vertical Bone Augmentation in a Canine Model.

    Science.gov (United States)

    Hsu, Yung-Ting; Al-Hezaimi, Khalid; Galindo-Moreno, Pablo; O'Valle, Francisco; Al-Rasheed, Abdulaziz; Wang, Hom-Lay

    2017-09-01

    Vertical bone augmentation (VBA) remains unpredictable and challenging for most clinicians. This study aims to compare hard tissue outcomes of VBA, with and without recombinant human bone morphogenetic protein (rhBMP)-2, under space-making titanium mesh in a canine model. Eleven male beagle dogs were used in the study. Experimental ridge defects were created to form atrophic ridges. VBA was performed via guided bone regeneration using titanium mesh and allografts. In experimental hemimandibles, rhBMP-2/absorbable collagen sponge was well mixed with allografts prior to procedures, whereas a control buffer was applied within controls. Dogs were euthanized after a 4-month healing period. Clinical and radiographic examinations were performed to assess ridge dimensional changes. In addition, specimens were used for microcomputed tomography (micro-CT) assessment and histologic analysis. Membrane exposure was found on five of 11 (45.5%) rhBMP-2-treated sites, whereas it was found on nine of 11 (81.8%) non-rhBMP-2-treated sites. Within 4 months of healing, rhBMP-2-treated sites showed better radiographic bone density, greater defect fill, and significantly more bone gain in ridge height (P 0.05). Under light microscope, predominant lamellar patterns were found in the specimen obtained from rhBMP-2 sites. With inherent limitations of the canine model and the concern of such a demanding surgical technique, current findings suggest that the presence of rhBMP-2 in a composite graft allows an increase of vertical gain, with formation of ectopic bone over the titanium mesh in comparison with non-rhBMP-2 sites.

  5. Protective effects of recombinant human brain natriuretic peptide in perioperative period during open heart surgery.

    Science.gov (United States)

    Xu, Yunbin; Li, Yong; Bao, Weiguo; Qiu, Shi

    2018-03-01

    The aim of the present study was to evaluate the protective effects and safety aspects of recombinant human brain natriuretic peptide (rhBNP) on cardiac functions of patients undergoing open-heart surgery during perioperative period. In total, 150 patients undergoing open heart surgery in the Second Hospital of Shandong Universty from August 2015 to July 2016 were randomly divided into control group and observation group each with 75 cases. Patients in control group were treated by routine rehabilitation while patients in the observation group were treated by both the routine rehabilitation and rhBNP. All the observations were made before operation, after operation and 7 days after operation. The changes of N-terminal pro-brain natriuretic peptide (NT-proBNP) of patients, the left ventricular ejection fraction (LVEF), cardiac function [Cardiac output (CO), pulmonary capillary wedge pressure (PAWP) and central venous pressure (CVP)] of patients were measured. Further, respirator support time, ICU stay time, incidence of complications and vital signs (BP, HR, SaO2) of patients in the two groups were also compared. NT-proBNP levels of all patients improved after operation but it decreased in both groups after 7 days of operation. The decrease of NT-proBNP levels in observation group was significantly higher than that of control group. Whereas, LVEF, CO, PAWP and CVP of patients in both the groups increased after operation but effects were significantly higher in the observation group after 7 days of medication. Respirator support time and ICU stay time of patients in observation group were significantly shorter than those in control group, and the incidence of postoperative complications of patients in the observation group were significantly lower than the control group. Moreover, BP, HR and SaO2 of patients in observation group were significantly elevated in comparison to control group (Popen heart surgery, and is safe as well as reliable.

  6. Concentration-dependent interactions of the organophosphates chlorpyrifos oxon and methyl paraoxon with human recombinant acetylcholinesterase

    International Nuclear Information System (INIS)

    Kaushik, R.; Rosenfeld, Clint A.; Sultatos, L.G.

    2007-01-01

    For many decades it has been thought that oxygen analogs (oxons) of organophosphorus insecticides phosphorylate the catalytic site of acetylcholinesterase by a mechanism that follows simple Michaelis-Menten kinetics. More recently, the interactions of at least some oxons have been shown to be far more complex and likely involve binding of oxons to a second site on acetylcholinesterase that modulates the inhibitory capacity of other oxon molecules at the catalytic site. The current study has investigated the interactions of chlorpyrifos oxon and methyl paraoxon with human recombinant acetylcholinesterase. Both chlorpyrifos oxon and methyl paraoxon were found to have k i 's that change as a function of oxon concentration. Furthermore, 10 nM chlorpyrifos oxon resulted in a transient increase in acetylthiocholine hydrolysis, followed by inhibition. Moreover, in the presence of 100 nM chlorpyrifos oxon, acetylthiocholine was found to influence both the K d (binding affinity) and k 2 (phosphorylation constant) of this oxon. Collectively, these results demonstrate that the interactions of chlorpyrifos oxon and methyl paraoxon with acetylcholinesterase cannot be described by simple Michaelis-Menten kinetics but instead support the hypothesis that these oxons bind to a secondary site on acetylcholinesterase, leading to activation/inhibition of the catalytic site, depending on the nature of the substrate and inhibitor. Additionally, these data raise questions regarding the adequacy of estimating risk of low levels of insecticide exposure from direct extrapolation of insecticide dose-response curves since the capacity of individual oxon molecules at low oxon levels could be greater than individual oxon molecules in vivo associated with the dose-response curve

  7. Nonclinical comparability studies of recombinant human arylsulfatase A addressing manufacturing process changes.

    Science.gov (United States)

    Wright, Teresa; Li, Aiqun; Lotterhand, Jason; Graham, Anne-Renee; Huang, Yan; Avila, Nancy; Pan, Jing

    2018-01-01

    Recombinant human arylsulfatase A (rhASA) is in clinical development for the treatment of patients with metachromatic leukodystrophy (MLD). Manufacturing process changes were introduced to improve robustness and efficiency, resulting in higher levels of mannose-6-phosphate and sialic acid in post-change (process B) compared with pre-change (process A) rhASA. A nonclinical comparability program was conducted to compare process A and process B rhASA. All doses were administered intrathecally. Pharmacodynamic comparability was evaluated in immunotolerant MLD mice, using immunohistochemical staining of lysosomal-associated membrane protein-1 (LAMP-1). Pharmacokinetic comparability was assessed in juvenile cynomolgus monkeys dosed once with 6.0 mg (equivalent to 100 mg/kg of brain weight) process A or process B rhASA. Biodistribution was compared by quantitative whole-body autoradiography in rats. Potential toxicity of process B rhASA was evaluated by repeated rhASA administration at doses of 18.6 mg in juvenile cynomolgus monkeys. The specific activities for process A and process B rhASA were 89 U/mg and 106 U/mg, respectively, which were both well within the target range for the assay. Pharmacodynamic assessments showed no statistically significant differences in LAMP-1 immunohistochemical staining in the spinal cord and in most of the brain areas assessed between process A and B rhASA-dosed mice. LAMP-1 staining was reduced with both process A and B rhASA compared with vehicle, supporting its activity. Concentration-time curves in cerebrospinal fluid and serum of cynomolgus monkeys were similar with process A and B rhASA. Process A and B rhASA were similar in terms of their pharmacokinetic parameters and biodistribution data. No process B rhASA-related toxicity was detected. In conclusion, manufacturing process changes did not affect the pharmacodynamic, pharmacokinetic or safety profiles of process B rhASA relative to process A rhASA.

  8. Improvement of in vivo efficacy of recombinant human erythropoietin by encapsulation in PEG–PLA micelle

    Directory of Open Access Journals (Sweden)

    Shi YN

    2012-12-01

    Full Text Available Yanan Shi,1,2,* Wan Huang,1,* Rongcai Liang,1–3 Kaoxiang Sun,2,3 Fangxi Zhang,2,3 Wanhui Liu,2,3 Youxin Li1–31College of Life Science, Jilin University, Changchun, China; 2State Key Laboratory of Long-acting and Targeting Drug Delivery System, Luye Pharmaceutical Co, Ltd, Yantai, China; 3School of Pharmacy, Yantai University, Yantai, China*These authors contributed equally to this workAbstract: To improve the pharmacokinetics and stability of recombinant human erythropoietin (rhEPO, rhEPO was successfully formulated into poly(ethylene glycol–poly(d,l-lactide (PEG–PLA di-block copolymeric micelles at diameters ranging from 60 to 200 nm with narrow polydispersity indices (PDIs; PDI < 0.3 and trace amount of protein aggregation. The zeta potential of the spherical micelles was in the range of −3.78 to 4.65 mV and the highest encapsulation efficiency of rhEPO in the PEG–PLA micelles was about 80%. In vitro release profiles indicated that the stability of rhEPO in the micelles was improved significantly and only a trace amount of aggregate was found. Pharmacokinetic studies in rats showed highly enhanced plasma retention time of the rhEPO-loaded PEG-PLA micelles in comparison with the native rhEPO group. Increased hemoglobin concentrations were also found in the rat study. Native polyacrylamide gel electrophoresis results demonstrated that rhEPO was successfully encapsulated into the micelles, which was stable in phosphate buffered saline with different pHs and concentrations of NaCl. Therefore, PEG–PLA micelles can be a potential protein drug delivery system.Keywords: rhEPO, PEG–PLA micelle, in vitro, pharmacokinetics, pharmacodynamics

  9. Experimental research on recombinant human endostatin-induced cardiomyocyte apoptosis in rats

    Directory of Open Access Journals (Sweden)

    Jing QIN

    2014-03-01

    Full Text Available Objective To explore the recombinant human endostatin (rh-ES-induced cardiotoxicity in rats and its mechanism. Methods Twenty four female Wistar rats were randomly divided into four groups (6 each. Rats in low, moderate and high dose group received rh-ES with a dosage of 3, 6 and 12mg/(kg·d, respectively, by intraperitoneal injection, and rats in control group received the same amount of normal saline alone. Half of rats in each group were sacrificed by spinal dislocation after 4 weeks and 8 weeks of the treatment. Pathomorphologic and ultrastructural changes in rat's myocardial tissue were evaluated by light microscopy and transmission electron microscopy. Cardiomyocyte apoptosis was detected with TdT-mediated dUTP nick end labeling (TUNEL assay. Microvessel density (MVD in myocardial tissue was measured by immunohistochemically marking endothelial cell with CD34. Results No pathomorphologic and ultrastrucural changes were found under light microscope and transmission electron microscope in the low dose and moderate dose groups, but cardiomyocyte damage were found in the high dose group. TUNEL assay revealed more apoptotic cells in high and moderate (only 8 weeks dose groups than in control group (P=0.033, P=0.000, and the apoptosis index was highest in the high dose group at 8 weeks. In addition, compared with the control group, MVD significantly increased in high dose groups at 4 weeks and 8 weeks (P<0.05. Conclusions rh-ES induces the cardiotoxicity in rats, and cardiomyocyte apoptosis is involved in the pathological course of cardiac toxicity. DOI: 10.11855/j.issn.0577-7402.2014.01.02

  10. Recombinant human erythropoietin stimulates angiogenesis and wound healing in the genetically diabetic mouse.

    Science.gov (United States)

    Galeano, Mariarosaria; Altavilla, Domenica; Cucinotta, Domenico; Russo, Giuseppina T; Calò, Margherita; Bitto, Alessandra; Marini, Herbert; Marini, Rolando; Adamo, Elena B; Seminara, Paolo; Minutoli, Letteria; Torre, Valerio; Squadrito, Francesco

    2004-09-01

    The effects of recombinant human erythropoietin (rHuEPO) in diabetes-related healing defects were investigated by using an incisional skin-wound model produced on the back of female diabetic C57BL/KsJ-m(+/+)Lept(db) mice (db(+)/db(+)) and their normoglycemic littermates (db(+/+)m). Animals were treated with rHuEPO (400 units/kg in 100 microl s.c.) or its vehicle alone (100 microl). Mice were killed on different days (3, 6, and 12 days after skin injury) for measurement of vascular endothelial growth factor (VEGF) mRNA expression and protein synthesis, for monitoring angiogenesis by CD31 expression, and for evaluating histological changes. Furthermore, we evaluated wound-breaking strength at day 12. At day 6, rHuEPO injection in diabetic mice resulted in an increase in VEGF mRNA expression (vehicle = 0.33 +/- 0.1 relative amount of mRNA; rHuEPO = 0.9 +/- 0.09 relative amount of mRNA; P < 0.05) and protein wound content (vehicle = 23 +/- 5 pg/wound; rHuEPO = 92 +/- 12 pg/wound; P < 0.05) and caused a marked increase in CD31 gene expression (vehicle = 0.18 +/- 0.05 relative amount of mRNA; rHuEPO = 0.98 +/- 0.21 relative amount of mRNA; P < 0.05) and protein synthesis. Furthermore, rHuEPO injection improved the impaired wound healing and, at day 12, increased the wound-breaking strength in diabetic mice (vehicle = 12 +/- 2 g/mm; rHuEPO 21 +/- 5 g/mm; P < 0.05). Erythropoietin may have a potential application in diabetes-related wound disorders.

  11. Recombinant human erythropoietin improves angiogenesis and wound healing in experimental burn wounds.

    Science.gov (United States)

    Galeano, Mariarosaria; Altavilla, Domenica; Bitto, Alessandra; Minutoli, Letteria; Calò, Margherita; Lo Cascio, Patrizia; Polito, Francesca; Giugliano, Giovanni; Squadrito, Giovanni; Mioni, Chiara; Giuliani, Daniela; Venuti, Francesco S; Squadrito, Francesco

    2006-04-01

    Erythropoietin interacts with vascular endothelial growth factor (VEGF) and stimulates endothelial cell mitosis and motility; thus it may be of importance in the complex phenomenon of wound healing. The purpose of this study was to investigate the effect of recombinant human erythropoietin (rHuEPO) on experimental burn wounds. Randomized experiment. Research laboratory. C57BL/6 male mice weighing 25-30 g. Mice were immersed in 80 degrees C water for 10 secs to achieve a deep-dermal second degree burn. Animals were randomized to receive either rHuEPO (400 units/kg/day for 14 days in 100 microL subcutaneously) or its vehicle alone (100 microl/day distilled water for 14 days subcutaneously). On day 14 the animals were killed. Burn areas were used for histologic examination, evaluation of neoangiogenesis by immunohistochemistry, and expression (Western blot) of the specific endothelial marker CD31 as well as quantification of microvessel density, measurement of VEGF wound content (enzyme-linked immunosorbent assay), expression (Western blot) of endothelial and inducible nitric oxide synthases, and determination of wound nitric oxide (NO) products. rHuEPO increased burn wound reepithelialization and reduced the time to final wound closure. These effects were completely abated by a passive immunization with specific antibodies against erythropoietin. rHuEPO improved healing of burn wound through increased epithelial proliferation, maturation of the extracellular matrix, and angiogenesis. The hematopoietic factor augmented neoangiogenesis as suggested by the marked increase in microvessel density and by the robust expression of the specific endothelial marker CD31. Furthermore, rHuEPO enhanced the wound content of VEGF caused a marked expression of endothelial and inducible nitric oxide synthases and increased wound content of nitric oxide products. Our study suggests that rHuEPO may be an effective therapeutic approach to improve clinical outcomes after thermal injury.

  12. Effects of recombinant human erythropoietin injections on physical self in endurance athletes.

    Science.gov (United States)

    Ninot, Grégory; Connes, Philippe; Caillaud, Corrine

    2006-04-01

    This study examined the time course of mean self-esteem and physical self scores in three groups: male endurance athletes treated with recombinant human erythropoietin (rHuEPO group, n = 6), a placebo group (n = 5) injected with a sodium chloride solution and a control group who did not receive any injection (n = 6). Each participant completed the Physical Self Inventory twice a day (between 07.00 and 09.00 h and between 19.00 and 21.00 h). Using a 10 cm visual analog scale, the participants assessed global self-esteem, physical self-worth and the sub-domains of physical condition, sport competence, attractive body and physical strength (Fox & Corbin, 1989). This was conducted over three consecutive periods: in the 2 weeks before the course of injections, during the 6 weeks of injections and for 4 weeks after the injections. Aerobic capacity was assessed before and after 4 weeks of treatment. The results showed a significant increase in aerobic physical fitness in the rHuEPO group and a significant increase in perceived physical condition and physical strength scores at the end of treatment. The main psychological result was that endurance athletes were highly sensitive to the effects of rHuEPO on physical fitness. The perception of increased physical condition may lead to a stronger commitment to training. The rHuEPO injections presented a dangerous hedonic effect linked to endurance training. These results confirm the need to tackle rHuEPO abuse at any time during the training season.

  13. The evaluation of lyophilized polymer matrices for administering recombinant human bone morphogenetic protein-2.

    Science.gov (United States)

    Duggirala, S S; Rodgers, J B; DeLuca, P P

    1996-07-01

    Novel unitary devices, prepared by lyophilization of viscous solutions of sodium carboxymethylcellulose (CMC) and methylcellulose (MC), were evaluated as sustained-release delivery systems for recombinant human bone morphogenetic protein-2 (rhBMP-2). In vitro characterization of the unitary devices, which contained rhBMP-2-loaded poly (d,l lactide-co-glycolide) (PLGA) bioerodible particles (BEPs), was conducted over a 2-month period. Determinations included buffer uptake, mass and molecular weight loss and rhBMP-2 release from the unitary devices. CMC devices imbibed approximately 16 times their weight of buffer, while with MC, equilibrium uptake was approximately 6 times the dry weight of the devices. Overall mass loss percentages were approximately 55 and 35%, respectively, for CMC and MC devices. rhBMP-2 release from the devices was essentially a triphasic process: an initial phase during which "free" protein (rhBMP-2 present on the surface and within the pores of the PLGA BEPs) was released, a lag period during which no release was discerned, and then release of "bound" rhBMP-2 (protein adsorbed to the BEPs). The release of bound protein correlated with the mass loss of the polymer which began after 3 weeks. Release from the unitary devices was lower than that from the BEPs alone, due to a retardation effect of the gelled CMC/MC polymers. In rabbits in which full-thickness cranial bone defects were created, the implants were well tolerated and induced significant new bone growth during an 8-week evaluation period. The CMC devices appear to have induced bone earlier (at 2 weeks), but this did not affect eventual 8-week results. CMC devices without rhBMP-2 appeared to provide some bone conduction, in contrast to the blank MC devices.

