Incidence of drug-induced torsades de pointes with intravenous amiodarone

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Jayaprakash Shenthar

2017-11-01

Conclusion: The incidence of drug-induced TdP with IV amiodarone is about 1.5%. Risk factors include female sex, left ventricular dysfunction, electrolyte abnormalities, baseline prolonged QTc, concomitant beta-blocker, and digoxin therapy. Amiodarone induced TdP has favorable prognosis if recognized and treated promptly, and these patients should not receive amiodarone by any route in future.

Analysis of computed tomography density of liver before and after amiodarone administration.

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Matsuda, Masazumi; Otaka, Aoi; Tozawa, Tomoki; Asano, Tomoyuki; Ishiyama, Koichi; Hashimoto, Manabu

2018-05-01

To evaluate CT density of liver changes between before and after amiodarone administration. Twenty-five patients underwent non-enhanced CT including the liver before and after amiodarone administration. We set regions of interest (ROIs) at liver S8, spleen, paraspinal muscle, and calculated average CT density in these ROIs, then compared CT density between liver and other organs. Statistical differences between CT density of liver and various ratios before and after administration were determined, along with correlations between cumulative dose of amiodarone and liver density after administration, density change of liver, and various ratios after administration. Liver density, liver-to-spleen ratio, and liver-to-paraspinal muscle ratio differed significantly between before and after amiodarone administration. No significant correlations were found between cumulative doses of amiodarone and any of liver density after administration, density change of liver, or various ratios after administration. CT density of liver after amiodarone administration was significantly higher than that before administration. No correlations were identified between cumulative dose of amiodarone and either liver density after administration or density change of liver. Amiodarone usage should be checked when radiologists identify high density of the liver on CT.

Amiodarone: Effects on thyroid function and the peripheral metabolism of the thyroid hormones

International Nuclear Information System (INIS)

Braverman, L.E.; Safran, M.; Bambini, G.; Pinchera, A.; Martino, E.

1985-01-01

In addition to the effects of Amiodarone on the peripheral metabolism of the thyroid hormones and on pituitary TSH secretion, a major complication of therapy is the relatively high frequency of iodide-induced thyroid dysfunction. The mean T 4 and T 3 concentration following Amiodarone application was measured in euthyroid, hypothyroid and hyperthyroid patients and in control patients with and without cardiac disorders. Furthermore, the serum TSH was determined in euthyroid Amiodarone-treated euthyroid patients. 131 I uptake was studied in patients with Amiodarone-associated thyrotoxicosis. The difficulties of the therapy of Amiodarone-induced hyper-thyroidism are outlined. Preliminary studied of the effect of Amiodarone and its analogues on the metabolism of thyroid hormones in the rat indicate that Amiodarone may act as a thyroid hormone agonist in the pituitary. (MG)

Amiodarone-induced thyroid dysfunction

African Journals Online (AJOL)

multinodular disease or Graves' disease, who on exposure to an excessive .... where the cardiovascular status permitted this, radioactive ... antedating the initiation of amiodarone therapy in the current cohort. .... an evidence-based protocol.

Effect of amiodarone therapy on mortality in patients with left ventricular dysfunction and asymptomatic complex ventricular arrhythmias: Argentine Pilot Study of Sudden Death and Amiodarone (EPAMSA).

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Garguichevich, J J; Ramos, J L; Gambarte, A; Gentile, A; Hauad, S; Scapin, O; Sirena, J; Tibaldi, M; Toplikar, J

1995-09-01

The efficiency of prophylactic antiarrhythmic treatment with amiodarone in reducing 1-year mortality in patients with reduced left ventricular ejection fraction ( < 35%) and asymptomatic ventricular arrhythmias (Lown classes 2 and 4) was investigated in a prospective, multicenter, randomized, controlled study. Among 127 patients who entered the study, 61 were assigned to no antiarrhythmic therapy (control group [CG] and 66 to amiodarone treatment (amiodarone group [AG]). Amiodarone was administered at a dosage of 800 mg/day for 2 weeks followed by 400 mg/day thereafter. A 12-month follow-up was completed for 106 patients (57 in the AG and 49 in the CG). Amiodarone reduced the overall mortality rate, which was 10.5% in the AG versus 28.6% in the CG (odds ratio [OR] 0.29; 95% confidence interval [CI] 0.10 to 0.84; log-rank test 0.02) and sudden death rate, which was 7.0% in the AG versus 20.4% in the CG (OR 0.29; 95% CI 0.08 to 1.00; log-rank test 0.04). Side effects were rare, and in only three patients did amiodarone treatment have to be discontinued.

Peripheral amiodarone-related phlebitis: an institutional nursing guideline to reduce patient harm.

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Spiering, Mary

2014-01-01

Intravenous amiodarone is one of the most widely used antiarrythmics for the treatment of atrial fibrillation with rapid ventricular response. Peripheral amiodarone infusion, however, often causes pain during infusion and subsequent phlebitis.Data collection on a cardiac telemetry unit revealed a high rate of phlebitis. A multidisciplinary team developed and implemented amiodarone peripheral infusion guidelines. The pre-guideline phlebitis rate was 85% and post-guideline rate was 38%, representing a 47% change or improvement. An additional finding was that the severity of phlebitis was reduced, as well. The results of this study suggest that the implementation of a peripheral amiodarone infusion guideline reduced the incidence and severity of amiodarone-related phlebitis in the cardiac population.

Amiodarone-Induced Thyrotoxic Thyroiditis: A Diagnostic and Therapeutic Challenge

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Umang Barvalia

2014-01-01

Full Text Available Amiodarone is an iodine-based, potent antiarrhythmic drug bearing a structural resemblance to thyroxine (T4. It is known to produce thyroid abnormalities ranging from abnormal thyroid function testing to overt hypothyroidism or hyperthyroidism. These adverse effects may occur in patients with or without preexisting thyroid disease. Amiodarone-induced thyrotoxicosis (AIT is a clinically recognized condition commonly due to iodine-induced excessive synthesis of thyroid, also known as type 1 AIT. In rare instances, AIT is caused by amiodarone-induced inflammation of thyroid tissue, resulting in release of preformed thyroid hormones and a hyperthyroid state, known as type 2 AIT. Distinguishing between the two states is important, as both conditions have different treatment implications; however, a mixed presentation is not uncommon, posing diagnostic and treatment challenges. We describe a case of a patient with amiodarone-induced type 2 hyperthyroidism and review the current literature on the best practices for diagnostic and treatment approaches.

Acute lung affection in an endurance-trained man under amiodarone medication

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Saurbier, Bernward

2005-06-01

Full Text Available Patients undergoing treatment with amiodarone can develop severe pulmonary side effects. This effect, which is often highly underestimated, can lead to dyspnea, pneumonitis, and further fibrosis. A recent change in the labeling of amdiodarone by the American Food and Drug Administration (FDA supports this suspicion. Tracing the symptoms back to the causing agent can be difficult, as shown in our report. The subject of this case report is an endurance-trained 65 year old male marathon runner who appeared with atrial fibrillation during a routine check up in autumn 2003. After medical cardioversion with flecainide a complaint free interval of 8 months was followed by a relapse, which resulted in a change of medication to amiodarone. Due to misunderstandings the patient kept on taking the amiodarone loading dose for six weeks and returned with severe dyspnea on exertion. Losses in CO diffusing capacity, a lowered macrophages count and a positive lymphocyte transformation test were the only first hand clinical evidence of amiodarone intoxication, despite the sensation of dyspnea. This case shows that special care has to be taken in treatment with amiodarone. Side effects can be hard to trace and do not evidently show a clear connection to amiodarone.

Assistance algorithm of nursing for amiodarone intravenous infusion

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Francimar Tinoco de Oliveira

2014-12-01

Full Text Available This study aimed at identifying scientific publication on phlebitis caused by amiodarone and proposes a nursing care algorithm for interventions in intravenous amiodarone administration grounded in the Infusion Nursing Society and the Center for Disease Control and Prevention. It is a descriptive study mediated by integrative review in MedLine, LILACS, IBECS, BDENF, Cochrane Library and Scielo bases, published from 2006 to 2013. The sample consisted of nine articles. The evidence pointed the incidence of phlebitis due to the infusion of amiodarone and the need to control this event. The algorithm proposed shows the materials to be used and the procedure of drug administration in order to minimize injury. Besides subsidizing the development of future studies, this algorithm also promotes the incorporation of the best recommendation for the interventionist clinical practice.

Treatment of amiodarone-induced thyrotoxicosis resistant to conventional therapy

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Nišić Tanja

2017-01-01

Full Text Available Introduction: Amiodarone as an antiarrhythmic medication is necessary in the prevention and treatment of malignant ventricular arrhythmias, however, it can induce thyroid dysfunction. Thyroid dysfunction may be either hypothyroidism or thyrotoxicosis, however, 50% of patients who have used amiodarone are euthyroid. Case report: A 27-year-old female patient, hospitalized at the Clinic for Endocrinology due to type 2 amiodarone-induced thyrotoxicosis. The patient had previously received amiodarone for two years. At age 25, the patient was diagnosed with dilated cardiomyopathy (EF 25%, EDD/ESD 56-57/47 mm with mild Ebstein’s anomaly, WPW Sy and recorded episodes of non-sustained VT. In order to reduce the risk of sudden death and prevent malignant ventricular arrhythmias, ICD-VR was implanted and amiodarone was prescribed. Treatment with propylthiouracil (PTU and dexamethasone was initiated after thyrotoxicosis was diagnosed. Three weeks after the introduction of PTU, hepatotoxicity was registered, thus the medication was discontinued. Thyrozol, which regulates the hepatotoxicity parameters, was introduced. Sodium perchlorate and glucocorticoid (per os, IV and intrathyroidal therapy was introduced. The treatment had lasted for fifty days and laboratory signs of thyrotoxicosis were still present, which is why a total of eight plasmapheresis sessions were performed. Each plasmapheresis resulted in a significant decrease in FT4 and a slight decrease in FT3. After seventy two days of treatment, an optimal hormonal status of the thyroid gland was established and total thyroidectomy was performed. Conclusion: Patient was treated for amiodarone-induced thyrotoxicosis (AIT type 2, which was resistant to conventional therapy for a long period of time. Successful treatment was achieved by applying plasmapheresis although the effect of perchlorate and glucocorticoids application cannot be disregarded.

Incidence and severity of phlebitis in patients receiving peripherally infused amiodarone.

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Boyce, Brenda A Brady; Yee, Barbara Homer

2012-08-01

Nurses noted that the rate of phlebitis was high when intravenous amiodarone was infused via a peripheral site. Hospital policy recommends a central vascular catheter, but this method is often not feasible because the drug is administered in emergent situations for short periods. To determine the rate and severity of phlebitis in patients given peripherally infused amiodarone. The literature, policy, and procedures for administration of amiodarone were reviewed; the pharmacy was consulted; and a data collection tool was developed. The tool was pilot tested and revised, and face validation was established. Data were collected during a 6-month period. A convenience sample was used. The study included a total of 12 patients. Each new infusion of intravenous amiodarone was considered a separate occurrence, for a total of 24 infusions. Various grades of phlebitis developed in 8 patients (67%). Phlebitis developed at 12 of the 24 infusion sites (50%). Patients receiving peripherally infused amiodarone are at high risk for phlebitis. This complication may lead to infection, additional medical intervention, delay in treatment, and prolonged hospitalization.

Amiodarone Cost Effectiveness in Preventing Atrial Fibrillation After Coronary Artery Bypass Graft Surgery

DEFF Research Database (Denmark)

Zebis, Lars Riber; Christensen, Thomas Decker; Kristiansen, Ivar S.