  14. Prolonged Particulate Hexavalent Chromium Exposure Suppresses Homologous Recombination Repair in Human Lung Cells.

    Science.gov (United States)

    Browning, Cynthia L; Qin, Qin; Kelly, Deborah F; Prakash, Rohit; Vanoli, Fabio; Jasin, Maria; Wise, John Pierce

    2016-09-01

    Genomic instability is one of the primary models of carcinogenesis and a feature of almost all cancers. Homologous recombination (HR) repair protects against genomic instability by maintaining high genomic fidelity during the repair of DNA double strand breaks. The defining step of HR repair is the formation of the Rad51 nucleofilament, which facilitates the search for a homologous sequence and invasion of the template DNA strand. Particulate hexavalent chromium (Cr(VI)), a human lung carcinogen, induces DNA double strand breaks and chromosome instability. Since the loss of HR repair increases Cr(VI)-induced chromosome instability, we investigated the effect of extended Cr(VI) exposure on HR repair. We show acute (24 h) Cr(VI) exposure induces a normal HR repair response. In contrast, prolonged (120 h) exposure to particulate Cr(VI) inhibited HR repair and Rad51 nucleofilament formation. Prolonged Cr(VI) exposure had a profound effect on Rad51, evidenced by reduced protein levels and Rad51 mislocalization to the cytoplasm. The response of proteins involved in Rad51 nuclear import and nucleofilament formation displayed varying responses to prolonged Cr(VI) exposure. BRCA2 formed nuclear foci after prolonged Cr(VI) exposure, while Rad51C foci formation was suppressed. These results suggest that particulate Cr(VI), a major chemical carcinogen, inhibits HR repair by targeting Rad51, causing DNA double strand breaks to be repaired by a low fidelity, Rad51-independent repair pathway. These results further enhance our understanding of the underlying mechanism of Cr(VI)-induced chromosome instability and thus, carcinogenesis. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  15. Recombinant erythropoietin in humans has a prolonged effect on circulating erythropoietin isoform distribution.

    Directory of Open Access Journals (Sweden)

    Niels Jacob Aachmann-Andersen

    Full Text Available The membrane-assisted isoform immunoassay (MAIIA quantitates erythropoietin (EPO isoforms as percentages of migrated isoforms (PMI. We evaluated the effect of recombinant human EPO (rhEPO on the distribution of EPO isoforms in plasma in a randomized, placebo-controlled, double-blinded, cross-over study. 16 healthy subjects received either low-dose Epoetin beta (5000 IU on days 1, 3, 5, 7, 9, 11 and 13; high-dose Epoetin beta (30.000 IU on days 1, 2 and 3 and placebo on days 5, 7, 9, 11 and 13; or placebo on all days. PMI on days 4, 11 and 25 was determined by interaction of N-acetyl glucosamine with the glycosylation dependent desorption of EPO isoforms. At day 25, plasma-EPO in both rhEPO groups had returned to values not different from the placebo group. PMI with placebo, reflecting the endogenous EPO isoforms, averaged 82.5 (10.3 % (mean (SD. High-dose Epoetin beta decreased PMI on days 4 and 11 to 31.0 (4.2% (p<0.00001 and 45.2 (7.3% (p<0.00001. Low-dose Epoetin beta decreased PMI on days 4 and 11 to 46.0 (12.8% (p<0.00001 and 46.1 (10.4% (p<0.00001. In both rhEPO groups, PMI on day 25 was still decreased (high-dose Epoetin beta: 72.9 (19.4% (p=0.029; low-dose Epoetin beta: 73.1 (17.8% (p=0.039. In conclusion, Epoetin beta leaves a footprint in the plasma-EPO isoform pattern. MAIIA can detect changes in EPO isoform distribution up til at least three weeks after administration of Epoetin beta even though the total EPO concentration has returned to normal.

  16. A repeated injection of polyethyleneglycol-conjugated recombinant human butyrylcholinesterase elicits immune response in mice

    International Nuclear Information System (INIS)

    Chilukuri, Nageswararao; Sun Wei; Parikh, Kalpana; Naik, Ramachandra S.; Tang Lin; Doctor, Bhupendra P.; Saxena, Ashima

    2008-01-01

    Human serum butyrylcholinesterase (Hu BChE) serves as an efficacious bioscavenger of highly toxic organophosphorus (OP) compounds. Since there is a concern that the supply of native Hu BChE may be limited, monomeric and tetrameric forms of recombinant Hu BChE (rHu BChE) were evaluated as replacements and found that they lacked sufficient stability in vivo. However, their in vivo stability could be significantly prolonged by conjugation with polyethyleneglycol-20K (PEG) suggesting that monomeric and tetrameric PEG-rHu BChE could function as bioscavengers. Here, the immunogenicity of PEG-rHu BChE was evaluated in mice following two injections given four weeks apart. In addition to pharmacokinetic parameters, such as mean residence time, maximal concentration, time to reach the maximal concentration, elimination half-life and area under the plasma concentration-time curve extrapolated to infinity, the presence of circulating anti-rHu BChE antibodies was also determined. Although the pharmacokinetic parameters were significantly improved for the first injection of monomeric and tetrameric PEG-rHu BChEs, they were much lower for the second injection. Anti-rHu BChE antibodies were detected in the blood of mice following the first and second enzyme injections and their levels were approximately higher by 5-fold and 2-fold in mice injected with monomeric and tetrameric PEG-rHu BChEs as compared to mice injected with unconjugated enzymes. The findings that the rapid clearance of a repeat injection of PEG-rHu BChEs in mice which coincides with the presence of circulating anti-rHu BChE antibodies suggest that PEG conjugation prolonged the circulatory stability of rHu BChE but failed to eliminate its immunogenicity in mice

  17. Tissue eosinophilia induced by recombinant human interleukin-5 in the hamster cheek pouch membrane

    Directory of Open Access Journals (Sweden)

    M. Minnicozzi

    1995-01-01

    Full Text Available Interleukin-5 (IL-5 is a cytokine that preferentially effects the development and function of eosinophils, and is considered important in the pathophysiology of allergic inflammation. In this study, we evaluated the ability of recombinant human IL-5 (rHu IL-5 to promote tissue eosinophilia and the importance of this eosinophilia to pathological alterations in vascular function. Repetitive subcutaneous administration for 18 days of rHu IL-5 resulted in a 7-fold increase in the number of eosinophils found in the ipsilateral hamster cheek pouch membrane. The contralateral cheek pouch membrane and peritoneum of these animals showed lesser but significant elevations in the number of eosinophils. In contrast, denatured rHu IL-5 did not elevate eosinophils in these tissues. Through the use of intravital microscopy and fluorometric analysis, rHu IL-5 treated hamster cheek pouch membranes were evaluated for alterations in microvascular permeability, using plasma clearance of FITC-dextran 150 as an index. Despite promoting a prominent tissue eosinophilia, the repetitive subcutaneous injections of rHu IL-5 did not alter the clearance of FITC-dextran 150. Topical application of rHu IL-5 to the cheek pouch, also, had no effect on the clearance of FITC-dextran 150. Immunofluorescence observations using an antibody to the granule protein, eosinophil peroxidase, indicated that the recruited cells had not degranulated. Our results support the importance of IL-5 in the recruitment of tissue eosinophils, but further stimulation is probably required to cause degranulation of these cells and the ensuing tissue damage.

  18. Pharmacokinetics and tissue distribution of recombinant human tumor necrosis factor-alpha in mice

    International Nuclear Information System (INIS)

    Ferraiolo, B.L.; Moore, J.A.; Crase, D.; Gribling, P.; Wilking, H.; Baughman, R.A.

    1988-01-01

    The serum pharmacokinetics and the major organs of accumulation of recombinant human tumor necrosis factor-alpha (rHuTNF) were determined in BDF1 mice after intravenous and intramuscular administration. Serum concentrations of immunoreactive protein were determined by enzyme-linked immunosorbent assay, and radioactivity was quantitated by beta and gamma scintigraphy. The serum pharmacokinetics of labeled and unlabeled rHuTNF were identical when administered by the intravenous route. After intravenous doses of 165 to 320 micrograms/kg, the clearance was 2.9-3.6 ml/hr, the initial volume of distribution was 1.4-1.6 ml (70-80 ml/kg), and the half-life was 18.5-19.2 min. Intramuscular administration of 320 micrograms/kg resulted in a peak serum concentration of 112 ng/ml. The time of the peak concentration was 1 hr, and the bioavailability of the intramuscular dose was 12%. The data suggest that the disposition of this protein may be biexponential. If this is the case, the terminal phase would appear to account for less than 1% of the total AUC. Since serum concentrations in the terminal phase are at the sensitivity limit of the assay, a single half-life is reported. 125I-Labeled and metabolically labeled 3H-rHuTNF were used to examine tissue distribution. After intravenous 125I-rHuTNF administration, the rank order of accumulation of the 125I-radiolabel in the major organs (per cent dose per organ over 1440 min) was: liver greater than kidney greater than lung greater than heart greater than spleen. This rank order of accumulation was confirmed by intravenous 3H-rHuTNF administration

  19. Dendritic Cell Activity Driven by Recombinant Mycobacterium bovis BCG Producing Human IL-18, in Healthy BCG Vaccinated Adults.

    Science.gov (United States)

    Szpakowski, Piotr; Biet, Franck; Locht, Camille; Paszkiewicz, Małgorzata; Rudnicka, Wiesława; Druszczyńska, Magdalena; Allain, Fabrice; Fol, Marek; Pestel, Joël; Kowalewicz-Kulbat, Magdalena

    2015-01-01

    Tuberculosis remains an enormous global burden, despite wide vaccination coverage with the Bacillus Calmette-Guérin (BCG), the only vaccine available against this disease, indicating that BCG-driven immunity is insufficient to protect the human population against tuberculosis. In this study we constructed recombinant BCG producing human IL-18 (rBCGhIL-18) and investigated whether human IL-18 produced by rBCGhIL-18 modulates DC functions and enhances Th1 responses to mycobacterial antigens in humans. We found that the costimulatory CD86 and CD80 molecules were significantly upregulated on rBCGhIL-18-infected DCs, whereas the stimulation of DCs with nonrecombinant BCG was less effective. In contrast, both BCG strains decreased the DC-SIGN expression on human DCs. The rBCGhIL-18 increased IL-23, IL-10, and IP-10 production by DCs to a greater extent than nonrecombinant BCG. In a coculture system of CD4(+) T cells and loaded DCs, rBCGhIL-18 favoured strong IFN-γ but also IL-10 production by naive T cells but not by memory T cells. This was much less the case for nonrecombinant BCG. Thus the expression of IL-18 by recombinant BCG increases IL-23, IP-10, and IL-10 expression by human DCs and enhances their ability to induce IFN-γ and IL-10 expression by naive T cells, without affecting the maturation phenotype of the DCs.

  20. Dendritic Cell Activity Driven by Recombinant Mycobacterium bovis BCG Producing Human IL-18, in Healthy BCG Vaccinated Adults

    Directory of Open Access Journals (Sweden)

    Piotr Szpakowski

    2015-01-01

    Full Text Available Tuberculosis remains an enormous global burden, despite wide vaccination coverage with the Bacillus Calmette-Guérin (BCG, the only vaccine available against this disease, indicating that BCG-driven immunity is insufficient to protect the human population against tuberculosis. In this study we constructed recombinant BCG producing human IL-18 (rBCGhIL-18 and investigated whether human IL-18 produced by rBCGhIL-18 modulates DC functions and enhances Th1 responses to mycobacterial antigens in humans. We found that the costimulatory CD86 and CD80 molecules were significantly upregulated on rBCGhIL-18-infected DCs, whereas the stimulation of DCs with nonrecombinant BCG was less effective. In contrast, both BCG strains decreased the DC-SIGN expression on human DCs. The rBCGhIL-18 increased IL-23, IL-10, and IP-10 production by DCs to a greater extent than nonrecombinant BCG. In a coculture system of CD4+ T cells and loaded DCs, rBCGhIL-18 favoured strong IFN-γ but also IL-10 production by naive T cells but not by memory T cells. This was much less the case for nonrecombinant BCG. Thus the expression of IL-18 by recombinant BCG increases IL-23, IP-10, and IL-10 expression by human DCs and enhances their ability to induce IFN-γ and IL-10 expression by naive T cells, without affecting the maturation phenotype of the DCs.

  1. Involvement of ERK, Bcl-2 family and caspase 3 in recombinant human activin A-induced apoptosis in A549

    International Nuclear Information System (INIS)

    Wang Baiding; Feng Yuling; Song Xingbo; Liu Qingqing; Ning Yunye; Ou Xuemei; Yang Jie; Zhang Xiaohong; Wen, Fuqiang

    2009-01-01

    Background: Activins are members of the transforming growth factor-β (TGF-β) superfamily. Previous studies have shown that activin A may have a central role in regulating both apoptosis and proliferation. However, direct studies of recombination human activin A on human NSCLC A549 cells have not yet been reported. The purpose of this study was to investigate whether activin A could induce apoptosis in A549 cells and the possible mechanisms via which it worked. Methods: Cellular apoptosis induced by activin A was detected by TUNEL assay and the levels of protein expression were detected by western blot. Results: Recombination human activin A induced apoptosis in human NSCLC A549 cells in a concentrate-dependent manner. Activin A-induced A549 apoptosis was accompanied by the up-regulation of Bax, Bad and Bcl-Xs and down-regulation of Bcl-2. Moreover, activin A treatment increased the expression of its typeII receptors, activated ERK and caspase 3 in A549. These results clearly demonstrate that the induction of apoptosis by activin-A involves multiple cellular/molecular pathways and strongly suggest that pro- and anti-apoptotic Bcl-2 family proteins and caspase 3 participate in activin A-induced apoptotic process in A549 cells. On the other hand, activin A treatment had little effect on primary human small airway epithelial cells (SAECs). Conclusion: Recombination human activin A induced apoptosis in A549 cells, at least partially, through ERK and mitochondrial pathway. The result that activin A did not affect the normal SAEC revealed activin A might be considered as a potential anticancer agent and worthy of further studies

  2. Environmental radioiodine in thyroids of grazing animals

    International Nuclear Information System (INIS)

    Van Middlesworth, L.

    1990-01-01

    Animal thyroids are excellent indicators of environmental contamination from radioiodine, a fact largely unappreciated before 1954. From continuous monitoring of excised thyroids in Nal well-detectors, we know that high-altitude releases of mixed fission products caused increases of 10 3 to 10 5 times the minimal detectable level of 131 I (3.7 mBq/g) in sheep thyroids from the United States, Europe, Asia, Australia, and New Zealand. Within 6 weeks after high-altitude releases, sheep thyroids from different continents of the same hemisphere contained comparable concentrations of 131 I: more than 10 4 times greater concentrations than cow's milk and 5 x 10 3 times greater concentrations than human thyroids from similar areas. In contrast, during the low-altitude Chernobyl release, average 131 I concentrations in animal thyroids, at distances greater than 1,200 km from Chernobyl, were distributed as the inverse square of the distance from the source. 129 I concentrations measured since 1984 with a germanium well-detector in thyroids of sheep slaughtered in Birmingham, England, ranged from 3 to 50 mBq/g organ weight. Similar quantities of 125 I were present intermittently in thyroids of sheep slaughtered in Birmingham since 1986. The source of the 125 I, which is not a fission product, is not clear. For comparison, deer thyroids from the Savannah River Reservation, USA, contained 10 to 7 x 10 5 mBq 129 I/g. Although this isotope is not a biological hazard, it is a long-lived tracer of fission products

  3. Analysis of the kinetic mechanism of recombinant human isoprenylcysteine carboxylmethyltransferase (Icmt

    Directory of Open Access Journals (Sweden)

    Baron Rudi A

    2004-12-01

    Full Text Available Abstract Background Isoprenylcysteine carboxyl methyltransferase (Icmt is the third of three enzymes that posttranslationally modify proteins that contain C-terminal CaaX motifs. The processing of CaaX proteins through this so-called prenylation pathway via a route initiated by addition of an isoprenoid lipid is required for both membrane targeting and function of the proteins. The involvement of many CaaX proteins such as Ras GTPases in oncogenesis and other aberrant proliferative disorders has led to the targeting of the enzymes involved in their processing for therapeutic development, necessitating a detailed understanding of the mechanisms of the enzymes. Results In this study, we have investigated the kinetic mechanism of recombinant human Icmt. In the reaction catalyzed by Icmt, S-adenosyl-L-methionine (AdoMet provides the methyl group that is transferred to the second substrate, the C-terminal isoprenylated cysteine residue of a CaaX protein, thereby generating a C-terminal prenylcysteine methyl ester on the protein. To facilitate the kinetic analysis of Icmt, we synthesized a new small molecule substrate of the enzyme, biotin-S-farnesyl-L-cysteine (BFC. Initial kinetic analysis of Icmt suggested a sequential mechanism for the enzyme that was further analyzed using a dead end competitive inhibitor, S-farnesylthioacetic acid (FTA. Inhibition by FTA was competitive with respect to BFC and uncompetitive with respect to AdoMet, indicating an ordered mechanism with SAM binding first. To investigate the order of product dissociation, product inhibition studies were undertaken with S-adenosyl-L-homocysteine (AdoHcy and the N-acetyl-S-farnesyl-L-cysteine methylester (AFCME. This analysis indicated that AdoHcy is a competitive inhibitor with respect to AdoMet, while AFCME shows a noncompetitive inhibition with respect to BFC and a mixed-type inhibition with respect to AdoMet. These studies established that AdoHcy is the final product released, and

  4. Polymyxin B as inhibitor of LPS contamination of Schistosoma mansoni recombinant proteins in human cytokine analysis

    Directory of Open Access Journals (Sweden)

    Pacífico Lucila G

    2007-01-01

    Full Text Available Abstract Background Recombinant proteins expressed in Escherichia coli vectors are generally contaminated with endotoxin. In this study, we evaluated the ability of Polymyxin B to neutralize the effect of LPS present as contaminant on Schistosoma mansoni recombinant proteins produced in E. coli in inducing TNF-α and IL-10. Peripheral blood mononuclear cells from individuals chronically infected with S. mansoni were stimulated in vitro with recombinant Sm22.6, Sm14 and P24 antigens (10 μg/mL in the presence of Polymyxin B (10 μg/mL. Results The levels of cytokines were measured using ELISA. There was greater than 90 % reduction (p S. mansoni recombinant proteins in the presence of Polymyxin B, a reduction in the levels of TNF-α and IL-10 was also observed. However, the percentage of reduction was lower when compared to the cultures stimulated with LPS, probably because these proteins are able to induce the production of these cytokines by themselves. Conclusion This study showed that Polymyxin B was able to neutralize the effect of endotoxin, as contaminant in S. mansoni recombinant antigens produced in E. coli, in inducing TNF-α and IL-10 production.

  5. Government-industry conference on airborne radioiodine

    International Nuclear Information System (INIS)

    Burchsted, C.A.

    1975-01-01

    The Working Group on Airborne Radioiodine met at AEC Headquarters on March 28, 1974. Dr. Alex Perge gave the introduction for the Division of Waste Management and Transportation, noting the Commission hopes that private industry will take a bigger share in the future in funding and initiating needed research; that there should be a greater effort in the direction of reducing the quantity of material that becomes contaminated as an avenue toward reducing the airborne radioiodine problem, and toward reducing the waste generated to a form suitable for direct storage; and that the Commission must ensure valid bases for future regulations governing airborne releases and contamination. Dr. First discussed the background of the review committee and its outgrowth from the earlier organization meeting. He noted that its function will be the coordination of efforts concerned with the radioiodine problem and the dissemination of information and research data. A major objective of this meeting was to identify subjects for discussion at the Government-Industry Conference of Adsorbers and Adsorbents which will be held in conjunction with the 13th AEC Air Cleaning Conference in August. Mr. Dempsey noted that the gaseous effluent program had been inherited by WMT from the Division of Operational Safety, and that an important function of these continuing meetings of the Working Group will be to guide WMT in the expenditure of funds and assignment of research related to the radioiodine problem. (U.S.)

  6. Immobilization of radioiodine in synthetic boracite

    Science.gov (United States)

    Babad, H.; Strachan, D.M.

    1982-09-23

    A nuclear waste storage product is disclosed in which radioiodine is incorporated in a synthetic boracite. The boracite may be prepared by reacting a transition metal iodide with an alkali horate under mild hydrothermal conditions, drying the reaction product, and then hot pressing.

  7. UV-dependent production of 25-hydroxyvitamin D2 in the recombinant yeast cells expressing human CYP2R1

    International Nuclear Information System (INIS)

    Yasuda, Kaori; Endo, Mariko; Ikushiro, Shinichi; Kamakura, Masaki; Ohta, Miho; Sakaki, Toshiyuki

    2013-01-01

    Highlights: •We produce 25-hydroxyvitamin D in the recombinant yeast expressing human CYP2R1. •Vitamin D2 is produced in yeast from endogenous ergosterol with UV irradiation. •We produce 25-hydroxyvitamin D2 in the recombinant yeast without added substrate. -- Abstract: CYP2R1 is known to be a physiologically important vitamin D 25-hydroxylase. We have successfully expressed human CYP2R1 in Saccharomyces cerevisiae to reveal its enzymatic properties. In this study, we examined production of 25-hydroxylated vitamin D using whole recombinant yeast cells that expressed CYP2R1. When vitamin D 3 or vitamin D 2 was added to the cell suspension of CYP2R1-expressing yeast cells in a buffer containing glucose and β-cyclodextrin, the vitamins were converted into their 25-hydroxylated products. Next, we irradiated the cell suspension with UVB and incubated at 37 °C. Surprisingly, the 25-hydroxy vitamin D 2 was produced without additional vitamin D 2 . Endogenous ergosterol was likely converted into vitamin D 2 by UV irradiation and thermal isomerization, and then the resulting vitamin D 2 was converted to 25-hydroxyvitamin D 2 by CYP2R1. This novel method for producing 25-hydroxyvitamin D 2 without a substrate could be useful for practical purposes

  8. Activity of the Human Rhinovirus 3C Protease Studied in Various Buffers, Additives and Detergents Solutions for Recombinant Protein Production.

    Directory of Open Access Journals (Sweden)

    Raheem Ullah

    Full Text Available Proteases are widely used to remove affinity and solubility tags from recombinant proteins to avoid potential interference of these tags with the structure and function of the fusion partner. In recent years, great interest has been seen in use of the human rhinovirus 3C protease owing to its stringent sequence specificity and enhanced activity. Like other proteases, activity of the human rhinovirus 3C protease can be affected in part by the buffer components and additives that are generally employed for purification and stabilization of proteins, hence, necessitate their removal by tedious and time-consuming procedures before proteolysis can occur. To address this issue, we examined the effect of elution buffers used for common affinity based purifications, salt ions, stability/solubility and reducing agents, and detergents on the activity of the human rhinovirus 3C protease using three different fusion proteins at 4°C, a temperature of choice for purification of many proteins. The results show that the human rhinovirus 3C protease performs better at 4°C than the frequently used tobacco etch virus protease and its activity was insensitive to most of the experimental conditions tested. Though number of fusion proteins tested is limited, we expect that these finding will facilitate the use of the human rhinovirus 3C protease in recombinant protein production for pharmaceutical and biotechnological applications.

  9. Activity of the Human Rhinovirus 3C Protease Studied in Various Buffers, Additives and Detergents Solutions for Recombinant Protein Production.

    Science.gov (United States)

    Ullah, Raheem; Shah, Majid Ali; Tufail, Soban; Ismat, Fouzia; Imran, Muhammad; Iqbal, Mazhar; Mirza, Osman; Rhaman, Moazur

    2016-01-01

    Proteases are widely used to remove affinity and solubility tags from recombinant proteins to avoid potential interference of these tags with the structure and function of the fusion partner. In recent years, great interest has been seen in use of the human rhinovirus 3C protease owing to its stringent sequence specificity and enhanced activity. Like other proteases, activity of the human rhinovirus 3C protease can be affected in part by the buffer components and additives that are generally employed for purification and stabilization of proteins, hence, necessitate their removal by tedious and time-consuming procedures before proteolysis can occur. To address this issue, we examined the effect of elution buffers used for common affinity based purifications, salt ions, stability/solubility and reducing agents, and detergents on the activity of the human rhinovirus 3C protease using three different fusion proteins at 4°C, a temperature of choice for purification of many proteins. The results show that the human rhinovirus 3C protease performs better at 4°C than the frequently used tobacco etch virus protease and its activity was insensitive to most of the experimental conditions tested. Though number of fusion proteins tested is limited, we expect that these finding will facilitate the use of the human rhinovirus 3C protease in recombinant protein production for pharmaceutical and biotechnological applications.

  10. A Randomized Case-Controlled Study of Recombinant Human Granulocyte Colony Stimulating Factor for the Treatment of Sepsis in Preterm Neutropenic Infants

    OpenAIRE

    Aktaş, Doğukan; Demirel, Bilge; Gürsoy, Tuğba; Ovalı, Fahri

    2015-01-01

    To investigate the efficacy and safety of recombinant human granulocyte colony-stimulating factor, recombinant human granulocyte-macrophage colony-stimulating factor (rhG-CSF) to treat sepsis in neutropenic preterm infants. Methods: Fifty-six neutropenic preterm infants with suspected or culture-proven sepsis hospitalized in Zeynep Kamil Maternity and Children's Educational and Training Hospital, Kozyatağı/Istanbul, Turkey between January 2008 and January 2010 were enrolled. Patients were ...

  11. Genetic analysis of a novel human adenovirus with a serologically unique hexon and a recombinant fiber gene.

    Directory of Open Access Journals (Sweden)

    Elizabeth B Liu

    Full Text Available In February of 1996 a human adenovirus (formerly known as Ad-Cor-96-487 was isolated from the stool of an AIDS patient who presented with severe chronic diarrhea. To characterize this apparently novel pathogen of potential public health significance, the complete genome of this adenovirus was sequenced to elucidate its origin. Bioinformatic and phylogenetic analyses of this genome demonstrate that this virus, heretofore referred to as HAdV-D58, contains a novel hexon gene as well as a recombinant fiber gene. In addition, serological analysis demonstrated that HAdV-D58 has a different neutralization profile than all previously characterized HAdVs. Bootscan analysis of the HAdV-D58 fiber gene strongly suggests one recombination event.

  12. Evidence for repair of ultraviolet light-damaged herpes virus in human fibroblasts by a recombination mechanism

    International Nuclear Information System (INIS)

    Hall, J.D.; Featherston, J.D.; Almy, R.E.

    1980-01-01

    Human cells were either singly or multiply infected with herpes simplex virus (HSV-1) damaged by ultraviolet (uv) light, and the fraction of cells able to produce infectious virus was measured. The fraction of virus-producing cells was considerably greater for multiply infected cells than for singly infected cells at each uv dose examined. These high survival levels of uv-irradiated virus in multiply infected cells demonstrated that multiplicity-dependent repair, possibly due to genetic exchanges between damaged HSV-1 genomes, was occurring in these cells. To test whether uv light is recombinogenic for HSV-1, the effect of uv irradiation on the yield of temperature-resistant viral recombinants in cells infected with pairs of temperature-sensitive mutants was also investigated. The results of these experiments showed that the defective functions in these mutant host cells are not required for multiplicity-dependent repair or uv-stimulated viral recombination in herpes-infected cells

  13. Near-lethal respiratory failure after recombinant human thyroid-stimulating hormone use in a patient with metastatic thyroid carcinoma.

    Science.gov (United States)

    Goffman, Thomas; Ioffe, Vladimir; Tuttle, Michael; Bowers, John T; Mason, M Elizabeth

    2003-08-01

    A patient with widely metastatic differentiated thyroid cancer who had been heavily pretreated with (131)I was given recombinant human thyroid stimulating hormone (rhTSH) prior to (131)I treatment. Clinical and physical data from both this case and the literature suggest that the recombinant hormone, not the (131)I, may have caused a significant portion of the tumor swelling, which in turn was the most likely cause of the patient's symptoms. The potential effect of (131)I-induced tumor swelling and direct radiation effect on the lung is also analyzed. We review the potential hazards associated with rhTSH in patients with metastasis and propose means of minimizing this risk.

  14. Recombinant human TSH and ablation of post-surgical thyroid remnants in differentiated thyroid cancer: the effect of pre-treatment with furosemide and furosemide plus lithium

    Energy Technology Data Exchange (ETDEWEB)

    Barbaro, Daniele; Lapi, Paola; Pasquini, Cristina; Orsini, Paola; Turco, Anna [General Hospital of Livorno, Endocrinology Unit, Livorno (Italy); Grosso, Mariano; Boni, Giuseppe; Mariani, Giuliano [University of Pisa Medical School, Regional Center of Nuclear Medicine, Pisa (Italy); Meucci, Giuseppe [General Hospital of Livorno, Division of General Surgery, Livorno (Italy); Marzola, Maria Cristina; Rubello, Domenico [' Santa Maria della Misericordia' Hospital, Department of Nuclear Medicine, PET Centre, Rovigo (Italy); Berti, Piero; Miccoli, Paolo [University of Pisa Medical School, Department of Surgery, Pisa (Italy)

    2010-02-15

    Recombinant human TSH (rhTSH) can be used for post-surgical radioiodine (I-131) thyroid remnants ablation in differentiated thyroid cancer (DTC) patients after surgery. Debate exists in literature about the optimal amount of I-131 that should be given for obtaining an effective ablation and about the role of iodine pool during treatment. Therefore, the aim of the present study was to assess whether I-131 ablation during rhTSH stimulus can be improved by reducing the circulating iodine pool and by increasing thyroid cell uptake and retention of I-131 obtained by administering furosemide and lithium. A total of 201 consecutive DTC patients were entered in the study: they were treated by total thyroidectomy and I-131 therapy during rhTSH stimulus to ablate thyroid remnants. Patients were divided into two groups according to the TNM stage: group 1 included patients in stage I-II who were treated with a low 30-mCi I-131 dose, while group 2 included patients in stage III-IV who were treated by a high 100-mCi I-131 dose. Moreover, both groups were further subdivided into three subgroups. Subgroup (a) included 45 patients from group 1 and 22 from group 2: they were treated with I-131 under rhTSH stimulus, following a short 4-day withdrawal of L-thyroxine (LT4). Subgroup (b) included 45 patients from group 1 and 22 from group 2: they were treated with I-131 under rhTSH stimulus, following a short 4-day withdrawal of L-T4, and after furosemide administration (25 mg/day orally) during the 3 days before I-131. Subgroup (c) included 45 patients from group 1 and 22 from group 2: they were treated with I-131 under rhTSH stimulus, following a short 4-day L-T4 withdrawal, and after administration of furosemide (25 mg/day orally) during the 3 days prior I-131 and lithium (450 mg/day orally) during the 3 days following I-131. Another group (group 3) of 20 patients characterized by a very low-risk cancer (unifocal tumor <1.0 cm in diameter, without extra-capsular extension, N0) was

  15. Recombinant human tripeptidyl peptidase-1 infusion to the monkey CNS: Safety, pharmacokinetics, and distribution

    Energy Technology Data Exchange (ETDEWEB)

    Vuillemenot, Brian R., E-mail: bvuillemenot@bmrn.com [BioMarin Pharmaceutical Inc., Novato, CA (United States); Kennedy, Derek [BioMarin Pharmaceutical Inc., Novato, CA (United States); Reed, Randall P.; Boyd, Robert B. [Northern Biomedical Research, Inc., Muskegon, MI (United States); Butt, Mark T. [Tox Path Specialists, LLC, Hagerstown, MD (United States); Musson, Donald G.; Keve, Steve; Cahayag, Rhea; Tsuruda, Laurie S.; O' Neill, Charles A. [BioMarin Pharmaceutical Inc., Novato, CA (United States)

    2014-05-15

    CLN2 disease is caused by deficiency in tripeptidyl peptidase-1 (TPP1), leading to neurodegeneration and death. The safety, pharmacokinetics (PK), and CNS distribution of recombinant human TPP1 (rhTPP1) were characterized following a single intracerebroventricular (ICV) or intrathecal-lumbar (IT-L) infusion to cynomolgus monkeys. Animals received 0, 5, 14, or 20 mg rhTPP1, ICV, or 14 mg IT-L, in artificial cerebrospinal fluid (aCSF) vehicle. Plasma and CSF were collected for PK analysis. Necropsies occurred at 3, 7, and 14 days post-infusion. CNS tissues were sampled for rhTPP1 distribution. TPP1 infusion was well tolerated and without effect on clinical observations or ECG. A mild increase in CSF white blood cells (WBCs) was detected transiently after ICV infusion. Isolated histological changes related to catheter placement and infusion were observed in ICV treated animals, including vehicle controls. The CSF and plasma exposure profiles were equivalent between animals that received an ICV or IT-L infusion. TPP1 levels peaked at the end of infusion, at which point the enzyme was present in plasma at 0.3% to 0.5% of CSF levels. TPP1 was detected in brain tissues with half-lives of 3–14 days. CNS distribution between ICV and IT-L administration was similar, although ICV resulted in distribution to deep brain structures including the thalamus, midbrain, and striatum. Direct CNS infusion of rhTPP1 was well tolerated with no drug related safety findings. The favorable nonclinical profile of ICV rhTPP1 supports the treatment of CLN2 by direct administration to the CNS. - Highlights: • TPP1 enzyme replacement therapy to the CNS is in development for CLN2 disease. • Toxicology, pharmacokinetics, and CNS distribution were assessed in monkeys. • TPP1 infusion directly to the brain did not result in any safety concerns. • A positive pharmacokinetic and distribution profile resulted from TPP1 infusion. • This study demonstrates the feasibility of ICV administered

  16. [Effect of recombinant human growth hormone therapy on metabolic parameters in patients with craniopharyngioma].

    Science.gov (United States)

    Mao, J F; Wang, X; Xiong, S Y; Zheng, J J; Yu, B Q; Nie, M; Wu, X Y; Qi, S T

    2017-11-14

    Objective: To investigate the effects of recombinant human growth hormone (rhGH) on metabolic parameters in patients with craniopharyngioma surgeries. Methods: Totallys 30 patients with craniopharyngioma were included in this retrospective study. They were divided into growth hormone (GH) group and control group according to whether they received rhGH therapy or not. The following parameters, including body mass index (BMI), weight, waist circumstance, transaminase, fasting blood glucose, lipid profile and high-sensitivity C-reactive protein (hsCRP) were compared after rhGH therapy for 4-6 months. Results: In GH group, patients were 18-46 (30.0±8.8) years old. The duration after craniopharyngioma surgery was (12.9±5.4) years. Before rhGH therapy, they had got sufficient thyroid and glucocorticoid hormone replacement. After rhGH therapy, the body weight decreased from (92.3±20.1) to (87.6 ±14.6) kg ( P =0.190), with a reduction of BMI from (30.1±5.9) to (28.2±3.7) kg/m(2) ( P =0.120). The waist circumference decreased from (104.4±9.4) cm to (98.8±10.6) cm ( P =0.002). Alanine aminotransferase (ALT) decreased from (52±34) to (28±19) U/L ( P =0.029), with a reduction of aspartate transaminase (AST) from (46±21) to (33±18) U/L ( P =0.035) and γ-glutamyl transpeptadase (GGT) from (59±42) to (29±15) U/L ( P =0.02). hsCRP decreased from (5.3±4.9) to (2.3±2.8) mg/L ( P =0.006) and triglyceride (TG) decreased from (1.8±0.7) to (1.5±0.6) mmol/L ( P =0.028). Fasting blood glucose, low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C) and free fat acid (FFA) were not significantly changed(all P >0.05). In the control group, the above mentioned parameters did not changed significantly during 4-6 months of observational period(all P >0.05). Conclusion: rhGH therapy improves metabolic parameters in patients after craniopharyngioma surgery by decreasing body weight, waist circumstance and fat deposit in liver, as well as

  17. [Clinical characteristics of human recombination activating gene 1 mutations in 8 immunodeficiency patients with diverse phenotypes].

    Science.gov (United States)

    Yu, G; Wang, W J; Liu, D R; Tao, Z F; Hui, X Y; Hou, J; Sun, J Q; Wang, X C

    2018-03-02

    Objective: To investigate the clinical characteristics of 8 immunodeficiency cases caused by human recombination activating gene 1 (RAG1) mutations, and to explore the relationship among genotypes, clinical manifestations and immunophenotypes. Methods: Clinical data were collected and analyzed from patients with RAG1 mutations who visited the Department of Clinical Immunology, Children's Hospital of Fudan University between October 2013 and June 2017. The data included clinical manifestations, immunophenotypes and genotypes. Results: A total of 8 patients were diagnosed with RAG1 deficiency (6 boys and 2 girls). The minimum age of onset was 2 months, and the maximum age was 4 months. The minimum age of diagnosis was 2 months, and the maximum age was 13 years. Four patients had a family history of infant death due to severe infections. Two cases were born to the same consanguineous parents. All cases had recurrent infections, including involvement of respiratory tract (8 cases), digestive tract (6 cases), urinary tract (1 case), and central nervous system (1 case). The pathogens of infection included bacteria, viruses and fungi. Rotavirus was found in 3 cases, cytomegalovirus (CMV) in 5 cases, bacillus Calmette-Guérin adverse reaction in 2 cases (1 of whom had a positive acid-fast smear from lymph node puncture fluid), fungal infection in 3 cases. One case had multiple nodular space-occupying lesions in lungs and abdominal cavity complicated with multiple bone destruction. The peripheral blood lymphocyte counts of all patients ranged between 0.1 ×10(9)/L and 3.3×10(9)/L (median, 0.65×10(9)/L). Eosinophilia was found in 3 cases (range, (0.48-1.69) ×10(9)/L). The patients were classified according to immunophenotype as severe combined immunodeficiency phenotype (4 cases), leaky severe combined immunodeficiency (2 cases), Omenn syndrome (1 case) and combined immunodeficiency (1 case) . Decreased serum IgG levels were found in 3 cases, increased serum IgM levels in

  18. Recombinant human tripeptidyl peptidase-1 infusion to the monkey CNS: Safety, pharmacokinetics, and distribution

    International Nuclear Information System (INIS)

    Vuillemenot, Brian R.; Kennedy, Derek; Reed, Randall P.; Boyd, Robert B.; Butt, Mark T.; Musson, Donald G.; Keve, Steve; Cahayag, Rhea; Tsuruda, Laurie S.; O'Neill, Charles A.