2008-01-01

patients were included to receive either 300 mg amiodarone or placebo (5% aqueous dextrose solution) administered intravenously over 20 minutes followed by 600 mg amiodarone/placebo orally twice a day (8 am and 8 pm) for the first 5 postoperative days. RESULTS: In the amiodarone group, there were 14 cases...

Bronchiolitis obliterans organising pneumonia in patients taking acebutolol or amiodarone.

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Camus, P; Lombard, J N; Perrichon, M; Piard, F; Guérin, J C; Thivolet, F B; Jeannin, L

1989-01-01

Two patients, treated with acebutolol and amiodarone respectively, developed a disease clinically, radiologically, and pathologically indistinguishable from bronchiolitis obliterans organising pneumonia. In one case recovery followed discontinuation of acebutolol; in the other case cessation of amiodarone had no effect, and corticosteroids were required. In addition to these patients, several cases of bronchiolitis obliterans organising pneumonia have been reported during treatment with gold salts, amiodarone, and miscellaneous other drugs. Taken together, this information supports the view that bronchiolitis obliterans organising pneumonia may be a form of response by the lungs to insult by drugs. Images PMID:2588206

Sorption of amiodarone to polyvinyl chloride infusion bags and administration sets.

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Weir, S J; Myers, V A; Bengtson, K D; Ueda, C T

1985-12-01

The loss of amiodarone from i.v. admixtures to flexible polyvinyl chloride (PVC) infusion bags and i.v. administration sets was studied. Admixtures containing amiodarone hydrochloride 600 micrograms/mL and either 5% dextrose injection or 0.9% sodium chloride injection were stored at room temperature in glass bottles (both with and without contact of the drug solution with the rubber bottle closure), in flexible PVC bags, or in rigid PVC bottles. After 120 hours, the contents of each flexible PVC bag were emptied and replaced by methanol, which was allowed to remain in the bag for an additional 120 hours and was then analyzed for amiodarone content. To determine availability of amiodarone after infusion through a 1.8-m PVC i.v. administration set, solutions stored in glass containers were run through the set at 0.5 mL/min for 90 minutes. Samples of drug solutions were collected at appropriate intervals and analyzed by a stability-indicating high-performance liquid chromatography (HPLC) assay. Admixtures containing 0.9% sodium chloride injection were not stable; visual incompatibility was evident after 24 hours of storage in glass bottles, and no further testing was performed. In admixtures containing 5% dextrose injection that were stored in 50-mL flexible PVC bags, 60% of the initial amiodarone concentration remained after 120 hours; approximately half of the lost drug was recovered with the methanol. In effluent collected from the PVC administration set, 82% of the initial amiodarone concentration remained. Amiodarone concentrations did not decrease appreciably, after storage in glass or rigid PVC bottles, indicating that drug loss was probably affected by the plasticizer, di-2-ethylhexyl phthalate.(ABSTRACT TRUNCATED AT 250 WORDS)

Phlebitis in amiodarone administration: incidence, contributing factors, and clinical implications.

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Norton, Linda; Ottoboni, Linda K; Varady, Ann; Yang-Lu, Chia-Yu; Becker, Nancy; Cotter, Theresa; Pummer, Eileen; Haynes, Annette; Forsey, Lynn; Matsuda, Kelly; Wang, Paul

2013-11-01

Intravenous amiodarone is an important treatment for arrhythmias, but peripheral infusion is associated with direct irritation of vessel walls and phlebitis rates of 8% to 55%. Objectives To determine the incidence and factors contributing to the development of amiodarone-induced phlebitis in the coronary care unit in an academic medical center and to refine the current practice protocol. Medical records from all adult patients during an 18-month period who received intravenous amiodarone while in the critical care unit were reviewed retrospectively. Route of administration, location, concentration, and duration of amiodarone therapy and factors associated with occurrence of phlebitis were examined. Descriptive statistics and regression methods were used to identify incidence and phlebitis factors. In the final sample of 105 patients, incidence of phlebitis was 40%, with a 50% recurrence rate. All cases of phlebitis occurred in patients given a total dose of 3 g via a peripheral catheter, and one-quarter of these cases (n = 10) developed at dosages less than 1 g. Pain, redness, and warmth were the most common indications of phlebitis. Total dosage given via a peripheral catheter, duration of infusion, and number of catheters were significantly associated with phlebitis. Amiodarone-induced phlebitis occurred in 40% of this sample at higher drug dosages. A new practice protocol resulted from this study. An outcome study is in progress.

Amiodarone-induced hypothyroidism. A common complication of prolonged therapy: a report of eight cases

International Nuclear Information System (INIS)

Hawthorne, G.C.; Campbell, N.P.; Geddes, J.S.; Ferguson, W.R.; Postlethwaite, W.; Sheridan, B.; Atkinson, A.B.

1985-01-01

Amiodarone is a widely used antiarrhythmic drug, which contains 75 mg of iodide per 200 mg of active substance. Eight patients receiving long-term amiodarone therapy became hypothyroid. Seven of these patients had no previous history of thyroid dysfunction or goiter. Antithyroid antibodies were absent, and standard perchlorate discharge tests were positive in seven patients when hypothyroidism was diagnosed. In one patient, amiodarone therapy was withdrawn; over the next nine months, the hypothyroidism resolved, and results of the perchlorate discharge test reverted to normal. The authors conclude that amiodarone-induced hypothyroidism is similar to previously described iodide-induced hypothyroidism. It may develop in the absence of a previous history of thyroid disease, and all patients receiving long-term amiodarone therapy should therefore be regularly monitored for hypothyroidism

The Characteristics of Amiodarone-induced Thyrotoxicosis in a Moderate Iodine Deficit Area

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Ancuța-Elena Cota

2013-08-01

Full Text Available Introduction: Amiodarone (AMI, a class III anti-arrhythmic drug, is associated with a number of side effects, including thyroid dysfunction (both hypo- and hyperthyroidism, which is due to amiodarone's high iodine content and its direct toxic effect on the thyroid. Objective: To evaluate the incidence of Amiodarone induced thyrotoxicosis (AIT (type, rate of occurrence and to identify the risk factors involved in its occurrence. Material and method: We examined patients treated with amiodarone, between January 2002 and December 2011, who presented to our Department of Endocrinology Târgu Mures for thyroid dysfunctions. Results: The retrospective study included 87 patients with thyroid dysfunctions; 58 (66.7% patients had AIT and 29 (33.3% had Amiodarone induced hypothyroidism (AIH. In the AIT group: 35 were women (60.3%, 23 were men (39.7%; the average age was 61.60 ± 12.39 years. Risk factors identified for the AIT group were male gender (RR = OR = 3.8; Chi-squer = 5.7, p = 0.004 and pre-existing thyroid abnormalities (RR = 2.5, Chi-square = 4.1, p = 0.005. The thyroid dysfunction occurrence was heterogeneous (0.2-183 months. The patients with previous thyroid abnormalities developed earlier thyroid dysfunction compared to those with an apparently normal thyroid gland (22.25 ± 4.14 months versus 32.09 ± 7.69 months, p = 0.02, T test. Conclusion: In the context of the specific iodine geoclimatic intake and the area of origin, amiodarone - induced thyroid dysfunction spectrum is dominated by thyrotoxicosis. Screening and monitoring of thyroid function for patiens under chronic amiodarone treatment is necessary

Dronedarone and Amiodarone Induce Dyslipidemia and Thyroid Dysfunction in Rats

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Li-Qin Jiang

2016-05-01

Full Text Available Background/Aims: Amiodarone, a thyroid hormone-like molecule, can induce dyslipidemia and thyroid dysfunction. However, the effects of dronedarone on lipid metabolism and of both dronedarone and amiodarone on thyroid function and lipid metabolism remain unknown. Methods: Fifty male Sprague-Dawley rats were randomly divided into 5 groups (10 in each group: normal control (NC, amiodarone-treated (AMT, dronedarone-treated (DRT, rats treated with amiodarone combined with polyene phosphatidylcholine (AC, and rats treated with dronedarone combined with polyene phosphatidylcholine (DC. Rats were given amiodarone (120 mg/kg/d, dronedarone (120 mg/kg/d, and polyene phosphatidylcholine (200 mg/kg/d for 13 weeks. At the end of weeks 4, 8, 12, and 13, plasma-free triiodothyronine (FT3, free thyroxine (FT4, triglycerides (TG, total cholesterol (TC, low-density lipoprotein cholesterol (LDL-c, and high-density lipoprotein cholesterol (HDL-c were determined. At the end of this protocol, rats were sacrificed and the thyroid glands were isolated, weighed, and examined histopathologically. The protein expression of Bcl-2 was measured by immunochemical staining. The mRNA expression of thyroglobulin (Tg, type-1 deiodinase (D1, and thyroid peroxidase (TPO were detected by polymerase chain reaction (PCR. Results: Compared with the NC group, FT3 and FT4 levels in the DRT and DC groups significantly increased at week 4 but declined thereafter. The AMT and AC groups had lower FT3 levels but comparable FT4 levels. The levels of TG, LDL-c, and HDL-c in the NC group were lower than those in the other groups whereas the LDL-c/HDL-c ratio was lowest in the AMT group. Bcl-2 expression significantly increased in the DRT group. The mRNA expression of Tg increased whereas the mRNA expression of D1 decreased. Dronedarone induced hyperthyroidism at the early stage and hypothyroidism at the late stage whereas amiodarone only caused hypothyroidism. Conclusion: Both dronedarone and

SAFETY OF AMIODARONE USAGE IN PATIENTS WITH WOLFF-PARKINSON-WHITE SYNDROME AND ATRIAL FIBRILLATION

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D. A. Kuzhel

2010-01-01

Full Text Available Amiodarone is one of the basic antiarrhytmic drugs for atrial fibrillation treatment. However application of amiodarone in patients with atrial fibrillation and Wolff-Parkinson-White syndrome can induce ventricular fibrillation. Amiodarone usage in these patients should be accompanied by readiness for performance of resuscitation. This is confirmed by clinical case presentation.

[Acute onset pulmonary toxicity associated to amiodarone].

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Ferreira, Pedro Gonçalo; Saraiva, Fátima; Carreira, Cláudia

2012-01-01

Amiodarone is a potent anti-arrhythmic drug with a well-known potential chronic pulmonary toxicity. We describe a case of acute pulmonary toxicity (APT) induced by amiodarone in a 57 year old patient submitted to a perfusion of 900 mg in just 6 hours, to control an auricular flutter with rapid ventricular response. During the administration, the patient developed hemodynamic instability and oxygen dessaturation that led to an electrical cardioversion with return of sinus rhythm. Still, the patient continued in progressive respiratory deterioration with acute bilateral infiltrates on chest x-ray and apparent normal cardiac filling pressures confirmed by echocardiography. Anon-cardiogenic pulmonar edema progressing to clinico-physiological ARDS criteria was diagnosed. Expeditive therapeutic measures were undertaken, namely by initiation of non-invasive positive airway pressure support, that attained a good result.Albeit rare, amiodarone-induced APT might have severe consequences, namely progression to ALI/ARDS with a high mortality index.As it is a frequently prescribed drug, there should be a high clinical suspicion towards this phenomenon, allowing precocious therapeutic measures to be taken in a timely fashion to prevent the associated unfavorable outcome.