    2014-01-01

    CLN2 disease is caused by deficiency in tripeptidyl peptidase-1 (TPP1), leading to neurodegeneration and death. The safety, pharmacokinetics (PK), and CNS distribution of recombinant human TPP1 (rhTPP1) were characterized following a single intracerebroventricular (ICV) or intrathecal-lumbar (IT-L) infusion to cynomolgus monkeys. Animals received 0, 5, 14, or 20 mg rhTPP1, ICV, or 14 mg IT-L, in artificial cerebrospinal fluid (aCSF) vehicle. Plasma and CSF were collected for PK analysis. Necropsies occurred at 3, 7, and 14 days post-infusion. CNS tissues were sampled for rhTPP1 distribution. TPP1 infusion was well tolerated and without effect on clinical observations or ECG. A mild increase in CSF white blood cells (WBCs) was detected transiently after ICV infusion. Isolated histological changes related to catheter placement and infusion were observed in ICV treated animals, including vehicle controls. The CSF and plasma exposure profiles were equivalent between animals that received an ICV or IT-L infusion. TPP1 levels peaked at the end of infusion, at which point the enzyme was present in plasma at 0.3% to 0.5% of CSF levels. TPP1 was detected in brain tissues with half-lives of 3–14 days. CNS distribution between ICV and IT-L administration was similar, although ICV resulted in distribution to deep brain structures including the thalamus, midbrain, and striatum. Direct CNS infusion of rhTPP1 was well tolerated with no drug related safety findings. The favorable nonclinical profile of ICV rhTPP1 supports the treatment of CLN2 by direct administration to the CNS. - Highlights: • TPP1 enzyme replacement therapy to the CNS is in development for CLN2 disease. • Toxicology, pharmacokinetics, and CNS distribution were assessed in monkeys. • TPP1 infusion directly to the brain did not result in any safety concerns. • A positive pharmacokinetic and distribution profile resulted from TPP1 infusion. • This study demonstrates the feasibility of ICV administered

  19. Recombinant human growth hormone treatment in short children with renal disease: Our first experience

    Directory of Open Access Journals (Sweden)

    Spasojević-Dimitrijeva Brankica

    2010-01-01

    Full Text Available Introduction. Growth retardation is a hallmark of chronic illnesses such as chronic kidney disease in children, and it is associated with increased morbidity and mortality. The growth hormone (GH resistance observed in uraemia can be overcome by supraphysiological doses of exogenous GH. Objective. We would like to present our first results of recombinant human growth hormone (rhGH treatment, mainly in children on haemodialysis. Methods. Sixteen children, aged 4.5-17.1 years (mean age 11.25±3.57 with height below -2.0 standard deviation score (SDS for age or height velocity below -2.0 SDS for age, were selected to receive rhGH therapy at our Nephrology and Haemodialysis Department. Most of them were on haemodialysis (14 children with mean spent time 2.88±2.68 years (0-9 years before the initiation of rhGH therapy. One half of patients were prepubertal (8 children and the second half were in early puberty (testicular volume between 4 and 8 ml for boys and breast development B2 or B3 in girls. All patients received 28-30IU/m² rhGH per week by daily subcutaneous injection. The year before rhGH therapy served as a control period. Results. During the first year of treatment, mean height velocity in haemodialysis patients increased from 2.25 cm/year to 6.59 cm/year (p<0.0001 and in the second year it was 5.25 cm/ year (p=0.004. The mean height SDS in haemodialysis children did not improve significantly during the first year of rhGH treatment (from -3.01 SDS to -2.77 SDS, p=0.063. Neither weight nor the body mass index varied compared with the pretreatment period. Two patients developed worsened secondary hyperparathyroidism and were excluded from the study, but the relationship with rhGH remains uncertain. Conclusion. Mean height velocity significantly improved during rhGH therapy in haemodialysis patients. No significant side-effects were observed in children during three-year treatment with GH.

  20. Height outcome of the recombinant human growth hormone treatment in Turner syndrome: a meta-analysis

    Directory of Open Access Journals (Sweden)

    Ping Li

    2018-04-01

    Full Text Available Objective: This study sought to determine the effect of the recombinant human growth hormone (rhGH treatment of Turner syndrome (TS on height outcome. Methods: We searched in MEDLINE, EMBASE and Cochrane Central Register of Controlled Trials and Cochrane Database of Systematic Reviews. A literature search identified 640 records. After screening and full-text assessment, 11 records were included in the systematic review. Methodological quality was assessed using the Cochrane Risk of Bias tool. RevMan 5.3 software was used for meta-analysis. We also assessed the quality of evidence with the GRADE system. Results: Compared with controls, rhGH therapy led to increased final height (MD = 7.22 cm, 95% CI 5.27–9.18, P < 0.001, I2 = 4%; P = 0.18, height standard deviation (HtSDS (SMD = 1.22, 95% CI 0.88–1.56, P < 0.001, I2 = 49%; P = 0.14 and height velocity (HV (MD 2.68 cm/year; 95% CI 2.34, 3.02; P < 0.001, I2 = 0%; P = 0.72. There was a small increase in bone age (SMD 0.32 years; 95% CI 0.1, 0.54; P = 0.004, I2 = 73%; P = 0.02 after rhGH therapy for 12 months. What is more, the rhGH/oxandrolone combination therapy suggested greater final height (MD 2.46 cm; 95% CI 0.73, 4.18; P = 0.005, I2 = 32%; P = 0.22, increase and faster HV (SMD 1.67 cm/year; 95% CI 1.03, 2.31; P < 0.03, I2 = 80%; P < 0.001, with no significant increase in HtSDS and bone maturation compared with rhGH therapy alone. Conclusions: For TS patients, rhGH alone or with concomitant use of oxandrolone treatment had advantages on final height.

  1. Mammary radioiodine accumulation due to functional sodium iodide symporter expression in a benign fibroadenoma

    International Nuclear Information System (INIS)

    Berger, F.; Unterholzner, S.; Diebold, J.; Knesewitsch, P.; Hahn, K.; Spitzweg, C.

    2006-01-01

    The sodium iodide symporter (NIS) has been characterized to mediate the active transport of iodide not only in the thyroid gland but also in various non-thyroidal tissues, including lactating mammary gland and the majority of breast cancers, thereby offering the possibility of diagnostic and therapeutic radioiodine application in breast cancer. In this report, we present a 57-year-old patient with multifocal papillary thyroid carcinoma, who showed focal radioiodine accumulation in a lesion in the right breast on a posttherapy 131 I scan following radioiodine therapy. CT and MR-mammography showed a focal solid lesion in the right breast suggestive of a fibroadenoma, which was confirmed by histological examination. Immunostaining of paraffin-embedded tumor tissue sections using a human NIS antibody demonstrated NIS-specific immunoreactivity confined to epithelial cells of mammary ducts. In conclusion, in a thyroid cancer patient we identified a benign fibroadenoma of the breast expressing high levels of functionally active NIS protein as underlying cause of focal mammary radioiodine accumulation on a posttherapy 131 I scan. These data show for the first time that functional NIS expression is not restricted to lactating mammary gland and malignant breast tissue, but can also be detected in benign breast lesions, such as fibroadenomata of the breast

  2. A review of countermeasures to reduce radioiodine in milk of dairy animals

    International Nuclear Information System (INIS)

    Howard, B.J.; Voigt, G.; Segal, M.G.

    1996-01-01

    The most effective countermeasure for radioiodine contamination of milk is to provide dairy animals with uncontaminated feed, with the added advantage that it will be effective for other radionuclides in the fallout. Another effective response is to process the milk into storable dairy products for an appropriate length of time to allow for physical decay. The use of additives given to ruminants to reduce radioiodine in milk is an alternative countermeasure which could be effective. Stable iodine administration is a practically feasible option which has the potential to reduce radioiodine levels in milk by at most a factor of three. Stable iodine supplementation should be at sufficiently high rates to be effective (and at least 1 g d -1 for dairy cows), particularly for ruminants already receiving high amounts of iodine in the diet. Currently available data are inadequate to recommend a suitable stable iodine administration rate for different species of ruminants. Other compounds, such as perchlorate and thiocyanate, also reduce the transfer to radioiodine to milk (and thyroid). Some of these compounds seem to be potentially equally as effective as stable iodine. However, currently there is inadequate information on their effectiveness and possible toxicity to both ruminants and humans for these compounds to be considered as suitable countermeasure additives. 85 refs., 2 figs., 3 tabs

  3. Production, purification and characterization of recombinant human antithrombin III by Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Maheswara Reddy Mallu

    2016-07-01

    Conclusions: The simple, cost-effective and economically viable nature of the process used in the present study for the production of rhAT will be highly beneficial for the healthcare sector. This may also be used to produce other value-added therapeutic recombinant proteins expressed in S. cerevisiae, with greater effectiveness and ease.

  4. Aberrant recombination and repair during immunoglobulin class switching in BRCA1-deficient human B cells

    DEFF Research Database (Denmark)

    Björkman, Andrea; Qvist, Per; Du, Likun

    2015-01-01

    of long microhomologies was found at recombination junctions derived from E3 ubiquitin-protein ligase RNF168-deficient, Fanconi anemia group J protein (FACJ, BRIP1)-deficient, or DNA endonuclease RBBP8 (CtIP)-compromised cells, whereas an increased frequency of S-region inversions was observed in breast...

  5. Emergence of new forms of human immunodeficiency virus type 1 intersubtype recombinants in central Myanmar.

    Science.gov (United States)

    Motomura, K; Kusagawa, S; Kato, K; Nohtomi, K; Lwin, H H; Tun, K M; Thwe, M; Oo, K Y; Lwin, S; Kyaw, O; Zaw, M; Nagai, Y; Takebe, Y

    2000-11-20

    We have previously shown that HIV-1 env subtypes B' (a Thai-B cluster within subtype B) and E (CRF01_AE) are distributed in Yangon, the capital city of Myanmar. However, HIV strains from the rest of country have not yet been genetically characterized. In the present study, we determined env (C2/V3) and gag (p17) subtypes of 25 specimens from central Myanmar (Mandalay). Phylogenetic analyses identified 5 subtype C (20%), in addition to 10 CRF01_AE (40%) and 4 subtype B' (16%). Interestingly, the remaining six specimens (24%) showed discordance between gag and env subtypes; three gag subtype B'/env subtype C, one gag subtype B'/env subtype E, one gag subtype C/env subtype B', and one gag subtype C/env subtype E. These discordant specimens were found frequently among injecting drug users (4 of 12, 33%) and female commercial sex workers (2 of 8, 25%) engaging in high-risk behaviors. The recombinant nature of these HIV-1 strains was verified in three specimens, indicating the presence of new forms of HIV-1 intersubtype C/B' and C/B'/E recombinants with different recombination breakpoints. The data suggest that multiple subtypes of B', C, and CRF01_AE are cocirculating in central Myanmar, leading to the evolution of new forms of intersubtype recombinants among the risk populations exhibiting one of the highest HIV infection rates in the region.

  6. Human histologic evaluation of anorganic bovine bone mineral combined with recombinant human platelet-derived growth factor BB in maxillary sinus augmentation: case series study.

    Science.gov (United States)

    Nevins, Myron; Garber, David; Hanratty, James J; McAllister, Bradley S; Nevins, Marc L; Salama, Maurice; Schupbach, Peter; Wallace, Steven; Bernstein, Simon M; Kim, David M

    2009-12-01

    The objective of this proof-of-principle study was to examine the potential for improved bone regenerative outcomes in maxillary sinus augmentation procedures when recombinant human platelet-derived growth factor BB (0.3 mg/mL) is combined with particulate anorganic bovine bone mineral. The surgical outcomes in all treated sites were uneventful at 6 to 8 months, with sufficient regenerated bone present to allow successful placement of maxillary posterior implants. Large areas of dense, well-formed lamellar bone were seen throughout the intact core specimens in more than half of the grafted sites. Abundant numbers of osteoblasts were noted in concert with significant osteoid in all sites, indicating ongoing osteogenesis. A number of cores demonstrated efficient replacement of the normally slowly resorbing anorganic bovine bone mineral matrix particles with newly formed bone when the matrix was saturated with recombinant human platelet-derived growth factor BB.

  7. Cytological studies of human meiosis: sex-specific differences in recombination originate at, or prior to, establishment of double-strand breaks.

    Directory of Open Access Journals (Sweden)

    Jennifer R Gruhn

    Full Text Available Meiotic recombination is sexually dimorphic in most mammalian species, including humans, but the basis for the male:female differences remains unclear. In the present study, we used cytological methodology to directly compare recombination levels between human males and females, and to examine possible sex-specific differences in upstream events of double-strand break (DSB formation and synaptic initiation. Specifically, we utilized the DNA mismatch repair protein MLH1 as a marker of recombination events, the RecA homologue RAD51 as a surrogate for DSBs, and the synaptonemal complex proteins SYCP3 and/or SYCP1 to examine synapsis between homologs. Consistent with linkage studies, genome-wide recombination levels were higher in females than in males, and the placement of exchanges varied between the sexes. Subsequent analyses of DSBs and synaptic initiation sites indicated similar male:female differences, providing strong evidence that sex-specific differences in recombination rates are established at or before the formation of meiotic DSBs. We then asked whether these differences might be linked to variation in the organization of the meiotic axis and/or axis-associated DNA and, indeed, we observed striking male:female differences in synaptonemal complex (SC length and DNA loop size. Taken together, our observations suggest that sex specific differences in recombination in humans may derive from chromatin differences established prior to the onset of the recombination pathway.

  8. Evolutionary and network analysis of virus sequences from infants infected with an Australian recombinant strain of human parechovirus type 3.

    Science.gov (United States)

    Alexandersen, Soren; Nelson, Tiffanie M; Hodge, Jason; Druce, Julian

    2017-06-20

    We present the near complete virus genome sequences with phylogenetic and network analyses of potential transmission networks of a total of 18 Australian cases of human parechovirus type 3 (HPeV3) infection in infants in the period from 2012-2015. Overall the results support our previous finding that the Australian outbreak strain/lineage is a result of a major recombination event that took place between March 2012 and November 2013 followed by further virus evolution and possibly recombination. While the nonstructural coding region of unknown provenance appears to evolve significantly both at the nucleotide and amino acid level, the capsid encoding region derived from the Yamagata 2011 lineage of HPeV3 appears to be very stable, particularly at the amino acid level. The phylogenetic and network analyses performed support a temporal evolution from the first Australian recombinant virus sequence from November 2013 to March/April 2014, onto the 2015 outbreak. The 2015 outbreak samples fall into two separate clusters with a possible common ancestor between March/April 2014 and September 2015, with each cluster further evolving in the period from September to November/December 2015.

  9. HUMAN PAPILLOMA VIRUS IMMUNOGEN CREATION ON THE BASE OF CHIMERIC RECOMBINANT PROTEIN L2E7

    Directory of Open Access Journals (Sweden)

    I. S. Malakhov

    2016-01-01

    Full Text Available The cervical cancer is one of the most common diseases in world. This malignancy is the seventh highest prevalence oncological disease worldwide and the second highest prevalence oncological disease of women in the world. Meanwhile women need to be infected by human papilloma virus (HPV is absolutely necessary for it further evolution, HPV DNA was found in 99.97% cases of disease. Except cervical cancer, HPV cause 85% of rectal cancer, 50% of the vulva, vagina and penis cancers, 20% of oropharyngeal cancer and 10% of larynx and esophagus cancers. In 2009, 14 000 women were diagnosed with cervical cancer in Russia. The growth in morbidity was 19% (in comparison with 1999. The most effective recognised measure for almost each infection prophylaxis is a vaccination. Two human papilloma virus vaccines are available in Russia nowadays — Gardasil and Cervarix, produced in Belgium and the Netherlands respectively. Cervarix is a bivalent vaccine based on virus-like particles (VLP of two types. Recombinant major capsid proteins L1 HPV 16 and HPV 18 express in baculovirus expression system and self-assembled into virus-like particles (about 70 percent of cervical cancers are caused by HPV 16 and HPV 18. VLP of each strain produced in different baculovirus vectors and then combined in single drug. Gardasil is like Cervarix with few exceptions. Producing organisms are fungi S. cerevisiae in this case, and this vaccine contains low-risk HPV 6 and HPV 11 VLP. Thus, Gardasil is quadrivalent HPV-6/11/16/18 vaccine. These vaccines are very effective in averting infection of disease and don’t have significant side-effects, however they have some disadvantages. Firstly, they have a high price because of necessity of their expression in eukaryotic cells. Secondly, they are strain-specific, so vaccines are completely effective only for virus’s strains which are represented in the vaccine. Thirdly, it`s the absence of therapeutic (treatment of established

  10. Guideline for radioiodine therapy for benign thyroid diseases (version 4)

    International Nuclear Information System (INIS)

    Dietlein, M.; Schicha, H.; Moser, E.; Reiners, C.; Schneider, P.; Schober, O.

    2007-01-01

    Version 4 of the guideline for radioiodine therapy for benign thyroid diseases includes an interdisciplinary consensus ondecision making for antithyroid drugs, surgical treatment and radioiodine therapy. The quantitative description of a specific goiter volume for radioiodine therapy or operation was cancelled. For patients with nodular goiter with or without autonomy, manifold circumstances are in favor of surgery (suspicion on malignancy, large cystic nodules, mediastinal goiter, severe compression of the trachea) or in favor of radioiodine therapy (treatment of autonomy, age of patient, co-morbidity, history of prior subtotal thyroidectomy, profession like teacher, speaker or singer). For patients with Graves' disease, radioiodine therapy or surgery are recommended in the constellation of high risk of relapse (first-line therapy), persistence of hyperthyroidism or relapse of hyperthyroidism. After counseling, the patient gives informed consent to the preferred therapy. The period after radioiodine therapy of benign disorders until conception of at least four months was adapted to the European recommendation. (orig.)