Comparative efficacy of amiodarone with ivabradin combination or amiodarone with bisoprolol combination in the prevention of atrial fibrillation recurrence in pa- tients with left ventricular diastolic dysfunction

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K. G. Adamyan

2015-01-01

Full Text Available Aim. To study the efficacy of use of amiodarone with ivabradine combination or amiodarone with bisoprolol combination in the prevention of atrial fibrillation (AF recurrence in patients (pts with left ventricular diastolic dysfunction (LVDD after conversion to sinus rhythm. Material and methods. 65 patients (40 males, 25 females aged 53±8 years with persistent AF and LVDD were included into the study and randomized into 3 groups to receive ivabradine and amiodarone (22 pts, bisoprolol and amiodarone (22 pts or amiodarone alone (21 pts. Left atrium (LA volume indices, LA longitudinal strain rate (LASR in systole, LV mass index, mean heart rate (HR, 24-hour HR variability and the incidence of AF by 96 h ECG monitoring were measured after the titration period, and after 3 and 6 months of follow-up. Results. After 6 months of follow-up group 1 revealed significantly lower maximum LA volume index (21.3±2.4 vs 25.2±3.0 and 28.7±3.6 ml/m2 in the 2nd and control groups, respectively, P-wave LA volume index (15.3±3.5 versus 18.1±3.8 and 20.4±4.0 ml/m2 in the 2nd and control groups, respectively, and LA systolic volume index (7.3±1.2 versus 9.4±1.6 and 9.6±1.7 ml/m2 in 2nd and control groups, respectively. The incidence of side effects in group 1 was significantly less than that in group 2 and was not different compared with control group. Conclusion. Ivabradine and amiodarone combination provides better prevention of AF recurrence and less side-effects in pts with LVDD and persistent AF after sinus rhythm restoration as compared with bisoprolol and amiodarone combination, it also reduces LA maximum, conduit and systolic volumes, and increases LASR.

Amiodarone Hepatotoxicity with Absent Phospholipidosis and Steatosis: A Case Report and Review of Amiodarone Toxicity in Various Organs

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Adela Cimic

2013-01-01

Full Text Available We present the first description of amiodarone toxicity in the liver without phospholipidosis or steatosis. In doing so, we will review the various effects of amiodarone toxicity in various organs. The patient is a young adult who had cardiac reconstruction as a child for transposition of the great vessels. A needle biopsy was taken due to elevated liver enzymes. Her ALT was 188 U/L (5–50 and AST 162 U/L (5–50. Alkaline phosphatase, total bilirubin, protein, and albumin were within normal limits. A serologic panel for viral hepatitis was negative. Antinuclear antibodies were positive at 260; however, anti-smooth muscle antibody and anti-mitochondrial antibody were negative. A protein electrophoresis showed a slightly elevated beta globulin 2 level of 0.5. Quantitative immunoglobulin levels were within normal limits except for a slightly elevated IgA 409 mg/dL (60–350. Liver ultrasound was unremarkable. The clinical differential was broad and included hepatic congestion along with autoimmune hepatitis. Sections showed only ballooned hepatocytes with Mallory-Denk bodies and perisinusoidal fibrosis. Arrival to the diagnosis was possible only after careful review of the patient’s medications. After discontinuation of amiodarone, the patient’s liver enzymes returned to normal levels.

The effects of sesame oil on the prevention of amiodarone-induced phlebitis

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Masoumeh Bagheri-Nesami

2015-01-01

Full Text Available Background: Phlebitis is the most common complication associated with peripheral intravenous infusion of amiodarone. The purpose of this study is to determine the effects of sesame oil on the prevention of amiodarone-induced phlebitis. Materials and Methods: This is a double-blind randomized controlled trial. Thirty-six patients hospitalized in a coronary care unit were randomly allocated into two groups using a convenience sampling method. Following peripheral intravenous cannulation, five drops of pure sesame oil were applied to the skin within a 10 cm radius of the infusion site prior to the administration of amiodarone in the intervention group. Sesame oil was rubbed on the skin at the infusion site every 6 h in the 24-h period of amiodarone infusion. In the control group, liquid paraffin, used as a placebo, was rubbed on the skin at the infusion site of amiodarone. Both groups were monitored for the development of phlebitis and its degree within the 24-h period of amiodarone infusion as well as 6 h after its administration. The incidence of phlebitis was confirmed and recorded by an assessor who was blind to the two groups. Data were analyzed using Statistical Package for Social Science (SPSS version 18, and descriptive and inferential statistics such as Chi-square test, Kaplan-Meier, Hazard ratio, independent t-test, and Fisher′s exact test. Results: There was a statistically significant difference between the two groups in their catheter survival after 30 h and 10 min ( P < 0.001. Over 60% of the patients (61.1% in the intervention group did not show any sign of phlebitis, while 16.7% and 22.2% of the patients manifested signs of grade 2 and 3 phlebitis, respectively. In the control group, 22.2% of the patients showed no signs of phlebitis, while 5.6%, 27.8%, and 44.4% of the patients exhibited signs of grade 2, 3, and 4 phlebitis, respectively. The statistical analysis showed significant differences in the degree of phlebitis ( P = 0

Drug-induced lupus: simvastatin or amiodarone? A case report in elderly

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Mauro Turrin

2013-03-01

Full Text Available Reports of systemic lupus erythematosus (SLE seen during treatment with amiodarone are rare in the literature. SLE or immunological abnormalities induced by treatment with statins are more frequent. In this issue we report a case of a 81-year-old male who, after a 2-year therapy with amiodarone, developed a clinical and serologic picture of drug-induced SLE (DILE. He was admitted for congestive heart failure in mechanical aortic valve prosthesis, permanent atrial fibrillation (anticoagulation with warfarin, hypercholesterolaemia, and hypothyroidism. Amiodarone was started two years earlier for polymorphic ventricular tachycardia, statin and L-thyroxine the following year. At admission he presented pleuro-pericardical effusion detected by CT-scan (also indicative of interstitial lung involvement and echocardiography. Serological main indicative findings were: elevation of inflammatory markers, ANA (Anti-Nuclear Antibodies titers = 1:320 (indirect immune-fluorescence – IIF – assay on HEp-2, homogeneous/fine speckled pattern, anti-dsDNA titers = 1:80 (IIF on Crithidia luciliae, negative ENA (Extractable Nuclear Antigens and antibodies anti-citrulline, rheumatoid factor = 253 KU/l, normal C3-C4, negative HbsAg and anti-HCV, negative anticardiolipin antibodies IgG and IgM, negative anti-beta2GPI IgG and IgM. Amiodarone was discontinued and methylprednisolone was started, since the patient was severely ill. At discharge, after a month, the patient was better and pleuro-pericardical effusion was reduced. Readmitted few weeks later for bradyarithmia and worsening of dyspnoea, pericardial effusion was further reduced but he died for refractory congestive heart failure and pneumonia. Clinical picture (sierositis, neither skin nor kidney involvement, other typical side effects of amiodarone (hypothyroidism and lung interstitial pathology and serological findings are suggestive of amiodarone-induced SLE.

The effect of the amiodarone-warfarin interaction on anticoagulation quality in a single, high-quality anticoagulation center.

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White, Ryan D; Riggs, Kyle W; Ege, Ed J; Petroski, Gregory F; Koerber, Scott M; Flaker, Greg

2016-03-01

Clinical trials have reported a low time in therapeutic range (TTR) in patients with atrial fibrillation treated with both warfarin andamiodarone. These trials included centers and countries with both high and low TTRs. What is the impact of amiodarone on the TTR in a single, high-quality anticoagulation clinic? TTR was assessed in amiodarone and nonamiodarone-treated patients from a University anticoagulation clinic. Baseline characteristics between patients ever-taking or never-taking amiodarone were similar, except more amiodarone patients were smokers (19.5 vs. 6.1%, P = 0.0031). The TTR calculated from 8901international normalized ratios (INRs) in 249 nonamiodarone patients with a mean follow-up of 34 ± 20 months (mean INR 36 ± 18) was 66 ± 16.6% compared with 61.3 ± 16.2% (P = 0.111) from 1455 INRs in 41 amiodarone-treated patients with a mean follow-up of 28 ± 20 months (mean INR 35 ± 22). Factors associated with a low TTR were male sex (P = 0.0013), smoker (P = 0.0048), and amiodarone use (P = 0.0374). A second on-treatment analysis, in which the TTR was calculated only during amiodarone therapy, resulted in similar findings; however, amiodarone did not emerge as a predictor of a low TTR. In 11 patients, the TTR prior to amiodarone (54.5 ± 22.2%) was not significantly different in the first 3 months (54.6 ± 33.4%) or after 3 months (67.2 ± 33.7%) of amiodarone. In a single high-quality anticoagulation center, anticoagulation quality, as measured by the TTR, can be comparable in amiodarone and nonamiodarone-treated patients.

Pulmonary hypertension and isolated right heart failure complicating amiodarone induced hyperthyroidism.

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Wong, Sean-Man; Tse, Hung-Fat; Siu, Chung-Wah

2012-03-01

Hyperthyroidism is a common side effect encountered in patients prescribed long-term amiodarone therapy for cardiac arrhythmias. We previously studied 354 patients prescribed amiodarone in whom the occurrence of hyperthyroidism was associated with major adverse cardiovascular events including heart failure, myocardial infarction, ventricular arrhythmias, stroke and even death [1]. We now present a case of amiodarone-induced hyperthyroidism complicated by isolated right heart failure and pulmonary hypertension that resolved with treatment of hyperthyroidism. Detailed quantitative echocardiography enables improved understanding of the haemodynamic mechanisms underlying the condition. Copyright © 2011 Australian and New Zealand Society of Cardiac and Thoracic Surgeons (ANZSCTS) and the Cardiac Society of Australia and New Zealand (CSANZ). Published by Elsevier B.V. All rights reserved.

Medical image of the week: acute amiodarone pulmonary toxicity

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Mazursky K

2015-10-01

Full Text Available No abstract available. Article truncated after 150 words. A 71 year old man with a medical history significant for chronic obstructive pulmonary disease, coronary artery disease with post-operative status coronary artery bypass grafting, heart failure with reduced ejection fraction (25% and atrial fibrillation/flutter underwent an elective ablation of the tachyarrhythmia at another facility and was prescribed amiodarone post procedure. He started complaining of cough and dyspnea one day post procedure and was empirically treated with 2 weeks of broad spectrum antibiotics. He subsequently was transferred to our facility due to worsening symptoms. He also complained of nausea, anorexia with resultant weight loss since starting amiodarone, which was stopped 5 days prior to transfer. Infectious work up was negative. On arrival to our facility, he was diagnosed with small sub-segmental pulmonary emboli, pulmonary edema and possible acute amiodarone toxicity. His was profoundly hypoxic requiring high flow nasal cannula or 100% non-rebreather mask at all times. His symptoms persisted despite ...

The incidence of phlebitis with intravenous amiodarone at guideline dose recommendations.

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Slim, Ahmad M; Roth, Jason E; Duffy, Benjamin; Boyd, Sheri Y N; Rubal, Bernard J

2007-12-01

Postoperative atrial fibrillation following cardiothoracic surgery is common and frequently managed with intravenous (IV) amiodarone. Phlebitis is the most common complication with peripheral infusion of this agent. Current practice guidelines for peripheral IV administration of phlebitis. The present study examines the incidence of phlebitis in a postoperative patient population given current dose recommendations. A total of 273 patient charts were reviewed. The incidence of phlebitis in patients given IV amiodarone (n = 36) was 13.9% (95% confidence interval, 2.6-25.2%; p = 0.001). Logistic regression analysis with backward elimination of other therapeutic risk factors suggests that the odds ratio for phlebitis using current dose regimens without IV filters is 19-fold greater than baseline risk in this population. Phlebitis remains a significant complication associated with peripheral infusion of amiodarone within recommended dosing limits.

Acutely Onset Amiodarone-Induced Angioedema in a Patient with New Atrial Fibrillation

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Hossein Vakili

2014-01-01

Full Text Available A 50-year-old man was admitted to our emergency department due to new episode of palpitation. He had history of angioplasty of right coronary artery (RCA with drug eluting stent 2 years ago. His electrocardiogram revealed atrial fibrillation (AF. Intravenous amiodarone 150 mg during 10 minutes and then 1 mg/min infusion were started to achieve rate control and pharmacologic conversion to sinus rhythm. After 60 minutes of starting amiodarone infusion, he developed swelling of the skin around his mouth and eyes, and also mucosa of the mouth, eyes and tongue. To conclude, angioedema should be considered a rare side effect of amiodarone which is used broadly in cardiovascular field.

Amiodarone-induced hyperthyroidism during massive weight loss following gastric bypass.

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Bourron, Olivier; Ciangura, Cécile; Bouillot, Jean-Luc; Massias, Laurent; Poitou, Christine; Oppert, Jean-Michel

2007-11-01

Gastric bypass is increasingly used in morbidly obese patients to achieve significant reduction of body weight and fat mass and concurrent improvement in co-morbidities. We report the case of a 53-year-old male patient (141 kg, BMI 50 kg/m2), successfully treated by amiodarone for supraventricular arrythmia, who underwent Roux-en-Y gastric bypass (RYGBP). 6 months after surgery, he had lost 45% of his preoperative weight (44.8% of weight loss was lean mass) and developed amiodarone-induced subclinical hyperthyroidism. We hypothesize the following sequence of events: weight loss after RYGBP, therefore fat loss, decrease in distribution volume of amiodarone inducing iodine overload and hyperthyroidism, reinforcing weight loss and particularly loss of lean mass. This report emphasizes the importance of careful monitoring of weight and body composition changes after RYGBP. In this situation, checking thyroid status is recommended, especially when there is a history of thyroid disease or potentially toxic thyroid medication.

Medication Use Evaluation of Dronedarone in Comparison to Amiodarone

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Adam Corey

2016-12-01

Full Text Available Amiodarone is the most effective rhythm-control for atrial fibrillation, but produces serious potential side effects. Dronedarone was designed to eliminate amiodarone toxicities, but increased the risk of mortality in clinical trials. This medication use evaluation compares one year of dronedarone use with a matched cohort of amiodarone patients at a single hospital in Greensboro, NC. Forty-eight patients were included with an average age of 71.8 years and 37.5% female population. No significant difference was found for the primary composite outcome of death, myocardial infarction, stroke, and systemic embolism (OR = 2.4, p = 0.148. Likewise, no statistical significance was demonstrated between the two groups for QTc prolongation, hypothyroidism, liver dysfunction or maintenance of normal sinus rhythm. In conclusion, the clinical decision process demonstrated no increased risk of death or other adverse events in the use of dronedarone. Conflict of Interest We declare no conflicts of interest or financial interests that the authors or members of their immediate families have in any product or service discussed in the manuscript, including grants (pending or received, employment, gifts, stock holdings or options, honoraria, consultancies, expert testimony, patents and royalties   Type: Student Project

The effects of sesame oil on the prevention of amiodarone-induced phlebitis.

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Bagheri-Nesami, Masoumeh; Shorofi, Seyed Afshin; Hashemi-Karoie, Seyedeh Zahra; Khalilian, Alireza

2015-01-01

Phlebitis is the most common complication associated with peripheral intravenous infusion of amiodarone. The purpose of this study is to determine the effects of sesame oil on the prevention of amiodarone-induced phlebitis. This is a double-blind randomized controlled trial. Thirty-six patients hospitalized in a coronary care unit were randomly allocated into two groups using a convenience sampling method. Following peripheral intravenous cannulation, five drops of pure sesame oil were applied to the skin within a 10 cm radius of the infusion site prior to the administration of amiodarone in the intervention group. Sesame oil was rubbed on the skin at the infusion site every 6 h in the 24-h period of amiodarone infusion. In the control group, liquid paraffin, used as a placebo, was rubbed on the skin at the infusion site of amiodarone. Both groups were monitored for the development of phlebitis and its degree within the 24-h period of amiodarone infusion as well as 6 h after its administration. The incidence of phlebitis was confirmed and recorded by an assessor who was blind to the two groups. Data were analyzed using Statistical Package for Social Science (SPSS) version 18, and descriptive and inferential statistics such as Chi-square test, Kaplan-Meier, Hazard ratio, independent t-test, and Fisher's exact test. There was a statistically significant difference between the two groups in their catheter survival after 30 h and 10 min (P phlebitis, while 16.7% and 22.2% of the patients manifested signs of grade 2 and 3 phlebitis, respectively. In the control group, 22.2% of the patients showed no signs of phlebitis, while 5.6%, 27.8%, and 44.4% of the patients exhibited signs of grade 2, 3, and 4 phlebitis, respectively. The statistical analysis showed significant differences in the degree of phlebitis (P = 0.006) and the onset of phlebitis development (P phlebitis.

Systematic review of the use of intravenous amiodarone and nifekalant for cardiopulmonary resuscitation in Japan

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Mari Amino, MD, PhD

2014-06-01

Conclusions: Amiodarone and nifekalant were equivalent in their prophylactic and defibrillation efficacy. Concerning the initial amiodarone dose, the 125 mg intravenous [i.v.] over 10 min seemed to be more appropriate for the Japanese population.

Severe congestive heart failure patient on amiodarone presenting with myxedemic coma: a case report.

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Shaheen, Mazen

2009-01-01

This is a case report of myxedema coma secondary to amiodarone-induced hypothyroidism in a patient with severe congestive heart failure (CHF). To our knowledge and after reviewing the literature there is one case report of myxedema coma during long term amiodarone therapy. Myxedema coma is a life threatening condition that carries a mortality reaching as high as 20% with treatment. The condition is treated with intravenous thyroxine (T4) or intravenous tri-iodo-thyronine (T3). Patients with CHF on amiodarone may suffer serious morbidity and mortality from hypothyroidism, and thus may deserve closer follow up for thyroid stimulating hormone (TSH) levels. This case report carries an important clinical application given the frequent usage of amiodarone among CHF patients. The myriad clinical presentation of myxedema coma and its serious morbidity and mortality stresses the need to suspect this clinical syndrome among CHF patients presenting with hypotension, weakness or other unexplained symptoms.

Amiodarone-Induced Liver Injury and Cirrhosis.

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Buggey, Jonathan; Kappus, Matthew; Lagoo, Anand S; Brady, Carla W

2015-01-01

We present a case report of an 80-year-old woman with volume overload thought initially to be secondary to heart failure, but determined to be amiodarone-induced acute and chronic liver injury leading to submassive necrosis and bridging fibrosis consistent with early cirrhosis. Her histopathology was uniquely absent of steatosis and phospholipidosis, which are commonly seen in AIC.

Effect of amiodarone-induced hyperthyroidism on left ventricular outflow obstruction after septal myectomy for hypertrophic cardiomyopathy.

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Pokorney, Sean D; Stone, Neil J; Passman, Rod; Oyer, David; Rigolin, Vera H; Bonow, Robert O

2010-12-01

Patients with obstructive hypertrophic cardiomyopathy who undergo septal myectomy are at risk for developing postoperative atrial fibrillation. Amiodarone is effective in treating this arrhythmia but is associated with multiple adverse effects, often with delayed onset. A novel case is described of a patient who developed type 2 amiodarone-induced hyperthyroidism that presented as recurrence of outflow obstruction after septal myectomy. The patient's symptoms and echocardiographic findings of outflow obstruction resolved substantially with the treatment of the amiodarone-induced hyperthyroidism. Amiodarone-induced hyperthyroidism of delayed onset can be a subtle diagnosis, requiring a high index of suspicion. In conclusion, recognition of this diagnosis in patients with recurrence of outflow obstruction by symptoms and cardiac imaging after septal myectomy may avoid unnecessary repeat surgical intervention. Copyright © 2010 Elsevier Inc. All rights reserved.

In Vitro Study on effects of Amiodarone and Ketoconazole on Leishmania infantum

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Mohammad Hosein Razi Jalali

2014-09-01

Full Text Available Background & objectives: The leishmaniases are considered among the major infectious diseases affecting public health in several regions. There are many chemical agents which are effective in treatment of visceral leishmaniasis. But, overall treatment of visceral leishmaniasis is often difficult. Thus, identification of new chemotherapeutic agents is important for treatment of disease. Since targeting of the ergosterol synthesis pathway of Leishmania may be useful therapeutically, the aim of this study was to investigate the effect of alone or in combination of amiodarone and ketoconazole on Leishmania infantum.   Methods : To obtain logarithmic promastigotes of L. infantum, the parasites were cultured in BHI medium with FCS 10% together with antibiotics of penicillin and streptomycin and incubated at 24° C. Amastigote forms were obtained in BHI medium supplemented with 20% FCS at pH of 5.5 which incubated in 37° C. L.infantum susceptibility to amiodarone and ketoconazole was evaluated by proliferation of parasites in the absence or presence of these drugs with MTT assay. For evaluation of antiproliferative synergism against promastigotes and axenic amastigotes, fractional inhibitory concentrations (FIC were calculated. An isobologram curve was constructed too.   Results: Amiodarone produced a marked reduction in the viability of L.infantum promastigotes and axenic amastigotes. On the other hand ketoconazole induced a dose dependent effect on the parasites proliferation for promastigotes and axenic amastigotes. When the drugs were used in combination, the results indicated clear synergistic as shown by a concave isobologram and FIC value.   Conclusion: The present study represents the evidence that the combination of amiodarone plus ketoconazole acts synergistically in controlling L. infantume in vitro. It is possible that amiodarone could be used in combination with ketoconazole to combat infection at low doses, thus reducing its side

Magnesium sulphate and amiodarone prophylaxis for prevention of postoperative arrhythmia in coronary by-pass operations

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Huysal Kagan

2009-02-01

Full Text Available Abstract Background The aim of this study was to investigate the use of prophylactic magnesium sulphate and amiodarone in treating arrhythmias that may occur following coronary bypass grafting operations. Methods The study population consisted of 192 consecutive patients who were undergoing coronary artery bypass grafting (CABG. Sixty-four patients were given 3 g of magnesium sulphate (MgSO4 [20 ml = 24.32 mEq/L Mg+2] in 100 cc of isotonic 0.9% solution over 2 hours intravenously at the following times: 12 hours prior to the operation, immediately following the operation, and on postoperative days 1, 2, and 3 (Group 1. Another group of 64 patients was given a preoperative infusion of amiodarone (1200 mg on first post-operative day (Group 2. After the operation amiodarone was administered orally at a dose of 600 mg/day. Sixty-four patients in group 3 (control group had 100 cc. isotonic 0.9% as placebo, during the same time periods. Results In the postoperative period, the magnesium values were significantly higher in Group 1 than in Group 2 for all measurements. The use of amiodarone for total arrhythmia was significantly more effective than prophylactic treatment with magnesium sulphate (p = 0.015. There was no difference between the two drugs in preventing supraventricular arrhythmia, although amiodarone significantly delayed the revealing time of atrial fibrillation (p = 0.026. Ventricular arrhythmia, in the form of ventricular extra systole, was more common in the magnesium prophylaxis group. The two groups showed no significant differences in other operative or postoperative measurements. No side effects of the drugs were observed. Conclusion Prophylactic use of magnesium sulphate and amiodarone are both effective at preventing arrhythmia that may occur following coronary by-pass operations. Magnesium sulphate should be used in prophylactic treatment since it may decrease arrhythmia at low doses. If arrhythmia should occur despite this

Treatment of amiodarone-induced hypothyroidism with potassium perchlorate

NARCIS (Netherlands)

van Dam, E. W.; Prummel, M. F.; Wiersinga, W. M.; Nikkels, R. E.