  11. The complete genome sequence of human adenovirus 84, a highly recombinant new Human mastadenovirus D type with a unique fiber gene.

    Science.gov (United States)

    Kaján, Győző L; Kajon, Adriana E; Pinto, Alexis Castillo; Bartha, Dániel; Arnberg, Niklas

    2017-10-15

    A novel human adenovirus was isolated from a pediatric case of acute respiratory disease in Panama City, Panama in 2011. The clinical isolate was initially identified as an intertypic recombinant based on hexon and fiber gene sequencing. Based on the analysis of its complete genome sequence, the novel complex recombinant Human mastadenovirus D (HAdV-D) strain was classified into a new HAdV type: HAdV-84, and it was designated Adenovirus D human/PAN/P309886/2011/84[P43H17F84]. HAdV-D types possess usually an ocular or gastrointestinal tropism, and respiratory association is scarcely reported. The virus has a novel fiber type, most closely related to, but still clearly distant from that of HAdV-36. The predicted fiber is hypothesised to bind sialic acid with lower affinity compared to HAdV-37. Bioinformatic analysis of the complete genomic sequence of HAdV-84 revealed multiple homologous recombination events and provided deeper insight into HAdV evolution. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Procedure guidelines for radioiodine therapy of differentiated thyroid cancer (version 3); Verfahrensanweisung zur Radioiodtherapie (RIT) beim differenzierten Schilddruesenkarzinom (Version 3)

    Energy Technology Data Exchange (ETDEWEB)

    Dietlein, M.; Schicha, H. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Koeln Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin; Dressler, J. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Nuklearmedizinsiche Klinik der Henriettenstiftung, Hannover (Germany); Eschner, W. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Deutsche Gesellschaft fuer Medizinische Physik (DGMP) (Germany); Koeln Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin; Gruenwald, F. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Frankfurt Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin; Lassmann, M. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Deutsche Gesellschaft fuer Medizinische Physik (DGMP) (Germany); Wuerzburg Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin; Leisner, B. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Allgemeines Krankenhaus St. Georg, Hamburg (Germany); Luster, M.; Reiners, C. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Wuerzburg Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin; Moser, E. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Radiologische Universitaetsklinik Freiburg (Germany); Schober, O. [Deutsche Gesellschaft fuer Nuklearmedizin (DGN) (Germany); Muenster Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin

    2007-07-01

    The procedure guideline for radioiodine therapy (RIT) of differentiated thyroid cancer (version 3) is the counterpart to the procedure guideline for {sup 131}I whole-body scintigraphy (version 3) and specify the interdisciplinary guideline for thyroid cancer of the Deutsche Krebsgesellschaft concerning the nuclear medicine part. Recommendation for ablative {sup 131}I therapy is given for all differentiated thyroid carcinoma (DTC) >1 cm. Regarding DTC {<=}1 cm {sup 131}I ablation may be helpful in an individual constellation. Preparation for {sup 131}I ablation requires low iodine diet for two weeks and TSH stimulation by withdrawal of thyroid hormone medication or by use of recombinant human TSH (rhTSH). The advantages of rhTSH (no symptoms of hypothyroidism, lowerblood activity) and the advantages of endogenous TSH stimulation (necessary for {sup 131}I-therapy in patients with metastases, higher sensitivity of {sup 131}I whole-body scan) are discussed. In most centers standard activities are used for {sup 131}I ablation. If pretherapeutic dosimetry is planned, the diagnostic administration of {sup 131}I should not exceed 1-10MBq, alternative tracers are {sup 123}I or {sup 124}I. The recommendations for contraception and family planning are harmonized with the recommendation of ATA and ETA. Regarding the best possible protection of salivary glands the evidence is insufficient to recommend a specific setting. To minimize the risk of dental caries due to xerostomia patients should use preventive strategies for dental hygiene. (orig.)

  13. In vitro radiosensitivity of human fresh T-lymphocytes by colony formation assay using PHA and recombinant Interleukin-2

    International Nuclear Information System (INIS)

    Tatsugawa, Kaori; Nakamura, Nori; Hakoda, Masayuki; Akiyama, Mitoshi.

    1988-05-01

    In vitro culture conditions for colony formation of human fresh peripheral T-cells using phytohemagglutinin (PHA) and recombinant Interleukin-2 are defined. Peripheral lymphocytes, from six individuals, were exposed to X or gamma rays in vitro, and dose-survival curves were obtained. The results showed typical sigmoid curves similar to those observed when other mammalian cells are exposed to radiation. The D 10 (dose required to kill 90 % of the cells) was found to be 3.0 to 3.5 Gy. (author)

  14. Recombinant human interleukin 2 directly provides signals for the proliferation and functional maturation of murine B lymphocytes

    OpenAIRE

    Moll, Heidrun; Emmrich, F.; Simon, Markus M.

    2009-01-01

    In this study the effect of recombinant human interleukin 2 (rec.hIL-2) on the proliferation and maturation of B lymphocytes was investigated. It was found that the presence of rec.hIL 2 results in proliferation of mitogen (LPS)-activated B cell blasts. In addition, it is shown that highly enriched murine B cells can be induced by rec.hIL-2 to proliferate and to develop into antibody-secreting cells (PFC) in the presence of antigen (SRBC). When tested for its effect on B cell preparations enr...

  15. Small-angle neutron scattering study of recombinant yeast-derived human hepatitis B virus surface antigen vaccine particle

    Science.gov (United States)

    Sato, M.; Ito, Y.; Kameyama, K.; Imai, M.; Ishikawa, N.; Takagi, T.

    1995-02-01

    The overall and internal structure of recombinant yeast-derived human hepatitis B virus surface antigen vaccine particles was investigated by small-angle neutron scattering using the contrast variation method. The vaccine is a nearly spherical particle, and its contrast-matching point was determined to be at about 24% D 2O content, indicating that a large part of the vaccine particle is occupied by lipids and carbohydrates from the yeast. The Stuhrmann plot suggests that the surface antigens exist predominantly in the peripheral region of the particle, which is favorable to the induction of anti-virus antibodies.

  16. Recombinant human melatonin receptor MT1 isolated in mixed detergents shows pharmacology similar to that in mammalian cell membranes.

    Directory of Open Access Journals (Sweden)

    Christel Logez

    Full Text Available The human melatonin MT1 receptor-belonging to the large family of G protein-coupled receptors (GPCRs-plays a key role in circadian rhythm regulation and is notably involved in sleep disorders and depression. Structural and functional information at the molecular level are highly desired for fine characterization of this receptor; however, adequate techniques for isolating soluble MT1 material suitable for biochemical and biophysical studies remain lacking. Here we describe the evaluation of a panel of constructs and host systems for the production of recombinant human MT1 receptors, and the screening of different conditions for their solubilization and purification. Our findings resulted in the establishment of an original strategy using a mixture of Fos14 and CHAPS detergents to extract and purify a recombinant human MT1 from Pichia pastoris membranes. This procedure enabled the recovery of relatively pure, monomeric and ligand-binding active MT1 receptor in the near-milligram range. A comparative study based on extensive ligand-binding characterization highlighted a very close correlation between the pharmacological profiles of MT1 purified from yeast and the same receptor present in mammalian cell membranes. The high quality of the purified MT1 was further confirmed by its ability to activate its cognate Gαi protein partner when reconstituted in lipid discs, thus opening novel paths to investigate this receptor by biochemical and biophysical approaches.

  17. Recombinant human melatonin receptor MT1 isolated in mixed detergents shows pharmacology similar to that in mammalian cell membranes.

    Science.gov (United States)

    Logez, Christel; Berger, Sylvie; Legros, Céline; Banères, Jean-Louis; Cohen, William; Delagrange, Philippe; Nosjean, Olivier; Boutin, Jean A; Ferry, Gilles; Simonin, Frédéric; Wagner, Renaud

    2014-01-01

    The human melatonin MT1 receptor-belonging to the large family of G protein-coupled receptors (GPCRs)-plays a key role in circadian rhythm regulation and is notably involved in sleep disorders and depression. Structural and functional information at the molecular level are highly desired for fine characterization of this receptor; however, adequate techniques for isolating soluble MT1 material suitable for biochemical and biophysical studies remain lacking. Here we describe the evaluation of a panel of constructs and host systems for the production of recombinant human MT1 receptors, and the screening of different conditions for their solubilization and purification. Our findings resulted in the establishment of an original strategy using a mixture of Fos14 and CHAPS detergents to extract and purify a recombinant human MT1 from Pichia pastoris membranes. This procedure enabled the recovery of relatively pure, monomeric and ligand-binding active MT1 receptor in the near-milligram range. A comparative study based on extensive ligand-binding characterization highlighted a very close correlation between the pharmacological profiles of MT1 purified from yeast and the same receptor present in mammalian cell membranes. The high quality of the purified MT1 was further confirmed by its ability to activate its cognate Gαi protein partner when reconstituted in lipid discs, thus opening novel paths to investigate this receptor by biochemical and biophysical approaches.

  18. Engineering the oxygen sensing regulation results in an enhanced recombinant human hemoglobin production by Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Martinez Ruiz, José Luis; Liu, Lifang; Petranovic, Dina

    2015-01-01

    Efficient production of appropriate oxygen carriers for transfusions (blood substitutes or artificial blood) has been pursued for many decades, and to date several strategies have been used, from synthetic polymers to cell-free hemoglobin carriers. The recent advances in the field of metabolic en...... the transcription factor HAP1, which resulted in an increase of the final recombinant active hemoglobin titer exceeding 7% of the total cellular protein....

  19. Recombinant human protein - new palette of drugs for the treatment of psoriasis

    OpenAIRE

    Kristl, Jernej

    2015-01-01

    Psoriasis is a chronic, inflammatory disease, which is estimated to affect 2,5% of the worldćs population. Clinically it is characterized by thickened, silvery white, scaly plaques, that can produce considerable discomfort and pain in patients. For the treatment of psoriasis are currently mainly used corticosteroids, vitamin D analogs, ultraviolet A radiation and retinoids, allof which are nonspecific and unable to block the outbreak of disease. Advances in recombinant DNA technology and incr...

  20. Current status of radioiodine (131I) treatment of hyperthyroidism

    International Nuclear Information System (INIS)

    Becker, D.V.; Hurley, J.R.

    1982-01-01

    Radioiodine appears to be increasingly selected as the primary form of therapy for most hyperthyroid adults. Some controversy exists concerning the use of I-131 in children and adolescents as well as in some patients with toxic nodular goiter. The technical aspects of radioiodine administration reviewed in this chapter include the biologic basis of I-131 use, dose strategies, measurement and evaluation of radioiodine parameters, and I-125 treatment of hyperthyroidism. The management of patients before and after I-131 treatment is discussed as well as posttreatment hypothyroidism and complications of radioiodine treatment

  1. Thyrotoxicosis and radioiodine therapy: Does the dose matter?

    Directory of Open Access Journals (Sweden)

    Andrew Collier

    2012-01-01

    Full Text Available There are 3 treatment options for thyrotoxicosis: Antithyroid drugs, Surgery and radioiodine. The choice of treatment varies geographically. Radioiodine therapy is preferred in the United States. The aim of radioiodine is to destroy sufficient thyroid tissue to cure the hyperthyroidism. There is a lack of consensus towards what dose of radioiodine should be used. Several methods are used to determine the dose. In our practice we administer 400 MBq to patients with Graves and in patients with large multinodular goiter, we would administer 800 MBq.

  2. METHODS FOR RECOMBINANT EXPRESSION AND FUNCTIONAL CHARACTERIZATION OF HUMAN CANNABINOID RECEPTOR CB2

    Directory of Open Access Journals (Sweden)

    Alexei A. Yeliseev

    2013-03-01

    Full Text Available Cannabinoid receptor CB2 is a seven transmembrane-domain integral membrane protein that belongs to a large superfamily of G protein-coupled receptors (GPCR. CB2 is a part of the endocannabinoid system that plays vital role in regulation of immune response, inflammation, pain sensitivity, obesity and other physiological responses. Information about the structure and mechanisms of functioning of this receptor in cell membranes is essential for the rational development of specific pharmaceuticals. Here we review the methodology for recombinant expression, purification, stabilization and biochemical characterization of CB2 suitable for preparation of multi-milligram quantities of functionally active receptor. The biotechnological protocols include expression of the recombinant CB2 in E. coli cells as a fusion with the maltose binding protein, stabilization with a high affinity ligand and a derivative of cholesterol in detergent micelles, efficient purification by tandem affinity chromatography, and reconstitution of the receptor into lipid bilayers. The purified recombinant CB2 receptor is amenable to functional and structural studies including nuclear magnetic resonance spectroscopy and a wide range of biochemical and biophysical techniques.

  3. Thirty six treatments with radiodine after the administration of TSH recombinant in 26 patients with carcinoma differentiated from thyroid gland

    International Nuclear Information System (INIS)

    Pitoia, F.; Passerieu, M.; Bruno, O.D.; Niepomniszcze, H.

    2004-01-01

    The released studies that confirm the safety and efficacy the TSH recombinant (rhTSH) led to an increase in the interest for the use in the patients' preparation with thyroid remanents normal or differenced thyroid carcinoma persistent/recurrent, before the administration of therapeutic radioiodine doses in some situations, when it is impossible the suspension of the hormonal therapy thyroid. The objective is to evaluate the effectiveness of the administration of therapeutic doses of radioiodine after the administration of rhTSH

  4. Direct application of radioiodinated aminoacyl tRNA for radiolabeling nascent proteins

    International Nuclear Information System (INIS)

    Scherberg, N.H.; Barokas, K.; Murata, Y.; Refetoff, S.

    1985-01-01

    A two-step procedure to incorporate 125 I-iodotyrosine into protein synthesized in a reticulocyte lysate is described. In the first step, the iodination of tyrosyl tRNA was catalyzed by a solid-state glycouril compound. More than one-third of 200 microCi of radioiodine became bound to 70 micrograms of aminoacyl tRNA after 15 min at 0 degrees C. The isotope was distributed in a three-to-one ratio of monoiodotyrosine to di-iodotyrosine. In the second step, the soluble product of the radioiodination was transferred directly into a nuclease-treated reticulocyte lysate coded with RNA isolated from the human hepatoma cell line Hep G2. Fractional recovery of radioiodine in nascent protein was maximally 7.6%. Reaction of the product of translation with antibody against alpha-antitrypsin separated an 125 I-containing protein having a molecular weight estimated as 47,000. The synthesis of unprocessed alpha-antitrypsin was confirmed by cleavage of the labeled protein with leader peptidase and by its displacement from immunocomplex formation with purified alpha-antitrypsin. The amount of 125 I incorporated into alpha-antitrypsin was proportionate to iodinated tRNA additions up to a concentration of 70 micrograms/ml. The synthesis of alpha-antitrypsin as detected in radioautograms after gel electrophoresis was more than twice as sensitive using radioiodinated aminoacyl tRNA as compared with [ 35 S]methionine. Iodine labeling of thyroxine-binding globulin was also demonstrated in the translation product of Hep G2 RNA. Since the specific activity of the radioiodine is high and the means for detection of the isotope efficient, the method described can facilitate the demonstration of quantitatively minor translation products

  5. Graves' disease and toxic nodular goiter - radioiodine therapy; Morbus Basedow und Autonomie - Radioiodtherapie

    Energy Technology Data Exchange (ETDEWEB)

    Schicha, H.; Dietlein, M. [Koeln Univ. (Germany). Klinik und Poliklinik fuer Nuklearmedizin

    2002-04-01

    At the 15th conference on the human thyroid in Heidelberg in 2001 the following aspects of the radioiodine therapy of benign thyroid disorders were presented: General strategies for therapy of benign thyroid diseases, criterions for conservative or definitive treatment of hyperthyroidism as first line therapy and finally preparation, procedural details, results, side effects, costs and follow-up care of radioiodine therapy as well as legal guidelines for hospitalization in Germany. The diagnosis Graves' hyperthyroidism needs the decision, if rather a conservative treatment or if primary radioiodine therapy is the best therapeutic approach. In the USA 70-90% of these patients are treated with radioiodine as first line therapy, whereas in Germany the conservative therapy for 1-1.5 years is recommended for 90%. This review describes subgroups of patients with Graves' disease showing a higher probability to relapse after conservative treatment. Comparing benefits, adverse effects, costs, and conveniences of both treatment strategies the authors conclude that radioiodine therapy should be preferred as first line therapy in 60-70% of the patients with Graves' hyperthyroidism. (orig.) [German] In dem vorliegenden Beitrag wird ueber die Radioiodtherapie gutartiger Schilddruesenerkrankungen referiert. Dies betrifft gesetzliche Regelungen in Deutschland, grundlegende Ueberlegungen zur Therapie benigner Schilddruesenerkrankungen, die Diskussion konservative versus definitive Therapieindikation der Hyperthyreose und schliesslich die Themenkomplexe Vorbereitung, Durchfuehrung, Ergebnisse, Nebenwirkungen und Komplikationen, Kosten sowie Nachsorge der Radioiodtherapie. Im Vordergrund steht die Abgrenzung der konservativen Therapieindikation gegenueber der primaeren Radioiodtherapie des Morbus Basedow in Deutschland. Waehrend in den USA 70-90% der Morbus-Basedow-Faelle primaer mit Radioiodtherapie behandelt werden, wird in Deutschland bei 90% zunaechst ein

  6. Investigation of {sup 99m}Tc-labelling of recombinant human interleukin-2 via hydrazinonicotinamide

    Energy Technology Data Exchange (ETDEWEB)

    Karczmarczyk, Urszula, E-mail: ukarczmarczyk@o2.p [Department of Radiopharmaceuticals, National Medicines Institute, 00-725 Warsaw (Poland); Garnuszek, Piotr; Maurin, Michal [Department of Radiopharmaceuticals, National Medicines Institute, 00-725 Warsaw (Poland); Di Gialleonardo, Valentina [Nuclear Medicine, Ospedale S. Andrea, Via di Grottarossa 1035, 00189 Rome (Italy); Department of Nuclear Medicine and Molecular Imaging, University of Groningen, 9700 RB Groningen (Netherlands); Galli, Filippo [Nuclear Medicine, Ospedale S. Andrea, Via di Grottarossa 1035, 00189 Rome (Italy); Signore, Alberto [Nuclear Medicine, Ospedale S. Andrea, Via di Grottarossa 1035, 00189 Rome (Italy); Department of Nuclear Medicine and Molecular Imaging, University of Groningen, 9700 RB Groningen (Netherlands); Mikolajczak, Renata [Institute of Atomic Energy, Radioisotope Centre POLATOM, 05-400 Swierk (Poland)

    2010-10-15

    Introduction: Interleukin-2 (IL-2) when radiolabelled with {sup 99m}Tc has been proved useful in imaging the side of lymphocytic infiltration in patients with autoimmune disorders and plays a significant role as a T-cell imaging agent. However, the labelling procedures used so far appeared to be rather complex and laborious. The aim of present study was to develop an efficient procedure of {sup 99m}Tc-labelling of recombinant human interleukin-2 (rhIL-2) via hydrazinonicotinamide (HYNIC) to develop a dry kit formulation. Methods: Various molar ratios of rhIL-2/HYNIC (from 1:2 to 1:12) were used at the conjugation step. The conjugates were purified on a PD-10 column to remove the excess of unbound HYNIC, as well as of any aggregates. The final peptide concentration was quantified by the BCA method, and the number of HYNIC molecules incorporated into a rhIL-2 molecule was determined based on the reaction with 2-sulfobenzaldehyde. The {sup 99m}Tc-labelling was optimized using various amounts of HYNIC-rhIL-2, {sup 99m}Tc, SnCl{sub 2}, tricine and nicotinic acid (NA). Quality control included GF-HPLC, ITLC, SDS-PAGE and biological assay. Biodistribution studies were performed in Swiss mice and Wistar rats. Results: Generally, the highest radiolabelling yields were achieved when the HYNIC-rhIL-2 conjugates of ca. 2-4 HYNIC molecule substitution ratios were used. The optimal pH of the reaction medium was found to be in the range of 6.5 to 7.0. GF-HPLC analysis indicated that monomer and aggregates of {sup 99m}Tc-HYNIC-rhIL-2 are formed during radiolabelling. At optimized conditions of wet radiolabelling, the {sup 99m}Tc-HYNIC-rhIL-2 monomer was obtained with radiochemical purity >99%, specific activity of ca. 4 GBq/mg rhIL-2 and overall yield of ca. 65%. The two-vial freeze-dried kit was prepared: the first vial contained 30 {mu}g HYNIC-rhIL-2, co-ligands, buffer and antioxidant; the second vial contained tricine and SnCl{sub 2}. The monomer of {sup 99m}Tc-HYNIC-rhIL-2

  7. Haemoglobin mass and running time trial performance after recombinant human erythropoietin administration in trained men.