1993-01-01

The antiarrhythmic drug, amiodarone, induces thyroid dysfunction, which is potentially dangerous in cardiac patients. After discontinuation of the drug it takes several months before euthyroidism is restored. The potent antithyroid drug, potassium perchlorate (KClO4), is used successfully to treat

Sternal Route More Effective than Tibial Route for Intraosseous Amiodarone Administration in a Swine Model of Ventricular Fibrillation.

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Burgert, James M; Martinez, Andre; O'Sullivan, Mara; Blouin, Dawn; Long, Audrey; Johnson, Arthur D

2018-01-01

The pharmacokinetics of IO administered lipid soluble amiodarone during ventricular fibrillation (VF) with ongoing CPR are unknown. This study measured mean plasma concentration over 5 minutes, maximum plasma concentration (Cmax), and time to maximum concentration (Tmax) of amiodarone administered by the sternal IO (SIO), tibial IO (TIO), and IV routes in a swine model of VF with ongoing CPR. Twenty-one Yorkshire-cross swine were randomly assigned to three groups: SIO, TIO, and IV. Ventricular fibrillation was induced under general anesthesia. After 4 minutes in VF, 300 mg amiodarone was administered as indicated by group assignment. Serial blood specimens collected at 30, 60, 90, 120, 150, 180, 240, and 300 seconds were analyzed using high performance liquid chromatography with tandem mass spectrometry. The mean plasma concentration of IV amiodarone over 5 minutes was significantly higher than the TIO group at 60 seconds (P = 0.02) and 90 seconds (P = 0.017) post-injection. No significant differences in Cmax between the groups were found (P <0.05). The Tmax of amiodarone was significantly shorter in the SIO (99 secs) and IV (86 secs) groups compared to the TIO group (215 secs); P = 0.002 and P = 0.002, respectively. The SIO and IV routes of amiodarone administration were comparable. The TIO group took nearly three times longer to reach Tmax than the SIO and IV groups, likely indicating depot of lipid-soluble amiodarone in adipose-rich tibial yellow bone marrow. The SIO route was more effective than the TIO route for amiodarone delivery in a swine model of VF with ongoing CPR. Further investigations are necessary to determine if the kinetic differences found between the SIO and TIO routes in this study affect survival of VF in humans.

[Successful treatment of fetal supraventricular tachycardia with a combination of digoxin and amiodarone].

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Hajdú, J; Szabó, I; Német, J

1996-10-06

The supraventricular tachycardia is a life threatening state in the intrauterine life. It can cause non-immune hydrops fetalis, intrauterine death or complications during the delivery. The unexplained tachycardia can cause fetal distress and premature delivery. Usually the digoxin is the first drug of choice for transplacental cardioversion. If digitalisation does not achieve cardioversion, the second line antiarrhythmic drugs should be instituted. Amiodarone has been suggested as a therapeutic alternative after failure of digoxin-verapamil combination. We give a drug in standard therapeutic doses for four-five days and after it we determine whether it is effective or not. We should determine the newer therapy or termination of pregnancy. The transplacental administration of amiodarone may be dangerous because of fetal cretinism. Our case is the first in Hungary-in our best knowledge- and we suggest the amiodarone for transplacental therapy.

Comparison of amiodarone vs magnesium sulphate in the prevention of atrial fibrillation after coronary artery bypass grafting surgery

International Nuclear Information System (INIS)

Bashir, I.; Abbas, S.

2013-01-01

Background: Atrial fibrillation (AF) is common in patients after coronary artery bypass grafting (CABG) and can result in increased morbidity and mortality, increased length of hospital stay, and increased cost. In this study we compared the efficacy of amiodarone versus magnesium sulphate in the prophylaxis of post-CABG atrial fibrillation. Objective: This study was carried out to assess the efficacy of amiodarone in comparison to magnesium sulphate in the prevention of atrial fibrillation after coronary artery bypass grafting. Study Design: Randomized controlled trials. Place and duration of study: The study was carried out at Armed Forces Institute of Cardiology and National Institute of Heart Diseases, Rawalpindi from July 2010 to December 2011 Patients and Methods: Total 240 patients were included in the study and randomly divided in two groups of 120 each using random number table. Patients in Group A (Amiodarone group) were given a loading dose of amiodarone 5 mg/Kg after induction of anesthesia which was then continued as infusion at 5 micro gm/Kg/minute on first postoperative day. This was followed by an oral dose of 600 mg/day postoperatively for 5 days. Those in Group B (Magnesium Sulphate group) received 2 g of magnesium sulphate in 100 ml of isotonic 0.9% solution intravenously over 1 hour at following times: preoperatively, immediately following the operation, and on postoperative days 1, 2, and 3. Results: Thirteen patients (10.8%) developed AF in Amiodarone group, compared to 31 patients (25.8%) in magnesium sulphate group. The results proved amiodarone to be more effective than magnesium sulphate in preventing post-CABG AF (p<0.001). Thirty one patients who developed AF postoperatively in the magnesium group were treated with amiodarone, and all patients recovered normal sinus rhythm. In the amiodarone prophylaxis group, 9 patients regained sinus rhythm in 6 - 8 hours, while for 4 remaining patients cardioversion was attempted out of which 2

2018 European Thyroid Association (ETA) Guidelines for the Management of Amiodarone-Associated Thyroid Dysfunction.

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Bartalena, Luigi; Bogazzi, Fausto; Chiovato, Luca; Hubalewska-Dydejczyk, Alicja; Links, Thera P; Vanderpump, Mark

2018-03-01

Treatment with amiodarone is associated with changes in thyroid function tests, but also with thyroid dysfunction (amiodarone-induced hypothyroidism, AIH, and amiodarone-induced thyrotoxicosis, AIT). Both AIH and AIT may develop in apparently normal thyroid glands or in the presence of underlying thyroid abnormalities. AIH does not require amiodarone withdrawal, and is treated with levothyroxine replacement if overt, whereas subclinical forms may be followed without treatment. Two main types of AIT are recognized: type 1 AIT (AIT 1), a form of iodine-induced hyperthyroidism occurring in nodular goitres or latent Graves disease, and type 2 AIT (AIT 2), resulting from destructive thyroiditis in a normal thyroid gland. Mixed/indefinite forms exist due to both pathogenic mechanisms. AIT 1 is best treated with thionamides that may be combined for a few weeks with sodium perchlorate to make the thyroid gland more sensitive to thionamides. AIT 2 is treated with oral glucocorticoids. Once euthyroidism has been restored, AIT 2 patients are followed up without treatment, whereas AIT 1 patients should be treated with thyroidectomy or radioiodine. Mixed/indefinite forms of AIT are treated with thionamides. Oral glucocorticoids can be added from the beginning if a precise diagnosis is uncertain, or after a few weeks if response to thionamides alone is poor. The decision to continue or to stop amiodarone in AIT should be individualized in relation to cardiovascular risk stratification and taken jointly by specialist cardiologists and endocrinologists. In the presence of rapidly deteriorating cardiac conditions, emergency thyroidectomy may be required for all forms of AIT.

Variable use of amiodarone is associated with a greater risk of recurrence of atrial fibrillation in the critically ill.

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Mitrić, Goran; Udy, Andrew; Bandeshe, Hiran; Clement, Pierre; Boots, Rob

2016-04-02

Atrial fibrillation is a common rhythm disturbance in the general medical-surgical intensive care unit. Amiodarone is a popular drug in this setting but evidence to inform clinical practice remains scarce. We aimed to identify whether variation in the clinical use of amiodarone was associated with recurrent atrial fibrillation. This was a retrospective audit of 177 critically ill patients who developed new-onset atrial fibrillation after admission to a tertiary level medical-surgical trauma intensive care unit. Patterns of amiodarone prescription (including dosage schedule and duration) were assessed in relation to recurrence of atrial fibrillation during the intensive care unit stay. Known recurrence risk factors, such as inotrope administration, cardiac disease indices, Charlson Comorbidity Index, magnesium concentrations, fluid balance, and potassium concentrations, were also included in adjusted analysis using forward stepwise logistic regression modelling. The cohort had a median (interquartile range) age of 69 years (60-75), Acute Physiology and Chronic Health Evalution II score of 22 (17-28) and Charlson Comorbidity Index of 2 (1-4). A bolus dose of amiodarone followed by infusion (P = 0.02), in addition to continuing amiodarone infusion through to discharge from the intensive care unit (P < 0.001), were associated with less recurrent dysrhythmia. Recurrence after successful treatment was associated with ceasing amiodarone while an inotrope infusion continued (P < 0.001), and was more common in patients with a prior history of congestive cardiac failure (P = 0.04), and a diagnosis of systemic inflammatory response syndrome (P = 0.02). Amiodarone should be administered as a bolus dose followed immediately with an infusion when treating atrial fibrillation in the medical-surgical intensive care unit. Consideration should be given to continuing amiodarone infusions in patients on inotropes until they are ceased.

Effect of single intraoperative dose of amiodarone in patients with rheumatic valvular heart disease and atrial fibrillation undergoing valve replacement surgery

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Selvaraj Thiruvenkadam

2009-01-01

Full Text Available Maintenance of sinus rhythm (SR is superior to rate control in atrial fibrillation (AF. In order to achieve SR, we administered single-dose intravenous amiodarone intraoperatively and evaluated its effect on conversion of rheumatic AF to SR in patients undergoing valvular heart surgery. Patients were randomly assigned to amiodarone ( n = 42 or control ( n = 40 group in a double blind manner. The amiodarone group received amiodarone (3 mg/kg intravenously prior to the institution of cardiopulmonary bypass and the control group received the same volume of normal saline. In the amiodarone group, the initial rhythm after the release of aortic cross clamp was noted to be AF in 14.3% ( n = 6 and remained so in 9.5% ( n = 4 of patients till the end of surgery. In the control group, the rhythm soon after the release of aortic cross clamp was AF in 37.5% ( n = 15 ( p = 0.035 and remained so in 32.5% ( n = 13 of patients till the end of surgery ( p = 0.01. At the end of first post-operative day 21.4% ( n = 9 of patients in amiodarone group and 55% ( n = 22 of patients in control group were in AF ( p = 0.002. The requirement of cardioversion/defibrillation was 1.5 (±0.54 in amiodarone group and 2.26 (±0.73 in the control group ( p = 0.014, and the energy needed was 22.5 (±8.86 joules in the amiodarone group and 40.53 (±16.5 in the control group ( p = 0.008. A single intraoperative dose of intravenous amiodarone increased the conversion rate of AF to normal sinus rhythm, reduced the need and energy required for cardioversion/defibrillation and reduced the recurrence of AF within one day.

Effect of antiarrhythmic therapy with intravenous loading dose of amiodarone: evidence for an altered response in diabetic patients.