    Directory of Open Access Journals (Sweden)

    Jérôme Durussel

    Full Text Available UNLABELLED: Recombinant human erythropoietin (rHuEpo increases haemoglobin mass (Hb(mass and maximal oxygen uptake (v O(2 max. PURPOSE: This study defined the time course of changes in Hb(mass, v O(2 max as well as running time trial performance following 4 weeks of rHuEpo administration to determine whether the laboratory observations would translate into actual improvements in running performance in the field. METHODS: 19 trained men received rHuEpo injections of 50 IU•kg(-1 body mass every two days for 4 weeks. Hb(mass was determined weekly using the optimized carbon monoxide rebreathing method until 4 weeks after administration. v O(2 max and 3,000 m time trial performance were measured pre, post administration and at the end of the study. RESULTS: Relative to baseline, running performance significantly improved by ∼6% after administration (10:30±1:07 min:sec vs. 11:08±1:15 min:sec, p<0.001 and remained significantly enhanced by ∼3% 4 weeks after administration (10:46±1:13 min:sec, p<0.001, while v O(2 max was also significantly increased post administration (60.7±5.8 mL•min(-1•kg(-1 vs. 56.0±6.2 mL•min(-1•kg(-1, p<0.001 and remained significantly increased 4 weeks after rHuEpo (58.0±5.6 mL•min(-1•kg(-1, p = 0.021. Hb(mass was significantly increased at the end of administration compared to baseline (15.2±1.5 g•kg(-1 vs. 12.7±1.2 g•kg(-1, p<0.001. The rate of decrease in Hb(mass toward baseline values post rHuEpo was similar to that of the increase during administration (-0.53 g•kg(-1•wk(-1, 95% confidence interval (CI (-0.68, -0.38 vs. 0.54 g•kg(-1•wk(-1, CI (0.46, 0.63 but Hb(mass was still significantly elevated 4 weeks after administration compared to baseline (13.7±1.1 g•kg(-1, p<0.001. CONCLUSION: Running performance was improved following 4 weeks of rHuEpo and remained elevated 4 weeks after administration compared to baseline. These field performance effects coincided with r

  8. Characterization of recombinant human HBP/CAP37/azurocidin, a pleiotropic mediator of inflammation-enhancing LPS-induced cytokine release from monocytes.

    Science.gov (United States)

    Rasmussen, P B; Bjørn, S; Hastrup, S; Nielsen, P F; Norris, K; Thim, L; Wiberg, F C; Flodgaard, H

    1996-07-15

    Neutrophil-derived heparin-binding protein (HBP) is a strong chemoattractant for monocytes. We report here for the first time the expression of recombinant HBP. A baculovirus containing the human HBP cDNA mediated in insect cells the secretion of a 7-residue N-terminally extended HBP form (pro-HBP). Deletion of the pro-peptide-encoding cDNA sequence resulted in correctly processed HBP at the N-terminus. Electrospray mass spectrum analysis of recombinant HBP yielded a molecular weight of 27.237 +/- 3 amu. Consistent with this mass is a HBP form of 225 amino acids (mature part +3 amino acid C-terminal extension). The biological activity of recombinant HBP was confirmed by its chemotactic action towards monocytes. Furthermore, we have shown that recombinant HBP stimulates in a dose-dependent manner the lipopolysaccharide (LPS)-induced cytokine release from human monocytes.

  9. Radioiodination of proteins by reductive alkylation

    International Nuclear Information System (INIS)

    Panuska, J.R.; Parker, C.W.

    1987-01-01

    The use of the aliphatic aldehyde, para-hydroxyphenylacetaldehyde as the reactive moiety in the radioiodination of proteins by reductive alkylation is described. The para-hydroxyphenyl group is radiolabeled with 125 I, reacted through its aliphatic aldehyde group with primary amino groups on proteins to form a reversible Schiff base linkage which can then be stabilized with the mild reducing agent NaCNBH 3 . The introduction of the methylene group between the benzene ring and the aldehyde group increases its reactivity with protein amino groups permitting efficient labeling at low aldehyde concentrations. Using this method, radioiodinated proteins with high specific activity can be produced. The reductive alkylation procedure is advantageous in that the labeling conditions are mild, the reaction is specific for lysyl residues, and the modification of the epsilon-ammonium group of lysine results in ionizable secondary amino groups avoiding major changes in protein charge

  10. Results of radioiodine therapy of hyperthyroidism

    International Nuclear Information System (INIS)

    Gabriel, A.

    1980-01-01

    Between 1971 and 1977, 102 hyperthyroidism patients were treated with 131 I. The average age of the patients was 64 years. In 78 patients, hyperthyroidism was due to decompensated autonomous adenoma, 24 patients presented with diffuse hyperthyroidism. The patients were observed for 14 to 24 months. Until early 1976, diagnosis and therapy success assessment were based on thyroid scintiscanning, radioiodine two-phase tests, PBI 131 , T 4 tests, and T 3 tests. Since 1976, the radioiodine two-phase test has been replaced by the TRH test, the TRH-RIA, and the T 4 -RIA. The patients were given 3 to 25 mCi 131 I. In 88 cases, euthyroidism was reached after a single therapy; in 8 cases, after two therapies; in 1 case, after three therapies; in 4 cases, after 4 therapies; in 1 case, after 5 therapies. Euthyroidism was thus reached in 96% of all cases after 131 I therapy of hyperthyroidism. (orig./MG) [de

  11. Transmission of radioiodine through sampling lines

    International Nuclear Information System (INIS)

    Unrein, P.J.; Pelletier, C.A.; Cline, J.E.; Voilleque, P.G.

    1985-01-01

    An experimental program to measure radioiodine transmission through sampling lines is described. The transmission depends upon both deposition on and resuspension from the walls of the line. The deposition and resuspension processes are themselves controlled by the length, diameter, and material of the line and the conditions under which it is operated. The operating conditions under study are sampling flow rate, temperature and relative humidity. Measurement results have been interpreted in terms of a four-compartment model of radioiodine deposition and resuspension. The model is applied to each of twenty or more segments of the line. Experimental measurements of short-term transmission fractions and the deposition velocities derived from them are presented for six lines. Data on resuspension rates for the same lines were obtained and are also discussed

  12. Radioiodination via isotope exchange in pivalic acid

    International Nuclear Information System (INIS)

    Weichert, J.P.; Van Dort, M.E.; Groziak, M.P.; Counsell, R.E.

    1986-01-01

    A variety of benzoic and aryl aliphatic mono and polyiodinated acids and esters (sterol, triglyceride) were radioiodinated in 55-99% radiochemical yield by isotope exchange with Na 125 i in a melt of pivalic acid. In general, the reaction was complete in 1 h at 155 0 C with little or no substrate decomposition. High specific activity studies afforded 125 I-labeled iopanoic acid with a specific activity of over 700 Ci/mmol. (author)

  13. Expression and characterization of recombinant human factor V and a mutant lacking a major portion of the connecting region

    International Nuclear Information System (INIS)

    Kane, W.H.; Devore-Carter, D.; Ortel, T.L.

    1990-01-01

    Human coagulation factor V is a protein cofactor that is an essential component of the prothrombinase complex. A full-length factor V cDNA has been subcloned into the mammalian expression vector pDX and used to transfect COS cells. Approximately 95 ± 4% of the recombinant human factor V (rHFV) synthesized in COS cells is secreted into the culture medium. Factor V activity determined by fibrometer assay increased approximately 5-fold from 0.027 ± 0.012 to 0.124 ± 0.044 unit/mL following activation by the factor V activating enzyme from Russell's viper venom (RVV-V). A chromogenic assay specific for factor Va indicated that recombinant factor V had 3.8 ± 1.3% of the activity of the activated protein. The estimated specific activity of the recombinant factor Va was approximately 1,800 ± 500 units/mg, which is similar to the specific activity of purified plasma factor Va of 1,700-2,000 units/mg. Immunoprecipitation of [ 35 S]methionine-labeled rHFV revealed a single high molecular mass component. Treatment of rHFV with thrombin or RVV-V resulted in the formation of proteolytic products that were similar to those seen with plasma factor V. The authors have also expressed a mutant, rHFV-des-B 811-1441 , that lacks a large portion of the highly glycosylated connecting region that is present in factor V. This mutant constitutively expressed 38 ± 7% of the activity of the RVV-V-activated protein. These results suggest that one of the functions of the large connecting region in factor V is to inhibit constitutive procoagulant activity

  14. Bioprocess development for extracellular production of recombinant human interleukin-3 (hIL-3) in Pichia pastoris.

    Science.gov (United States)

    Dagar, Vikas Kumar; Adivitiya; Devi, Nirmala; Khasa, Yogender Pal

    2016-10-01

    Human interleukin-3 (hIL-3) is a therapeutically important cytokine involved in the maturation and differentiation of various cells of the immune system. The codon-optimized hIL-3 gene was cloned in fusion with the N-terminus α-mating factor signal peptide of Saccharomyces cerevisiae under an inducible alcohol oxidase 1 (AOX1) and constitutive glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter. A Zeocin concentration up to 2000 mg/L was used to select hyper-producers. The shake flask cultivation studies in the Pichia pastoris GS115 host resulted a maximum recombinant hIL-3 expression level of 145 mg/L in the extracellular medium under the control of AOX1 promoter. The batch fermentation strategy allowed us to attain a fairly pure glycosylated hIL-3 protein in the culture supernatant at a final concentration of 475 mg/L with a high volumetric productivity of 4.39 mg/L/h. The volumetric product concentration achieved at bioreactor level was 3.28 folds greater than the shake flask results. The 6x His-tagged protein was purified using Ni-NTA affinity chromatography and confirmed further by western blot analysis using anti-6x His tag antibody. The glycosylation of recombinant hIL-3 protein was confirmed in a PNGase F deglycosylation reaction where it showed a molecular weight band pattern similar to E. coli produced non-glycosylated hIL-3 protein. The structural properties of recombinant hIL-3 protein were confirmed by CD and fluorescence spectroscopy where protein showed 40 % α-helix, 12 % β-sheets with an emission maxima at 343 nm. MALDI-TOF-TOF analysis was used to establish the protein identity. The biological activity of purified protein was confirmed by the human erythroleukemia TF-1 cell proliferation assay.

  15. Construction of a recombinant eukaryotic human ZHX1 gene expression plasmid and the role of ZHX1 in hepatocellular carcinoma.

    Science.gov (United States)

    Wang, Jianping; Liu, Dejie; Liang, Xiaohong; Gao, Lifen; Yue, Xuetian; Yang, Yang; Ma, Chunhong; Liu, Jun

    2013-11-01

    The zinc-fingers and homeoboxes protein 1 (ZHX1) consists of 873 amino acid residues, is localized in the cell nucleus and appears to act as a transcriptional repressor. Previous studies have shown that ZHX1 interacts with nuclear factor Y subunit α (NF-YA), DNA methyltransferases (DNMT) 3B and ZHX2, all of which are involved in tumorigenesis. However, the exact role of ZHX1 in tumorigenesis remains unknown. The aim of the current study was to construct a recombinant eukaryotic expression plasmid containing the human ZHX1 (hZHX1) gene and to investigate the biological activities of ZHX1 in hepatocellular carcinoma (HCC). Reverse transcription-polymerase chain reaction (RT‑PCR) was used to amplify the N- and C-terminal fragments (ZHX1‑N and ZHX1‑C, respectively) of the hZHX1 gene. The two PCR fragments were cloned into the pEASY-T1 vector and subcloned into the pcDNA3 plasmid to generate a recombinant pcDNA3‑ZHX1 plasmid. Following identification by enzyme digestion and DNA sequencing, the recombinant pcDNA3‑ZHX1 plasmid was transfected into SMMC-7721 cells. The level of ZHX1 expression was detected by RT-PCR and western blot analysis. Cell growth curve assays were used to evaluate the effect of ZHX1 on cell proliferation. Moreover, the differential expression of ZHX1 between cancer and adjacent cirrhotic liver tissue was investigated by quantitative PCR (qPCR). Enzyme digestion and DNA sequencing confirmed the successful construction of the recombinant plasmid, pcDNA3‑ZHX1. qPCR and western blot analysis demonstrated that ZHX1 was efficiently expressed in SMMC-7721 cells and overexpression of ZHX1 may inhibit the proliferation of SMMC-7721 cells. In addition, reduced ZHX1 expression is widespread among cancer tissues from HCC patients. In conclusion, a recombinant eukaryotic expression plasmid, pcDNA3‑ZHX1, was successfully constructed. In addition, the current results indicate that a low expression of ZHX1 may be responsible for hepatocarcinogenesis.

  16. Prosthetic radioiodination of interleukin-8 ([{sup 123/131}I]-IL-8): biological behavior in a mouse infection model

    Energy Technology Data Exchange (ETDEWEB)

    Amartey, J.K. [Cyclotron and Radiopharmaceuticals Department, King Faisal Specialist Hospital and Research Centre, MBC-03, P.O. Box 3354, Riyadh 11211 (Saudi Arabia)]. E-mail: amarty@kfshrc.edu.sa; Esguerra, C. [Cyclotron and Radiopharmaceuticals Department, King Faisal Specialist Hospital and Research Centre, MBC-03, P.O. Box 3354, Riyadh 11211 (Saudi Arabia); Al-Otaibi, B. [Cyclotron and Radiopharmaceuticals Department, King Faisal Specialist Hospital and Research Centre, MBC-03, P.O. Box 3354, Riyadh 11211 (Saudi Arabia); Al-Jammaz, I. [Cyclotron and Radiopharmaceuticals Department, King Faisal Specialist Hospital and Research Centre, MBC-03, P.O. Box 3354, Riyadh 11211 (Saudi Arabia); Al-Qahtani, M. [Cyclotron and Radiopharmaceuticals Department, King Faisal Specialist Hospital and Research Centre, MBC-03, P.O. Box 3354, Riyadh 11211 (Saudi Arabia); Parhar, R.S. [Cyclotron and Radiopharmaceuticals Department, King Faisal Specialist Hospital and Research Centre, MBC-03, P.O. Box 3354, Riyadh 11211 (Saudi Arabia)

    2005-01-01

    Numerous molecular entities with diverse structures have been radiolabeled and investigated as potential infection and inflammation detection agents. However, none of these molecules have gained the acceptance of gallium citrate or radiolabeled autologous white blood cells. We have radioiodinated interleukin-8 using two different methods and tested the biological behavior of the products in mice. As expected, the direct radioiodinated material displayed extensive in vivo deiodination. The use of pyridine-based prosthetic label yielded a product with better kinetics than the direct radioiodination method and showed a better target to non-target ratio. Nonetheless, this method is not suited for labeling of bioactive peptides such as the title peptide because of the very high specific activity required to prevent cytotoxic effects in a human application.

  17. Prosthetic radioiodination of interleukin-8 ([123/131I]-IL-8): biological behavior in a mouse infection model

    International Nuclear Information System (INIS)

    Amartey, J.K.; Esguerra, C.; Al-Otaibi, B.; Al-Jammaz, I.; Al-Qahtani, M.; Parhar, R.S.

    2005-01-01

    Numerous molecular entities with diverse structures have been radiolabeled and investigated as potential infection and inflammation detection agents. However, none of these molecules have gained the acceptance of gallium citrate or radiolabeled autologous white blood cells. We have radioiodinated interleukin-8 using two different methods and tested the biological behavior of the products in mice. As expected, the direct radioiodinated material displayed extensive in vivo deiodination. The use of pyridine-based prosthetic label yielded a product with better kinetics than the direct radioiodination method and showed a better target to non-target ratio. Nonetheless, this method is not suited for labeling of bioactive peptides such as the title peptide because of the very high specific activity required to prevent cytotoxic effects in a human application

  18. Poly(norepinephrine)-coated open tubular column for the separation of proteins and recombination human erythropoietin by capillary electrochromatography.