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Iervasi, G; Clerico, A; Bonini, R; Nannipieri, M; Manfredi, C; Sabatino, L; Biagini, A; Donato, L

1998-01-01

Amiodarone, a potent class III antiarrhythmic agent with adrenergic antagonism properties, is administered increasingly to diabetic patients with cardiac arrhythmias refractory to all other available forms of therapy. Because a large percentage of diabetic patients show a perturbed autonomic regulation of the cardiovascular system, including a pertubed regulation of heart rate, we studied the antiarrhythmic response as well as the early effects (within 5 days) on heart rate of an intravenous amiodarone loading dose in diabetic patients. Seven type II (noninsulin-dependent) diabetic patients (age 64.7 +/- 9.7 years), affected by uncontrolled atrial fibrilation or atrial flutter, were enrolled for the study and a group of 12 well-matched (for age, sex and arrhythmia) nondiabetic patients served as a control group. It was found that before amiodarone administration, nondiabetic patients showed significantly wider variations in the circadian rhythm of heart rate values than diabetic patients (p = 0.0062, unpaired t-test). In all patients but one (who was nondiabetic), amiodarone treatment resulted in a cardioversion to sinus rhythm. After amiodarone administration, nondiabetic patients showed a significantly greater decrease (p = 0.0011) in heart rate values in comparison with the diabetic group (-35% vs. -20% on average, at the end of the study). Furthermore, in nondiabetic patients there was also an earlier significant fall (within the first 4 h after the start of treatment with amiodarone, p atrial fibrilation or atrial flutter may be delayed in comparison with nondiabetic patients. This altered response may be (at least in part) due to the diabetic autonomic neuropathy. Our study indicates that the presence of diabetes mellitus always must be taken into account when patients are enrolled for large, prospective, randomized trials, planned to evaluate the antiarrhythmic effects of amiodarone given intravenously.

Amiodarone is a cost-neutral way of preventing atrial fibrillation after surgery for lung cancer

DEFF Research Database (Denmark)

Riber, Lars P.; Christensen, Thomas D.; Pilegaard, Hans K.

2014-01-01

OBJECTIVES: Our aim was to estimate the costs and health benefits of routinely administered postoperative amiodarone as a prophylactic agent in reducing the risk of atrial fibrillation in patients undergoing surgery for lung cancer. METHODS: This was a cost-effectiveness study, based.......23). There were no signs of adverse developments referable to amiodarone in this prophylactic regime. CONCLUSIONS: For patients undergoing surgery for lung cancer, routine use of postoperative prophylactic intravenous bolus and five subsequent days of oral amiodarone therapy reduces the risk of atrial...... on the randomized, controlled, double-blinded PASCART study, using avoidance of atrial fibrillation as the measure of benefit. Two hundred and fifty-four eligible, consecutively enrolled patients, undergoing surgery for lung cancer at the department of Cardiothoracic and Vascular Surgery, Aarhus University Hospital...

Role of Pre-incision, Intravenous Prophylactic Amiodarone to Control Arrhythmias in Patients with Rheumatic Valvular Heart Disease undergoing Mitral Valve Replacement

International Nuclear Information System (INIS)

Ahmad, K.; Naqvi, S.

2013-01-01

Objective: To evaluate the effect of intra-operative single intra venous dose of amiodarone on post operative cardiac arrhythmias in patients undergoing valvular heart surgery. Study Design: Randomized controlled trials. Place and Duration of surgery: This study was performed at Armed forces Institute of Cardiology Rawalpindi from Jan 01, 2011 to Dec 31, 2011. Patients and Methods: In this study 80 patients with rheumatic valvular heart disease and undergoing elective mitral valve replacement were randomly divided into two groups. Group I, n = 40 (Amiodarone group) was given single intravenous dose of amiodarone (5 mg/kg in 100 ml of saline over 30 min) before sternotomy incision. Group II, n = 40(control / placebo group) was given 100 ml of saline over 30 min. Result: In the amiodarone group, after removal of aortic cross clamp 75% patients had sinus rhythm compared to 47.5% in control group. p=0.045. Similarly 15% had AF, 5% JR and 5% VT/VF in amiodarone group in contrast to 32.5% with AF, 12.5% JR and 7.5% Vt/VF in control group. (p=0.045). Response to cardioversion was positive in 75% of the patients requiring shocks in amiodarone group as against 43.75% in the control group. (p=0.044). Conclusion: A single intravenous bolus dose of amiodarone is effective in decreasing the incidence of cardiac arrhythmias after mitral valve replacement in patients with rheumatic MVD. (author)

A randomized active-controlled study comparing the efficacy and safety of vernakalant to amiodarone in recent-onset atrial fibrillation

DEFF Research Database (Denmark)

Camm, A John; Capucci, Alessandro; Hohnloser, Stefan H

2011-01-01

with 32.8% of amiodarone patients; p = 0.0012). Serious adverse events or events leading to discontinuation of study drug were uncommon. There were no cases of torsades de pointes, ventricular fibrillation, or polymorphic or sustained ventricular tachycardia. Conclusions Vernakalant demonstrated efficacy...... amiodarone infusion, or a 60-min infusion of amiodarone (5 mg/kg) followed by a maintenance infusion (50 mg) over an additional 60 min, plus a sham vernakalant infusion. Results Conversion from AF to sinus rhythm within the first 90 min (primary end point) was achieved in 60 of 116 (51.7%) vernakalant...

Amiodarone and Catheter Ablation as Cardiac Resynchronization Therapy for Children with Dilated Cardiomyopathy and Wolff-Parkinson-White Syndrome

Science.gov (United States)

Kim, Sung Hoon; Jeong, Soo In; Kang, I-Seok; Lee, Heung Jae

2013-01-01

Preexcitation by accessory pathways (APs) is known to cause dyssynchrony of the ventricle, related to ventricular dysfunction. Correction of ventricular dyssynchrony can improve heart failure in cases of dilated cardiomyopathy (DCMP) with preexcitation. Here, we report the first case of a child with DCMP and Wolff-Parkinson-White (WPW) syndrome treated with amiodarone and radiofrequency catheter ablation (RFCA) in Korea. A 7-year-old boy, who suffered from DCMP and WPW syndrome, showed improved left ventricular function and clinical functional class after treatment with amiodarone to eliminate preexcitation. QRS duration and left ventricular ejection fraction (LVEF) were inversely correlated with amiodarone dosage. After confirming the reduction of preexcitation effects in DCMP, successful RFCA of the right anterior AP resulted in LVEF improvement, along with the disappearance of preexcitation. Our findings suggest that ventricular dyssynchrony, caused by preexcitation in DCMP with WPW syndrome, can worsen ventricular function and amiodarone, as well as RFCA, which should be considered as a treatment option, even in young children. PMID:23407697

Efficacy and safety of oral amiodarone in controlling heart rate in patients with persistent atrial fibrillation who have undergone digitalisation.

Science.gov (United States)

Kochiadakis, George E; Kanoupakis, Emmanuel M; Igoumenidis, Nikolaos E; Mavrakis, Hercules E; Kafarakis, Panagiotis K; Vardas, Panos E

2005-01-01

Oral amiodarone has been suggested by some authors for rate control in patients with persistent atrial fibrillation. In this study we evaluated the efficacy and safety of oral amiodarone versus placebo for rate control during exercise and daily activities in patients with chronic atrial fibrillation who had undergone digitalisation. The study group consisted of 53 patients (35 men, mean age 65 +/- 9 years) with persistent atrial fibrillation (mean duration 17 +/- 7 months). All patients had therapeutic levels of digitalis and were under anticoagulation treatment with acenocoumarol. Twenty-eight of them were treated with amiodarone (200 mg per day orally) and 25 received placebo. All patients were assessed with 24-hour ECG monitoring, a maximal symptom-limited cardiopulmonary exercise test and evaluation of adverse events. The mean exercise duration was similar in both groups. Amiodarone produced a lower heart rate than placebo at all exercise levels (p<0.0001 for all). VO2 was similar in both groups whereas O2 pulse was higher in the amiodarone group at all exercise levels. During daily life, heart rate showed a significant circadian pattern in both groups, with higher values during the day than at night (time effect for both p<0.001). The mean value of heart rate under amiodarone was lower than for placebo (75 +/- 10 vs. 86 +/- 12/min, p<0.001) but this difference was due to a significant difference during the day (p<0.001) that was not present during the night (p =0.48). Oral amiodarone is very effective when combined with digoxin for control of heart rate in patients with chronic atrial fibrillation and it should be considered as an alternative treatment when more traditional drugs, such as Ca(+2) inhibitors or b-blockers have proven ineffective or are contraindicated.

Effects of injection-site splinting on the incidence of phlebitis in patients taking peripherally infused amiodarone: A randomized clinical trial.

Science.gov (United States)

Ayat-Isfahani, Farah; Pashang, Mina; Davoudi, Bita; Sadeghian, Saeed; Jalali, Arash

2017-03-01

Intravenous amiodarone is considered an effective treatment option for cardiac ventricular and atrial arrhythmias. Peripheral infusion of amiodarone may cause blood vessels irritation and phlebitis that is the most common complication of this drug by this route even when it is administered within recommended dosing limits. The effect of injection-site splinting on the occurrence of phlebitis among a group of cardiac arrhythmia patients receiving peripherally infused amiodarone. This research is a clinical trial on patients of Tehran Heart Center who were hospitalized due to cardiac arrhythmias. A sample of 60 patients with mean age 65 ± 14 years were randomly divided into control and test groups. In the experimental group with close splint and restrict the movement of the injection site until the end of the infusion and control groups without closing brace, at the same time received amiodarone. Injection protocol was similar for both groups. The results were analyzed with Spss18. The results of this research still significantly reduced the incidence of amiodarone injection-site phlebitis in the injection time (P = .005). Copyright © 2016 Society for Vascular Nursing, Inc. Published by Elsevier Inc. All rights reserved.

Efficacy of amiodarone and lidocaine for preventing ventricular fibrillation after aortic cross-clamp release in open heart surgery: a meta-analysis of randomized controlled trials.

Science.gov (United States)

Zheng, Yong; Gu, Qiang; Chen, Hong-Wu; Peng, Huai-Ming; Jia, Dong-Yu; Zhou, Yu; Xiang, Mei-Xiang

The relative preventative efficacy of amiodarone and lidocaine for ventricular fibrillation (VF) after release of an aortic cross-clamp (ACC) during open heart surgery has not been determined. This meta-analysis was designed to systematically evaluate the influence of amiodarone, lidocaine, or placebo on the incidence of VF after ACC. Prospective randomized controlled trials (RCTs) that compared the VF-preventative effects of amiodarone with lidocaine, or amiodarone or lidocaine with placebo were included. PubMed, EMBASE, and the Cochrane Library were searched for relevant RCTs. Fixed or randomized effect models were applied according to the heterogeneity of the data from the selected studies. We included eight RCTs in the analysis. Pooled results suggested that the preventative effects of amiodarone and lidocaine were comparable (relative risk (RR)=1.12, 95% confidence interval (CI): 0.70 to 1.80, P=0.63), but both were superior to the placebo (amiodarone, RR=0.71, 95% CI: 0.51 to 1.00, P=0.05; lidocaine, RR=0.63, 95% CI: 0.46 to 0.88, P=0.006). The percentage of patients requiring electric defibrillation counter shocks (DCSs) did not differ significantly among patients administered amiodarone (RR=0.21, 95% CI: 0.04 to 1.19, P=0.08), lidocaine (RR=2.44, 95% CI: 0.13 to 44.02, P=0.55), or the placebo (RR=0.56, 95% CI: 0.25 to 1.25, P=0.16). Amiodarone and lidocaine are comparably effective in preventing VF after ACC, but the percentage of patients who subsequently require DCSs does not differ among those administered amiodarone, lidocaine, or placebo.

Pharmacokinetics and pharmacodynamic effects of amiodarone in plasma of ponies after single intravenous administration

International Nuclear Information System (INIS)

Trachsel, D.; Tschudi, P.; Portier, C.J.; Kuhn, M.; Thormann, W.; Scholtysik, G.; Mevissen, M.