    Science.gov (United States)

    Xiao, Xue; Zhang, Yamin; Wu, Jia; Jia, Li

    2017-12-01

    Recombinant human erythropoietin is an important therapeutic protein with high economic interest due to the benefits provided by its clinical use for the treatment of anemias associated with chronic renal failure and chemotherapy. In this work, a poly(norepinephrine)-coated open tubular column was successfully prepared based on the self-polymerization of norepinephrine under mild alkaline condition, the favorable film forming and easy adhesive properties of poly(norepinephrine). The poly(norepinephrine) coating was characterized by scanning electron microscopy and measurement of the electro-osmotic flow. The thickness of the coating was about 431 nm. The electrochromatographic performance of the poly(norepinephrine)-coated open tubular column was evaluated by separation of proteins. Some basic and acidic proteins including two variants of bovine serum albumin and two variants of β-lactoglobulin achieved separation in the poly(norepinephrine)-coated open tubular column. More importantly, the column demonstrated separation ability for the glycoforms of recombinant human erythropoietin. In addition, the column demonstrated good repeatability with the run-to-run, day-to-day, and column-to-column relative standard deviations of migration times of proteins less than 3.40%. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Growth Hormone Safety Workshop Position Paper: a critical appraisal of recombinant human growth hormone therapy in children and adults

    DEFF Research Database (Denmark)

    Allen, David B; Backeljauw, Philippe; Bidlingmaier, Martin

    2015-01-01

    Recombinant human GH (rhGH) has been in use for 30 years, and over that time its safety and efficacy in children and adults has been subject to considerable scrutiny. In 2001, a statement from the GH Research Society (GRS) concluded that 'for approved indications, GH is safe'; however, the statem...... (PES) convened a meeting to reappraise the safety of rhGH. The ouput of the meeting is a concise position statement.......Recombinant human GH (rhGH) has been in use for 30 years, and over that time its safety and efficacy in children and adults has been subject to considerable scrutiny. In 2001, a statement from the GH Research Society (GRS) concluded that 'for approved indications, GH is safe'; however......, the statement highlighted a number of areas for on-going surveillance of long-term safety, including cancer risk, impact on glucose homeostasis, and use of high dose pharmacological rhGH treatment. Over the intervening years, there have been a number of publications addressing the safety of rhGH with regard...

  20. Influence of low- and high-dose radioiodine therapy on oxidative modification of fibrinogen

    International Nuclear Information System (INIS)

    Schweeger-Exeli, I.J.

    2001-10-01

    Fibrinogen plays a central role in the course of thrombosis and hemostasis. It is soluble in blood and tissue extracts and transformed into the insoluble fibrin network structure in the presence of thrombin. Fibrinogen in circulating blood consists of a population of slightly different molecules with a half-life of 3.5-4.5 days. Various environmental conditions may cause different types of modifications of the molecule leading to a different functional behavior. Introduction of carbonyl groups in amino acid side chains is known as a marker for protein oxidation. Radioiodine therapy, applied in patients suffering from hyperthyroidism or differentiated thyroid carcinoma, may cause an oxidative modification of fibrinogen by formation of free radicals in blood exposed to the radioactive agent 131I. The topic of my thesis was i. to develop a simple and not time consuming method for isolation of fibrinogen from small volumes of human plasma (∼ 6ml), ii. to assess, whether radioiodine therapy causes detectable introduction of carbonyl groups into the fibrinogen molecule, and iii. to analyze an association between thyroid hormone function, fibrinogen levels and protein oxidation by means of carbonyl content. Purification of fibrinogen from human plasma was possible by three different methods (ammonium sulphate/ethanol; glycine/ethanol; glycine). Plasma levels of fibrinogen (Clauss method) and protein carbonyl group content (2,4-DNPH - assay) were determined before and after radioiodine therapy. The results demonstrate a significant increase (p = 0.05) in carbonyl content of human fibrinogen in cancer patients treated with 131I. However, in patients with diagnosed hyperthyroidism values were not significantly altered. In carcinoma patients, baseline fT4 levels and the relative increase in carbonyl content of fibrinogen after radioiodine therapy were correlated (r = 0.83; p 0.005), whereas no such correlation was found in patients with hyperthyroidism. Plasma fibrinogen

  1. Anti-Human Endoglin (hCD105 Immunotoxin—Containing Recombinant Single Chain Ribosome-Inactivating Protein Musarmin 1

    Directory of Open Access Journals (Sweden)

    Begoña Barriuso

    2016-06-01

    Full Text Available Endoglin (CD105 is an accessory component of the TGF-β receptor complex, which is expressed in a number of tissues and over-expressed in the endothelial cells of tumor neovasculature. Targeting endoglin with immunotoxins containing type 2 ribosome-inactivating proteins has proved an effective tool to reduce blood supply to B16 mice tumor xenografts. We prepared anti-endoglin immunotoxin (IT—containing recombinant musarmin 1 (single chain ribosome-inactivating proteins linked to the mouse anti-human CD105 44G4 mouse monoclonal antibody via N-succinimidyl 3-(2-pyridyldithio propionate (SPDP. The immunotoxin specifically killed L929 fibroblast mouse cells transfected with the short form of human endoglin with IC50 values in the range of 5 × 10−10 to 10−9 M.

  2. Cloning of human tumor necrosis factor (TNF) receptor cDNA and expression of recombinant soluble TNF-binding protein

    International Nuclear Information System (INIS)

    Gray, P.W.; Barrett, K.; Chantry, D.; Turner, M.; Feldmann, M.

    1990-01-01

    The cDNA for one of the receptors for human tumor necrosis factor (TNF) has been isolated. This cDNA encodes a protein of 455 amino acids that is divided into an extracellular domain of 171 residues and a cytoplasmic domain of 221 residues. The extracellular domain has been engineered for expression in mammalian cells, and this recombinant derivative binds TNFα with high affinity and inhibits its cytotoxic activity in vitro. The TNF receptor exhibits similarity with a family of cell surface proteins that includes the nerve growth factor receptor, the human B-cell surface antigen CD40, and the rat T-cell surface antigen OX40. The TNF receptor contains four cysteine-rich subdomains in the extracellular portion. Mammalian cells transfected with the entire TNF receptor cDNA bind radiolabeled TNFα with an affinity of 2.5 x 10 -9 M. This binding can be competitively inhibited with unlabeled TNFα or lymphotoxin (TNFβ)

  3. Anti-Human Endoglin (hCD105) Immunotoxin-Containing Recombinant Single Chain Ribosome-Inactivating Protein Musarmin 1.

    Science.gov (United States)

    Barriuso, Begoña; Antolín, Pilar; Arias, F Javier; Girotti, Alessandra; Jiménez, Pilar; Cordoba-Diaz, Manuel; Cordoba-Diaz, Damián; Girbés, Tomás

    2016-06-10

    Endoglin (CD105) is an accessory component of the TGF-β receptor complex, which is expressed in a number of tissues and over-expressed in the endothelial cells of tumor neovasculature. Targeting endoglin with immunotoxins containing type 2 ribosome-inactivating proteins has proved an effective tool to reduce blood supply to B16 mice tumor xenografts. We prepared anti-endoglin immunotoxin (IT)-containing recombinant musarmin 1 (single chain ribosome-inactivating proteins) linked to the mouse anti-human CD105 44G4 mouse monoclonal antibody via N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP). The immunotoxin specifically killed L929 fibroblast mouse cells transfected with the short form of human endoglin with IC50 values in the range of 5 × 10(-10) to 10(-9) M.

  4. Recombinant Programming

    OpenAIRE

    Pawlak , Renaud; Cuesta , Carlos; Younessi , Houman

    2004-01-01

    This research report presents a promising new approach to computation called Recombinant Programming. The novelty of our approach is that it separates the program into two layers of computation: the recombination and the interpretation layer. The recombination layer takes sequences as inputs and allows the programmer to recombine these sequences through the definition of cohesive code units called extensions. The output of such recombination is a mesh that can be used by the interpretation la...

  5. Common RNA replication signals exist among group 2 coronaviruses: evidence for in vivo recombination between animal and human coronavius molecules

    International Nuclear Information System (INIS)

    Wu, H.-Y.; Guy, James S.; Yoo, Dongwan; Vlasak, Reinhard; Urbach, Ena; Brian, David A.

    2003-01-01

    5' and 3' UTR sequences on the coronavirus genome are known to carry cis-acting elements for DI RNA replication and presumably also virus genome replication. 5' UTR-adjacent coding sequences are also thought to harbor cis-acting elements. Here we have determined the 5' UTR and adjacent 289-nt sequences, and 3' UTR sequences, for six group 2 coronaviruses and have compared them to each other and to three previously reported group 2 members. Extensive regions of highly similar UTR sequences were found but small regions of divergence were also found indicating group 2 coronaviruses could be subdivided into those that are bovine coronavirus (BCoV)-like (BCoV, human respiratory coronavirus-OC43, human enteric coronavirus, porcine hemagglutinating encephalomyelitis virus, and equine coronavirus) and those that are murine hepatitis virus (MHV)-like (A59, 2, and JHM strains of MHV, puffinosis virus, and rat sialodacryoadenitis virus). The 3' UTRs of BCoV and MHV have been previously shown to be interchangeable. Here, a reporter-containing BCoV DI RNA was shown to be replicated by all five BCoV-like helper viruses and by MHV-H2 (a human cell-adapted MHV strain), a representative of the MHV-like subgroup, demonstrating group 2 common 5' and 3' replication signaling elements. BCoV DI RNA, furthermore, acquired the leader of HCoV-OC43 by leader switching, demonstrating for the first time in vivo recombination between animal and human coronavirus molecules. These results indicate that common replication signaling elements exist among group 2 coronaviruses despite a two-cluster pattern within the group and imply there could exist a high potential for recombination among group members

  6. Comparison of the Influence on the Liver Function Between Thyroid Hormone Withdrawal and rh-TSH Before High-Dose Radioiodine Therapy in Patients with Well-Differentiated Thyroid Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Han, Yeon-Hee; Lim, Seok Tae; Yun, Kuk-No; Yim, Sung Kyun; Kim, Dong Wook; Jeong, Hwan-Jeong; Sohn, Myung-Hee [Chonbuk National Univ. Medical School and Hospital, Jeonju (Korea, Republic of)

    2012-06-15

    An elevated thyroid stimulating hormone level (TSH) is essential to stimulate the uptake of radioiodine into thyroid remnants and metastases and metastases of thyroid cancer when a patient under-goes high-dose radioiodine therapy. Nowadays, recombinant human thyroid stimulating hormone (rh-TSH) is increasingly used instead of the classic method of thyroid hormone withdrawal (THW). However, beyond the therapeutic effects, clinical differences between the two methods have not yet been clearly demonstrated. The aim of this work was to investigate the effects of the two methods, especially on liver function. We identified 143 evaluable patients who were further divided into two groups: THW and rh-TSH. We first reviewed the aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels, which were measured during the admission period for total thyroidectomy. We called these liver enzyme levels 'base AST' and 'base ALT.' We also assessed other chemistry profiles, including AST, ALT, total cholesterol, LDL cholesterol, alkaline phosphatase (ALP), total bilirubin (TB), and triglyceride (TG), which were measured on admission day for high-dose radioiodine therapy. We called these liver enzyme levels 'follow-up AST'and 'follow-up ALT.' We compared the changes in base and follow-up liver enzyme levels and the other chemistry profiles between the two groups. The base AST and base ALT levels of the two groups were within normal range, and there was no significant difference between the two groups. In contrast to these base liver enzyme levels, follow-up AST and ALT levels than did the rh-TSH group. Patients in the THW group. Patients in the THW group also had higher levels of total cholesterol and LDL cholesterol than did the patients in the rh-TSH group. However there were no statistically significant differences in ALP, total bilirubin, and triglyceride levels between the two groups. In this retrospective analysis of liver

  7. Characterization of deamidation at Asn138 in L-chain of recombinant humanized Fab expressed from Pichia pastoris.

    Science.gov (United States)

    Ohkuri, Takatoshi; Murase, Eri; Sun, Shu-Lan; Sugitani, Jun; Ueda, Tadashi

    2013-10-01

    A method was previously established for evaluating Asn deamidation by matrix-assisted laser desorption/ionization time of flight-mass spectrometry using endoproteinase Asp-N. In this study, we demonstrated that this method could be applied to the identification of the deamidation site of the humanized fragment antigen-binding (Fab). First, a system for expressing humanized Fab from methylotrophic yeast Pichia pastoris was constructed, resulting in the preparation of ∼30 mg of the purified humanized Fab from 1 l culture. Analysis of the L-chain derived from recombinant humanized Fab that was heated at pH 7 and 100°C for 1 h showed the deamidation at Asn138 in the constant region. Then, we prepared L-N138D Fab and L-N138A Fab and examined their properties. The circular dichroism (CD) spectrum of the L-N138D Fab was partially different from that of the wild-type Fab. The measurement of the thermostability showed that L-N138D caused a significant decrease in the thermostability of Fab. On the other hand, the CD spectrum and thermostability of L-N138A Fab showed the same behaviour as the wild-type Fab. Thus, it was suggested that the introduction of a negative charge at position 138 in the L-chain by the deamidation significantly affected the stability of humanized Fab.

  8. Long-term intravenous treatment of Pompe disease with recombinant human alpha-glucosidase from milk.

    Science.gov (United States)

    Van den Hout, Johanna M P; Kamphoven, Joep H J; Winkel, Léon P F; Arts, Willem F M; De Klerk, Johannes B C; Loonen, M Christa B; Vulto, Arnold G; Cromme-Dijkhuis, Adri; Weisglas-Kuperus, Nynke; Hop, Wim; Van Hirtum, Hans; Van Diggelen, Otto P; Boer, Marijke; Kroos, Marian A; Van Doorn, Pieter A; Van der Voort, Edwin; Sibbles, Barbara; Van Corven, Emiel J J M; Brakenhoff, Just P J; Van Hove, Johan; Smeitink, Jan A M; de Jong, Gerard; Reuser, Arnold J J; Van der Ploeg, Ans T

    2004-05-01

    Recent reports warn that the worldwide cell culture capacity is insufficient to fulfill the increasing demand for human protein drugs. Production in milk of transgenic animals is an attractive alternative. Kilogram quantities of product per year can be obtained at relatively low costs, even in small animals such as rabbits. We tested the long-term safety and efficacy of recombinant human -glucosidase (rhAGLU) from rabbit milk for the treatment of the lysosomal storage disorder Pompe disease. The disease occurs with an estimated frequency of 1 in 40,000 and is designated as orphan disease. The classic infantile form leads to death at a median age of 6 to 8 months and is diagnosed by absence of alpha-glucosidase activity and presence of fully deleterious mutations in the alpha-glucosidase gene. Cardiac hypertrophy is characteristically present. Loss of muscle strength prevents infants from achieving developmental milestones such as sitting, standing, and walking. Milder forms of the disease are associated with less severe mutations and partial deficiency of alpha-glucosidase. In the beginning of 1999, 4 critically ill patients with infantile Pompe disease (2.5-8 months of age) were enrolled in a single-center open-label study and treated intravenously with rhAGLU in a dose of 15 to 40 mg/kg/week. Genotypes of patients were consistent with the most severe form of Pompe disease. Additional molecular analysis failed to detect processed forms of alpha-glucosidase (95, 76, and 70 kDa) in 3 of the 4 patients and revealed only a trace amount of the 95-kDa biosynthetic intermediate form in the fourth (patient 1). With the more sensitive detection method, 35S-methionine incorporation, we could detect low-level synthesis of -glucosidase in 3 of the 4 patients (patients 1, 2, and 4) with some posttranslation modification from 110 kDa to 95 kDa in 1 of them (patient 1). One patient (patient 3) remained totally deficient with both detection methods (negative for cross

  9. HIV-specific humoral and cellular immunity in rabbits vaccinated with recombinant human immunodeficiency virus-like gag-env particles

    International Nuclear Information System (INIS)

    Haffar, O.K.; Smithgall, M.D.; Moran, P.A.; Travis, B.M.; Zarling, J.M.; Hu, S.L.

    1991-01-01

    Recombinant human immunodeficiency virus type-1 (HIV-1)-like gag-env particles produced in mammalian cells were inoculated into two New Zealand white rabbits. In parallel, two control rabbits were inoculated with the homologous HIV-1 virions inactivated by ultraviolet light (uv) and psoralen treatments. The humoral and cellular immune responses to HIV-1 were evaluated for both groups of animals. Recombinant particles elicited humoral immunity that was specific for all the viral structural proteins. The antibodies recognized both denatured and nondenatured proteins. Moreover, the sera neutralized the in vitro infectivity of the homologous virus in CEM cells. Importantly, the recombinant particles also generated a T helper response by priming with the HIV proteins. Similar results were observed with inactivated virus immunization. Therefore, the authors results suggest that the recombinant HIV-like particles elicit functional humoral immunity as well as cellular immunity and represent a novel vaccine candidate for AIDS

  10. Follow up of Graves' Opthalmopathy after radioiodine therapy

    International Nuclear Information System (INIS)

    Miah, M.S.R.; Paul, A. K.; Rahman, H.A.

    2002-01-01

    Graves' ophthalmopathy may first appear or worsen during or after treatment for hyperthyroidism. We followed up 158 Graves' hyperthyroid patients treated with radioiodine of which 49 had Grave's' ophthalmopathy during presentation in Nuclear Medicine Centre, Khulna during the period from 1995 to 2000. The aim of our study is to see the effect of radioiodine in Graves' ophthalmopathy. All the patients received radioiodine at fixed dose regime ranged from 7 mCi to 12 mCi. The duration of follow up was at least 12 months Graves' ophthalmopathy patients, 4 (4/49 i.e., 8.2%) showed exaggeration of ophthalmopathy and the rest (45/49 i.e., 91.8%) remained unchanged. None of ophthalmopathy developed among any of Graves' hyperthyroid or disappeared after radioiodine treatment during follow up period. From the study we concluded that eye changes in Graves' hyperthyroidism remain unchanged or exaggerated after radioiodine therapy and needs ophthalmologist care.(author)

  11. Pore architecture and cell viability on freeze dried 3D recombinant human collagen-peptide (RHC)–chitosan scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Jing; Zhou, Aimei; Deng, Aipeng [School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China); Yang, Yang [Faculty of Engineering, University of Nottingham, Nottingham NG7 2RD (United Kingdom); Gao, Lihu; Zhong, Zhaocai [School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China); Yang, Shulin, E-mail: yshulin@njust.edu.cn [School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China)

    2015-04-01

    Pore architecture of 3D scaffolds used in tissue engineering plays a critical role in the maintenance of cell survival, proliferation and further promotion of tissue regeneration. We investigated the pore size and structure, porosity, swelling as well as cell viability of a series of recombinant human collagen-peptide–chitosan (RHCC) scaffolds fabricated by lyophilization. In this paper, freezing regime containing a final temperature of freezing (T{sub f}) and cooling rates was applied to obtain scaffolds with pore size ranging from 100 μm to 120 μm. Other protocols of RHC/chitosan suspension concentration and ratio modification were studied to produce more homogenous and appropriate structural scaffolds. The mean pore size decreased along with the decline of T{sub f} at a slow cooling rate of 0.7 °C/min; a more rapid cooling rate under 5 °C/min resulted to a smaller pore size and more homogenous microstructure. High concentration could reduce pore size and lead to thick well of scaffold, while improved the ratio of RHC, lamellar and fiber structure coexisted with cellular pores. Human umbilical vein endothelial cells (HUVECs) were seeded on these manufactured scaffolds, the cell viability represented a negative correlation to the pore size. This study provides an alternative method to fabricate 3D RHC–chitosan scaffolds with appropriate pores for potential tissue engineering. - Highlights: • Fabrication of recombinant human collagen-chitosan scaffolds by freezing drying • Influence of freeze drying protocols on lyophilized scaffolds • Pore size, microstructure, porosity, swelling and cell viability were compared. • The optimized porous scaffold is suitable for cell (HUVEC) seeding.