2004-01-01

Atrial fibrillation is a well-known heart disease in horses. The common therapy consists of administration of quinidine. More potent antiarrhythmic drugs have become available for human therapy and the use of these as alternatives to quinidine for equine antiarrhythmic therapy is a matter of interest. Amiodarone (AMD) is used in human medicine for treatment of many arrhythmias, including atrial fibrillation. Its disposition in horses has not yet been investigated. The purpose of this study was to measure the effect of single intravenous doses of amiodarone (5 and 7 mg/kg) on the surface electrocardiogram (ECG) of healthy minishetland ponies during the first 2 days after drug administration and to calculate pharmacokinetic parameters with a physiologically based pharmacokinetic model (PBPK) using amiodarone and desethylamiodarone (DAMD) plasma levels that were determined by high-performance liquid chromatography (HPLC). As expected for a K + -channel-blocker, the main effect on the measured ECG could be seen on the ventricular complex, as the QT interval and the T wave showed statistically significant alterations. The doses investigated were well tolerated clinically. Results from the pharmacokinetic model were found to compare well with literature data of rats, dogs, and humans. It showed a rapid distribution in the tissue, beginning with the rapidly perfused tissue, like the heart, followed by slowly perfused tissues, and finally an accumulation in fat. The half-life for total elimination was calculated to be 16.3 days with 99% eliminated by 97 days. The model predicts that approximately 96% of amiodarone is eliminated as desethylamiodarone in urine, 2% eliminated as desethylamiodarone in bile, and 2% as other metabolites

Amiodarone use after acute myocardial infarction complicated by heart failure and/or left ventricular dysfunction may be associated with excess mortality

DEFF Research Database (Denmark)

Thomas, Kevin L; Al-Khatib, Sana M; Lokhnygina, Yuliya

2008-01-01

, a randomized comparison of valsartan, captopril, or both in patients with acute myocardial infarction with HF and/or left ventricular systolic dysfunction. We compared baseline characteristics of 825 patients treated with amiodarone at randomization with 13,875 patients not treated with amiodarone. Using Cox...

Amiodarona y disfunción tiroidea Amiodarone and thyroid dysfunction

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Leonardo F. L. Rizzo

2012-02-01

Full Text Available La amiodarona es un análogo estructural de la hormona tiroidea, y algunas de sus propiedades antiarrítmicas como así también su toxicidad son atribuibles a su interacción con los receptores nucleares de las hormonas tiroideas. Por ser muy lipofílica, la amiodarona se concentra en muchos tejidos y se elimina, por consecuencia, muy lentamente. Se emplea preferentemente para el tratamiento de arritmias graves tales como fibrilación y taquicardia ventriculares. Otras indicaciones incluyen la fibrilación auricular y el aleteo, la insuficiencia cardíaca congestiva grave, la prevención de la fibrilación auricular recurrente y situaciones de emergencia médica como la prevención de muerte súbita cardiaca¹. Nuestro objetivo es proporcionar un enfoque actualizado sobre la amiodarona y su influencia sobre la fisiología tiroidea y discutir y analizar en profundidad sus potenciales efectos adversos como el hipotiroidismo y la tirotoxicosis.Amiodarone is a structural analogue of thyroid hormone, and some of its anti-arrhythmic actions and toxicity are attributable to its interaction with nuclear receptors of thyroid hormones. Being highly lipophilic, amiodarone is concentrated in many tissues and is eliminated, consequently, very slowly. It is preferably employed to manage life-threatening arrhythmias, including ventricular fibrillation and unstable ventricular tachycardia. Other indications include atrial fibrillation and flutter, severe congestive heart failure, prevention of atrial fibrillation recurrence, and even in emergency medical situations to prevent sudden cardiac death. The aim of this review is to provide an updated approach on amiodarone and its influence on thyroid physiology and to discuss and analyze in depth its potential and not infrequent thyroidal adverse effects such as hypothyroidism and thyrotoxicosis.

Cholinergic Manipulations Bidirectionally Regulate Object Memory Destabilization

Science.gov (United States)

Stiver, Mikaela L.; Jacklin, Derek L.; Mitchnick, Krista A.; Vicic, Nevena; Carlin, Justine; O'Hara, Matthew; Winters, Boyer D.

2015-01-01

Consolidated memories can become destabilized and open to modification upon retrieval. Destabilization is most reliably prompted when novel information is present during memory reactivation. We hypothesized that the neurotransmitter acetylcholine (ACh) plays an important role in novelty-induced memory destabilization because of its established…

Randomised trial of low-dose amiodarone in severe congestive heart failure. Grupo de Estudio de la Sobrevida en la Insuficiencia Cardiaca en Argentina (GESICA)

Science.gov (United States)

Doval, H C; Nul, D R; Grancelli, H O; Perrone, S V; Bortman, G R; Curiel, R

1994-08-20

In severe heart failure many deaths are sudden and are presumed to be due to ventricular arrhythmias. The GESICA trial evaluated the effect of low-dose amiodarone on two-year mortality in patients with severe heart failure. Our prospective multicentre trial included 516 patients on optimal standard treatment for heart failure. Patients were randomised to 300 mg/day amiodarone (260) or to standard treatment (256). Intention-to-treat analysis showed 87 deaths in the amiodarone group (33.5%) compared with 106 in the control group (41.4%) (risk reduction 28%; 95% CI 4%-45%; log rank test p = 0.024). There were reductions in both sudden death (risk reduction 27%; p = 0.16) and death due to progressive heart failure (risk reduction 23%; p = 0.16). Fewer patients in the amiodarone group died or were admitted to hospital due to worsening heart failure (119 versus 149 in the control group; risk reduction 31%; 95% CI 13-46%; p = 0.0024). The decrease in mortality and hospital admission was present in all subgroups examined and independent of the presence of non-sustained ventricular tachycardia. Side-effects were reported in 17 patients (6.1%); amiodarone was withdrawn in 12. Low-dose amiodarone proved to be an effective and reliable treatment, reducing mortality and hospital admission in patients with severe heart failure independently of the presence of complex ventricular arrhythmias.

Interaction of rivaroxaban with amiodarone, verapamil and diltiazem in patients with atrial fibrillation: terra incognita

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S. N. Bel'diev

2016-01-01

Full Text Available Currently there are no generally accepted guidelines for the use of rivaroxaban together with amiodarone, verapamil or diltiazem in patients with creatinine clearance (CrCl<80 ml/min. Some researchers suggest that in renal failure amiodarone, verapamil and diltiazem contribute to a significant increase in plasma concentrations of rivaroxaban that is accompanied by increased risk of bleeding. According to preliminary calculations, it seems rational to reduce the dose of rivaroxaban when co-administered with these drugs: to 15 mg/day in patients with ClCr 50-79 ml/min and to 10 mg/day in patients with ClCr<50 ml/min.

Dysfunction of the thyroid gland during amiodarone therapy: a study of 297 cases

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Czarnywojtek A

2016-04-01

Full Text Available Agata Czarnywojtek,1,2,* Maria Teresa Płazińska,3,* Małgorzata Zgorzalewicz-Stachowiak,4 Kosma Woliński,1 Adam Stangierski,1 Izabela Miechowicz,5 Joanna Waligórska-Stachura,1 Paweł Gut,1 Leszek Królicki,3 Maja Zioncheck,6 Marek Ruchała1 1Department of Endocrinology, Metabolism and Internal Medicine, 2Department of Pharmacology, Poznan University of Medical Sciences, Poznan, 3Nuclear Medicine Department, Medical University of Warsaw, Warsaw, 4Department of Health Prophylaxis, Laboratory of Medical Electrodiagnostics, 5Department of Computer Science and Statistics, 6Poznan University of Medical Sciences, Poznan, Poland *These authors contributed equally to this work Aim: This study aims to explore and compare the efficacy of radioiodine treatment (RIT in hyperthyroid and euthyroid patients who have been treated with amiodarone (AM in the past or are currently undergoing AM treatment. Clinical observation of a group of patients with amiodarone-induced hypothyroidism during a 12-month follow-up period was used for comparison.Design: This was a observational, two-centered study. Patients were assessed at baseline and at 2 months, 6 months, 8 months, and 12 months after RIT.Patients: Group A: At baseline (61 males [M] and 17 females [F], mean age 50±19 years, there were 78 euthyroid patients with cardiac arrhythmias, who were treated with AM and developed amiodarone-induced thyrotoxicosis, and currently require retreatment with AM. Group B: Hyperthyroid patients (92 M and 26 F, mean age 72±11.8 years after AM therapy in the past. Group C: Hyperthyroid patients (66 M and 13 F, mean age 63.9±13.2 years currently treated by AM. Group D: Hypothyroid patients (6 M and 16 F, mean age 61.4±10.4 years after AM therapy. The patients from Groups A, B, and C were retreated with AM after ~3–6 weeks of RIT.Results: In Group A, after 12 months of RIT therapy, recurrent thyrotoxicosis was observed in six (7.7% cases, and persistent

Long-term use of amiodarone before heart transplantation significantly reduces early post-transplant atrial fibrillation and is not associated with increased mortality after heart transplantation

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Rivinius R

2016-02-01

Full Text Available Rasmus Rivinius,1 Matthias Helmschrott,1 Arjang Ruhparwar,2 Bastian Schmack,2 Christian Erbel,1 Christian A Gleissner,1 Mohammadreza Akhavanpoor,1 Lutz Frankenstein,1 Fabrice F Darche,1 Patrick A Schweizer,1 Dierk Thomas,1 Philipp Ehlermann,1 Tom Bruckner,3 Hugo A Katus,1 Andreas O Doesch1 1Department of Cardiology, Angiology and Pneumology, 2Department of Cardiac Surgery, Heidelberg University Hospital, 3Institute for Medical Biometry and Informatics, University of Heidelberg, Heidelberg, Germany Background: Amiodarone is a frequently used antiarrhythmic drug in patients with end-stage heart failure. Given its long half-life, pre-transplant use of amiodarone has been controversially discussed, with divergent results regarding morbidity and mortality after heart transplantation (HTX.Aim: The aim of this study was to investigate the effects of long-term use of amiodarone before HTX on early post-transplant atrial fibrillation (AF and mortality after HTX.Methods: Five hundred and thirty patients (age ≥18 years receiving HTX between June 1989 and December 2012 were included in this retrospective single-center study. Patients with long-term use of amiodarone before HTX (≥1 year were compared to those without long-term use (none or <1 year of amiodarone. Primary outcomes were early post-transplant AF and mortality after HTX. The Kaplan–Meier estimator using log-rank tests was applied for freedom from early post-transplant AF and survival.Results: Of the 530 patients, 74 (14.0% received long-term amiodarone therapy, with a mean duration of 32.3±26.3 months. Mean daily dose was 223.0±75.0 mg. Indications included AF, Wolff–Parkinson–White syndrome, ventricular tachycardia, and ventricular fibrillation. Patients with long-term use of amiodarone before HTX had significantly lower rates of early post-transplant AF (P=0.0105. Further, Kaplan–Meier analysis of freedom from early post-transplant AF showed significantly lower rates of AF in this

Inclusion complex of amiodarone hydrochloride with cyclodextrins: preparation, characterization and dissolution rate evaluation

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Alexandre Machado Rubim

2017-06-01

Full Text Available ABSTRACT This study aimed to improve the water solubility of amiodarone hydrochloride (AMH via inclusion complexes with β-cyclodextrin, methyl-β-cyclodextrin and 2-hydroxypropyl-β-cyclodextrin. Inclusion complexes were developed by physical mixture, coevaporation, spray-drying and freeze-drying. Solid state analysis was performed using X-ray powder diffraction, differential scanning calorimetry and scanning electronic microscopy. Thermodynamic studies demonstrate that the inclusion complexes of drug into different cyclodextrins were an exothermic process that occurred spontaneously. Water solubility and drug dissolution rates were significantly increased after the formation of inclusion complexes with the cyclodextrins evaluated in relation to the physical mixture and pure drug. The present study provides useful information for the potential application of complexation with amiodarone HCl. This may be a good strategy for the development of solid pharmaceutical dosage forms.