  12. High-efficiency generation of induced pluripotent mesenchymal stem cells from human dermal fibroblasts using recombinant proteins.

    Science.gov (United States)

    Chen, Fanfan; Zhang, Guoqiang; Yu, Ling; Feng, Yanye; Li, Xianghui; Zhang, Zhijun; Wang, Yongting; Sun, Dapeng; Pradhan, Sriharsa

    2016-07-30

    Induced pluripotent mesenchymal stem cells (iPMSCs) are novel candidates for drug screening, regenerative medicine, and cell therapy. However, introduction of transcription factor encoding genes for induced pluripotent stem cell (iPSC) generation which could be used to generate mesenchymal stem cells is accompanied by the risk of insertional mutations in the target cell genome. We demonstrate a novel method using an inactivated viral particle to package and deliver four purified recombinant Yamanaka transcription factors (Sox2, Oct4, Klf4, and c-Myc) resulting in reprogramming of human primary fibroblasts. Whole genome bisulfite sequencing was used to analyze genome-wide CpG methylation of human iPMSCs. Western blot, quantitative PCR, immunofluorescence, and in-vitro differentiation were used to assess the pluripotency of iPMSCs. The resulting reprogrammed fibroblasts show high-level expression of stem cell markers. The human fibroblast-derived iPMSC genome showed gains in DNA methylation in low to medium methylated regions and concurrent loss of methylation in previously hypermethylated regions. Most of the differentially methylated regions are close to transcription start sites and many of these genes are pluripotent pathway associated. We found that DNA methylation of these genes is regulated by the four iPSC transcription factors, which functions as an epigenetic switch during somatic reprogramming as reported previously. These iPMSCs successfully differentiate into three embryonic germ layer cells, both in vitro and in vivo. Following multipotency induction in our study, the delivered transcription factors were degraded, leading to an improved efficiency of subsequent programmed differentiation. Recombinant transcription factor based reprogramming and derivatization of iPMSC offers a novel high-efficiency approach for regenerative medicine from patient-derived cells.

  13. Decreased prothrombotic effects of pegylated recombinant human megakaryocyte growth and development factor in thrombocytopenic state in a rat thrombosis model.

    Science.gov (United States)

    Nishiyama, U; Kuwaki, T; Akahori, H; Kato, T; Ikeda, Y; Miyazaki, H

    2005-02-01

    Previous in vitro studies demonstrated that thrombopoietin (TPO) acts on platelets to activate a variety of intracellular signaling pathways and to enhance platelet sensitivity to multiple agonists. Little is known, however, about whether TPO exerts prothrombotic effects in vivo. The aim of this study was to examine the effects of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF), a pegylated N-terminal domain of human TPO, in a rat model of venous thrombosis. A microthrombus was photochemically induced on the vessel wall of a mesenteric venule, but the vessel was not occluded by it. A single intravenous injection of PEG-rHuMGDF (3 microg kg(-1)) after the thrombus generation into normal rats enhanced the thrombus size, resulting in transient thrombotic occlusion in the majority of rats. Stimulatory effects on thrombus growth were also observed following administration of glycosylated recombinant human full-length TPO (6 microg kg(-1)). In rats rendered thrombocytopenic by total body irradiation, however, PEG-rHuMGDF, even at 300 microg kg(-1), did not induce a significant increase in thrombus size or thrombotic occlusion. Platelets from thrombocytopenic rats had decreased surface levels of c-Mpl and decreased sensitivity to PEG-rHuMGDF in an in vitro aggregation response. Thus, decreased prothrombotic effects of PEG-rHuMGDF in thrombocytopenic rats might be the result not only of low platelet counts but also of decreased platelet reactivity to PEG-rHuMGDF. These results indicate that PEG-rHuMGDF has little effect on venous thrombus formation in thrombocytopenic states associated with high endogenous TPO levels.

  14. Pore architecture and cell viability on freeze dried 3D recombinant human collagen-peptide (RHC)–chitosan scaffolds

    International Nuclear Information System (INIS)

    Zhang, Jing; Zhou, Aimei; Deng, Aipeng; Yang, Yang; Gao, Lihu; Zhong, Zhaocai; Yang, Shulin

    2015-01-01

    Pore architecture of 3D scaffolds used in tissue engineering plays a critical role in the maintenance of cell survival, proliferation and further promotion of tissue regeneration. We investigated the pore size and structure, porosity, swelling as well as cell viability of a series of recombinant human collagen-peptide–chitosan (RHCC) scaffolds fabricated by lyophilization. In this paper, freezing regime containing a final temperature of freezing (T f ) and cooling rates was applied to obtain scaffolds with pore size ranging from 100 μm to 120 μm. Other protocols of RHC/chitosan suspension concentration and ratio modification were studied to produce more homogenous and appropriate structural scaffolds. The mean pore size decreased along with the decline of T f at a slow cooling rate of 0.7 °C/min; a more rapid cooling rate under 5 °C/min resulted to a smaller pore size and more homogenous microstructure. High concentration could reduce pore size and lead to thick well of scaffold, while improved the ratio of RHC, lamellar and fiber structure coexisted with cellular pores. Human umbilical vein endothelial cells (HUVECs) were seeded on these manufactured scaffolds, the cell viability represented a negative correlation to the pore size. This study provides an alternative method to fabricate 3D RHC–chitosan scaffolds with appropriate pores for potential tissue engineering. - Highlights: • Fabrication of recombinant human collagen-chitosan scaffolds by freezing drying • Influence of freeze drying protocols on lyophilized scaffolds • Pore size, microstructure, porosity, swelling and cell viability were compared. • The optimized porous scaffold is suitable for cell (HUVEC) seeding

  15. Evolution of R5 and X4 human immunodeficiency virus type 1 gag sequences in vivo: evidence for recombination

    International Nuclear Information System (INIS)

    Rij, Ronald P. van; Worobey, Michael; Visser, Janny A.; Schuitemaker, Hanneke

    2003-01-01

    Human immunodeficiency virus type 1 (HIV-1) infection is in general established by CCR5-utilizing (R5) virus variants, which persist throughout the course of infection. R5 HIV-1 variants evolve into CXCR4-utilizing (X4) HIV-1 variants in approximately half of the infected individuals. We have previously observed an ongoing genetic evolution with a continuous divergence of envelope gp120 sequences of coexisting R5 and X4 virus variants over time. Here, we studied evolution of gag p17 sequences in two patients who developed X4 variants in the course of infection. In contrast to the envelope gp120 sequences, gag p17 sequences of R5 and X4 virus populations intermingled in phylogenetic trees and did not diverge from each other over time. Statistical evaluation using the Shimodaira-Hasegawa test indicated that the different genomic regions evolved along different topologies, supporting the hypothesis of recombination. Therefore, our data imply that recombination between R5 and X4 HIV-1 variants occurs in vivo

  16. Investigations into, and development of, a lyophilized and formulated recombinant human factor IX produced from CHO cells.

    Science.gov (United States)

    Almeida, Aline G; Pinto, Rodrigo C V; Smales, C Mark; Castilho, Leda R

    2017-08-01

    To develop a recombinant human factor IX (rFIX) formulation equivalent to commercially available products in terms of cake appearance, residual moisture, proportion of soluble aggregates and activity maintenance for 3 months at 4-8 °C. NaCl and low bulking agent/cryoprotectant mass ratio had a negative impact on cake quality upon lyophilisation for a wide range of formulations tested. Particular devised formulations maintained rFIX activity after lyophilization with a similar performance when compared with the rFIX formulated using the excipients reported for a commercially available FIX formulation (Benefix). rFIX remained active after 3 months when stored at 4 °C, though this was not the case with samples stored at 40 °C. Interestingly, particular formulations had an increase in residual moisture after 3 months storage, but not above a 3% threshold. All four formulations tested were equivalent to the Benefix formulation in terms of particle size distribution and cake appearance. Three specific formulations, consisting of surfactant polysorbate-80, sucrose or trehalose as cryoprotectant, mannitol or glycine as bulking agent, L-histidine as buffering agent, and NaCl added in the reconstitution liquid at 0.234% (w/v) were suitable for use with a CHO cell-derived recombinant FIX.

  17. Analysis of antigenic cross-reactivity between subgroup C avian pneumovirus and human metapneumovirus by using recombinant fusion proteins.

    Science.gov (United States)

    Luo, L; Sabara, M I; Li, Y

    2009-10-01

    Avian pneumovirus subgroup C (APV/C) has recently been reported to be more closely related to human metapneumovirus (hMPV) as determined by sequence analysis. To examine the antigenic relationship between APV/C and hMPV, the APV/C fusion (F) gene was cloned and expressed as an uncleaved glycoprotein in a baculovirus system. The reactivity of the APV/C F protein with antibodies against APV subgroups A, B, C, and hMPV was examined by Western blot analysis. The results showed that the expressed APV/C F protein was not only recognized by APV/C-specific antibodies but also by antibodies raised against hMPV. Previously expressed recombinant hMPV F protein also reacted with APV/C-specific antibodies, suggesting that there was significant antigenic cross-reactivity and a potential evolutionary relationship between hMPV and APV/C. Interestingly, the recombinant F proteins from APV/C and hMPV were not recognized by polyclonal antibodies specific to APV subgroups A and B.

  18. Studies on mechanism of treatment of granulocyte colony-stimulating factor, recombinant human interleukin-11 and recombinant human interleukin-2 on hematopoietic injuries induced by 4.5 Gy γ-rays irradiation in beagles

    International Nuclear Information System (INIS)

    Li Ming; Ou Hongling; Xing Shuang; Huang Haixiao; Xiong Guolin; Xie Ling; Zhao Yanfang; Zhao Zhenhu; Wang Ning; Wang Jinxiang; Miao Jingcheng; Zhu Nankang; Luo Qingliang; Cong Yuwen; Zhang Xueguang

    2010-01-01

    Objective: To investigate the mechanism of treatment of granulocyte colony-stimulating factor (rhG-CSF), recombinant human interleukin-11 (rhIL-11) and recombinant human interleukin-2 (rhIL-2) on hematopoietic injuries induced by 4.5 Gy 60 Co γ-ray irradiation in beagles, and to provide experimental evidence for the clinical treatment of extremely severe myeloid acute radiation sickness (ARS). Methods: Sixteen beagle dogs were given 4.5 Gy 60 Co γ-ray total body irradiation (TBI), then randomly assigned into irradiation control group, supportive care group or cytokines + supportive care (abbreviated as cytokines) group. In addition to supportive care, rhG-CSF, rhIL-11 and rhIL-2 were administered subcutaneously to treat dogs in cytokines group. The percentage of CD34 + cells, cell cycle and apoptosis of nucleated cells in peripheral blood were examined by Flow cytometry. Results: After 4.5 Gy 60 Co γ-ray irradiation, the CD34 + cells in peripheral blood declined obviously (61.3% and 52.1% of baseline for irradiation control and supportive care group separately). The cell proportion of nucleated cells in G 0 /G 1 phase was increased notably notably (99.27% and 99.49% respectively). The rate of apoptosis (26.93% and 21.29% separately) and necrosis (3.27% and 4.14%, respectively) of nucleated cells were elevated significantly when compared with values before irradiation (P 0 /G 1 phase blockage of nucleated cells became more serious (99.71%). The rate of apoptosis (5.66%) and necrosis (1.60%) of nucleated cells were significantly lower than that of irradiation control and supportive care groups 1 d after exposure. Conclusions: Cytokines maybe mobilize CD34 + cells in bone marrow to peripheral blood, indce cell block at G 0 /G 1 phase and reduce apoptosis, and eventually cure hematopoietic injuries induced by irradiation. (authors)

  19. Value of the Serum Thyroglobulin Level Alteration at the First High Dose Radioiodine Treatment in Patients with Differentiated Thyroid Carcinoma

    International Nuclear Information System (INIS)

    Nam, Hyun Yeol; Kim, In Joo; Kim, Yong Ki; Kim, Seong Jang; Jun, Sung Min; Kim, Bum Soo

    2009-01-01

    The purpose of this study was to evaluate if short-term serum thyroglobulin (Tg) elevation after radioiodine administration can predict successful radioiodine remnant ablation (RRA) and whether comparable RRA effectiveness is exhibited between a group administered with recombinant human thyrotropin (rhTSH) and a group experiencing thyroid hormone withdrawal (THW), in preparation for RRA. A retrospective chart review was performed on 39 patients in the rhTSH group and 46 patients in the THW group. They were treated for differentiated thyroid carcinoma by total or near total thyroidectomy, and referred for RRA between 2003 and 2006 (the rhTSH group) and between January and June of 2006 (the THW group). They were assessed for serum Tg levels just before I-131 administration (TgD0), reassessed 9 days later (TgD9), and again 6-12 months later. RRA was successful in 64 (37 from the THW group and 27 from the rhTSH group) of the total 85 patients. The success rates of RRA had no statistically significant differences between the two groups. In both groups, TgD9/TgD0 values were significantly higher in the RRA success group (the rhTSH group; P=0.03, the THW group; P=0.04). By combining cutoff values of TgD0 and TgD9/TgD0, the successful RRA value was determined to be 96.7% (29/30) with TgD0≤5.28 ng/mL and TgD9/TgD0>4.37 in both groups (the rhTSH group; 100% (16/16), the THW group; 92.9% (13/14)). Using logistic multivariate analysis, only TgD0 was independently associated with successful RRA. We may predict successful ablation by evaluating short-term serum Tg elevation after I-131 administration for RRA, in both rhTSH and THW patients

  20. Air Monitoring to Control the Intake of Airborne Radioiodine-131 Contaminants by Nuclear Medicine Workers

    International Nuclear Information System (INIS)

    Jiemwutthisak, P.; Sritongkul, N.; Chaudakshetrin, P.; Kanchanaphiboon, P.; Tuntawiroon, M.

    2012-01-01

    were less than the DAC of 416.67 Bq/m 3 assigned for I-131. This demonstrated that the work place condition is satisfactory and the level of protection provided to the worker is sufficient to minimize the internal dose equivalent. The inhalation of radioiodine vapors is the important routes of entry of radioiodine into the human body. The level of airborne I-131 radioactivity should be maintained well below allowable levels to keep the internal dose equivalent contribution to the total effective dose equivalent small. (author)

  1. Differentiated thyroid carcinoma referred for radioiodine therapy

    International Nuclear Information System (INIS)

    Al-Balawi, Ibrahim A.; Meir, Hadir M.; Yousef, Mohammad K.; Nayel, Hala A.; Al-Mobarak, Mohammad F.

    2001-01-01

    The current work was conducted to study the disease status and treatment results of patients with differentiated thyroid carcinoma referred for radioactive iodine therapy. Retrospective review of 78 patients with differentiated thyroid carcinoma referred for radioiodine therapy in the Nuclear Medicine Unit, King Abdulaziz Hospital and Oncology Center, Jeddah, Kingdom of Saudi Arabia. Analysis of the clinicopathologic characteristics, age correlation to different risk factors, treatment protocol and results were performed. Seventy seven percent were female and the female to male ratio was 3.5:1. The age of patients ranged between 13-63 years with a median age of 36 years. Cervical lymph node involvement was detected in 22 patients (25%). Papillary carcinoma was encountered in 78 patients (90%) and follicular carcinoma in 9 patients (10%). Analysis of the clinicopathologic characteristics showed no statistically significant difference between patients in the different age groups except for extrathyroid extension and lymph node involvement. Patients older than 45 years had a statistically significant lower incidence of nodal involvement and higher incidence of extra thyroid extension (P<0.02). In the current study we used a high dose method (Radioiodine-131 dose 75-100mCi) for thyroid remnant ablation after thyroidectomy (total or near total) in 67 patients. An Iodine 131 dose of 150 mCi was used in 12 patients with radioiodine-avid cervical lymph nodes and in 3 patients with gross residual tumor. In 4 patients with distant metastases an Iodine 131 dose of 200 mCi was used. For the whole study group the 5 year overall survival and disease-free survival was 96% and 88%. The current study, as with many other retrospective studies, concluded that despite the fact that differentiated thyroid carcinoma is among the most curable cancers, some patients are still at high risk for recurrent disease and associated mortality. (author)

  2. Endogenous acute phase serum amyloid A lacks pro-inflammatory activity, contrasting the two recombinant variants that activate human neutrophils through different receptors

    Directory of Open Access Journals (Sweden)

    Karin eChristenson

    2013-04-01

    Full Text Available Most notable among the acute phase proteins is serum amyloid A (SAA, levels of which can increase 1000-fold during infections, aseptic inflammation, and/or trauma. Chronically elevated SAA levels are associated with a wide variety of pathological conditions, including obesity and rheumatic diseases. Using a recombinant hybrid of the two human SAA isoforms (SAA1 and 2 that does not exist in vivo, numerous in vitro studies have given rise to the notion that acute phase SAA is a pro-inflammatory molecule with cytokine-like properties. It is however unclear whether endogenous acute phase SAA per se mediates pro-inflammatory effects. We tested this in samples from patients with inflammatory arthritis and in a transgenic mouse model that expresses human SAA1. Endogenous human SAA did not drive production of pro-inflammatory IL-8/KC in either of these settings. Human neutrophils derived from arthritis patients displayed no signs of activation, despite being exposed to severely elevated SAA levels in circulation, and SAA-rich sera also failed to activate cells in vitro. In contrast, two recombinant SAA variants (the hybrid SAA and SAA1 both activated human neutrophils, inducing L-selectin shedding, production of reactive oxygen species, a