Destabilization of emulsions by natural minerals.

Science.gov (United States)

Yuan, Songhu; Tong, Man; Wu, Gaoming

2011-09-15

This study developed a novel method to destabilize emulsions and recycle oils, particularly for emulsified wastewater treatment. Natural minerals were used as demulsifying agents, two kinds of emulsions collected from medical and steel industry were treated. The addition of natural minerals, including artificial zeolite, natural zeolite, diatomite, bentonite and natural soil, could effectively destabilize both emulsions at pH 1 and 60 °C. Over 90% of chemical oxygen demand (COD) can be removed after treatment. Medical emulsion can be even destabilized by artificial zeolite at ambient temperature. The mechanism for emulsion destabilization by minerals was suggested as the decreased electrostatic repulsion at low pH, the enhanced gathering of oil microdroplets at elevated temperature, and the further decreased surface potential by the addition of minerals. Both flocculation and coalescence were enhanced by the addition of minerals at low pH and elevated temperature. Copyright © 2011 Elsevier B.V. All rights reserved.

Simultaneous determination of amiodarone and its metabolite desethylamiodarone by high-performance liquid chromatography with chemiluminescent detection

International Nuclear Information System (INIS)

Perez-Ruiz, Tomas; Martinez-Lozano, Carmen; Garcia-Martinez, Maria Dolores

2008-01-01

A novel method was developed for the determination of amiodarone and desethylamiodarone by high-performance liquid chromatography (HPLC) coupled with chemiluminescent (CL) detection. The procedure is based on the post-column photolysis of the analytes into photoproducts which are active in the tris(2,2'-bipyridyl)ruthenium(III) [Ru(bpy) 3 3+ ] CL system. Ru(bpy) 3 3+ was on-line generated by photo-oxidation of the Ru(II) complex in the presence of peroxydisulfate. The separation was carried out on a Mediterranea C 18 column with isocratic elution using a mixture of methanol and 0.017 mol L -1 ammonium sulfate buffer of pH 6.8. Under the optimum conditions, analytical curves, based on standard solutions, were linear over the range 0.1-50 μg mL -1 for amiodarone and 0.5-25 μg mL -1 for desethylamiodarone. The detection limits of amiodarone and desethylamiodarone were 0.02 and 0.11 μg mL -1 , respectively. Intra- and inter-day precision values of 0.9% relative standard deviation (R.S.D.) (n = 10) and 1.6% R.S.D. (n = 15), respectively, were obtained. The method was applied successfully to the determination of these compounds in serum and pharmaceutical formulations

Effect of Lactobacillus casei on the Pharmacokinetics of Amiodarone in Male Wistar Rats

Czech Academy of Sciences Publication Activity Database

Matušková, Z.; Anzenbacher, P.; Večeřa, R.; Siller, M.; Tlaskalová-Hogenová, Helena; Strojil, J.; Anzenbacherová, E.

2017-01-01

Roč. 42, č. 1 (2017), s. 29-36 ISSN 0378-7966 R&D Projects: GA ČR(CZ) GAP303/12/0535 Institutional support: RVO:61388971 Keywords : Lactobacillus casei * Amiodarone * Wistar Rats Subject RIV: EE - Microbiology, Virology OBOR OECD: Microbiology Impact factor: 1.400, year: 2016

Effects of triiodothyronine and amiodarone on the promoter of the human LDL receptor gene

NARCIS (Netherlands)

Bakker, O.; Hudig, F.; Meijssen, S.; Wiersinga, W. M.

1998-01-01

Treatment of patients with amiodarone, a potent anti arrhythmic drug, increases plasma LDL cholesterol levels, similar to that seen during hypothyroidism. This increase is the result of a decreased expression of the hepatic LDL receptor gene. We investigated the effects of thyroid hormone,

Amiodarone biokinetics, the formation of its major oxidative metabolite and neurotoxicity after acute and repeated exposure of brain cell cultures.

Science.gov (United States)

Pomponio, Giuliana; Zurich, Marie-Gabrielle; Schultz, Luise; Weiss, Dieter G; Romanelli, Luca; Gramowski-Voss, Alexandra; Di Consiglio, Emma; Testai, Emanuela

2015-12-25

The difficulty in mimicking nervous system complexity and cell-cell interactions as well as the lack of kinetics information has limited the use of in vitro neurotoxicity data. Here, we assessed the biokinetic profile as well as the neurotoxicity of Amiodarone after acute and repeated exposure in two advanced rodent brain cell culture models, consisting of both neurons and glial cells organized in 2 or 3 dimensions to mimic the brain histiotypic structure and function. A strategy was applied to evidence the abiotic processes possibly affecting Amiodarone in vitro bioavailability, showing its ability to adsorb to the plastic devices. At clinically relevant Amiodarone concentrations, known to induce neurotoxicity in some patients during therapeutic treatment, a complete uptake was observed in both models in 24 h, after single exposure. After repeated treatments, bioaccumulation was observed, especially in the 3D cell model, together with a greater alteration of neurotoxicity markers. After 14 days, Amiodarone major oxidative metabolite (mono-N-desethylamiodarone) was detected at limited levels, indicating the presence of active drug metabolism enzymes (i.e. cytochrome P450) in both models. The assessment of biokinetics provides useful information on the relevance of in vitro toxicity data and should be considered in the design of an Integrated Testing Strategy aimed to identify specific neurotoxic alerts, and to improve the neurotoxicity assay predictivity for human acute and repeated exposure. Copyright © 2015 Elsevier Ltd. All rights reserved.

Ready-to-use parenteral amiodarone : A feasibility study towards a long-term stable product formulation

NARCIS (Netherlands)

Jacobs, Maartje S.; Luinstra, Marianne; Moes, Jan Reindert; Chan, Tiffany C. Y.; Minovic, Isidor; Frijlink, Henderik W.; Woerdenbag, Herman J.

Objectives To determine the feasibility of preparing a long-term stable ready-to-use parenteral amiodarone formulation using cyclodextrins as dissolution enhancer. Methods A preformulation study was performed with different molar ratios of hydroxypropyl-beta-cyclodextrin (HP-BCD) or

Moduli destabilization via gravitational collapse

Energy Technology Data Exchange (ETDEWEB)

Hwang, Dong-il [Sogang Univ., Seoul (Korea, Republic of). Center for Quantum Spacetime; Pedro, Francisco G. [Deutsches Elektronen-Synchrotron DESY, Hamburg (Germany). Theory Group; Yeom, Dong-han [Sogang Univ., Seoul (Korea, Republic of). Center for Quantum Spacetime; Kyoto Univ. (Japan). Yukawa Inst. for Theoretical Physics

2013-06-15

We examine the interplay between gravitational collapse and moduli stability in the context of black hole formation. We perform numerical simulations of the collapse using the double null formalism and show that the very dense regions one expects to find in the process of black hole formation are able to destabilize the volume modulus. We establish that the effects of the destabilization will be visible to an observer at infinity, opening up a window to a region in spacetime where standard model's couplings and masses can differ significantly from their background values.

Comparative in vitro and in vivo evaluation of three tablet formulations of amiodarone in healthy subjects

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J Emami

2010-09-01

Full Text Available "nBackground and the purpose of the study:The relative in vivo bioavailability and in vitro dissolution studies of three chemically equivalent amiodarone generic products in healthy volunteers was evaluated in three separate occasions. The possibility of a correlation between in vitro and in vivo performances of these tablet formulations was also evaluated. "nMethods: The bioequivalence studies were conducted based on a single dose, two-sequence, cross over randomized design. The bioavailability was compared using AUC0-72, AUC0-∞, Cmax and Tmax. Similarity factor, dissolution efficiency (DE, and mean dissolution time (MDT was used to compare the dissolution profiles. Polynomial linear correlation models were tested using either MDT vs mean residence time (MRT or fraction of the drug dissolved (FRD vs fraction of the drug absorbed (FRA. "nResults: Significant differences were found in the dissolution performances of the tested formulations and therefore they were included in the development of the correlation. The 90% confidence intervals of the log-transformed AUC0-72, AUC0-∞, and Cmax of each two formulations in each bioequivalence studies were within the acceptable range of 80-125%. Differences were not observed between the untransformed Tmax values. Poor correlation was found between MRT and MDT of the products. A point-to-point correlation which is essential for a reliable correlation was not obtained between pooled FRD and FRA. The dissolution condition which was used for amiodarone tablets failed for formulations which were bioequivalent in vivo and significant difference between the dissolution characteristics of products (f2<50 did not reflect their in vivo properties. Major conclusions: Bioequivalence studies should be considered as the only acceptable way to ensure the interchangeability and in vivo equivalence of amiodarone generic drug products. The dissolution conditions used of the present study could be used for routine and in

Amiodarona causa vasodilatação dependente do endotélio em artérias coronárias caninas Amiodarone causes endothelium-dependent vasodilation in canine coronary arteries

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Alfredo José Rodrigues

2005-03-01

Full Text Available OBJETIVO: Avaliar os efeitos vasodilatadores da amiodarona em artérias coronárias caninas empregando soluções de amiodarona dissolvida em polisorbato 80 ou em água. MÉTODOS: Anéis de artéria coronária, com e sem o endotélio íntegro, foram imersos em solução de krebs e conectadas a um transdutor para aferição de força isométrica promovida por contração vascular. As artérias foram expostas a concentrações crescentes de polisorbato 80, amiodarona dissolvida em água, amiodarona dissolvida em polisorbato 80 e uma apresentação comercial da amiodarona (Cordarone®. Os experimentos foram conduzidos na presença e na ausência dos seguintes bloqueadores enzimáticos: apenas indometacina, Nômega-nitro-L-arginina associada à indometacina e apenas Nômega-nitro-L-arginina. RESULTADOS: O polisorbato 80 causou pequeno relaxamento não dependente do endotélio. O Cordarone®, a amiodarona dissolvida em água e em polisorbato 80 promoveram relaxamento dependente do endotélio, que foi de maior magnitude para a amiodarona dissolvida em polisorbato e para o Cordarone®. Apenas a associação de indometacina com a Nômega-nitro-L-arginina foi capaz de abolir o relaxamento dependente do endotélio provocado pela amiodarona dissolvida em polisorbato 80. CONCLUSÃO: Os resultados obtidos indicam que a vasodilatação promovida pela amiodarona em artérias coronárias caninas é causada principalmente pela estimulação da liberação de óxido nítrico e fatores endoteliais relaxantes dependentes das ciclo-oxigenases.OBJECTIVE: To assess the vasodilating effects of amiodarone on canine coronary arteries by using solutions of amiodarone dissolved in polysorbate 80 or water. METHODS: Rings of coronary arteries, with or without intact endothelium, were immersed in Krebs solution and connected to a transducer for measuring the isometric force promoted by a vascular contraction. The arteries were exposed to increasing concentrations of

Heart rate is a marker of amiodarone mortality reduction in severe heart failure. The GESICA-GEMA Investigators. Grupo de Estudio de la Sobrevida en la Insuficiencia Cardiaca en Argentina-Grupo de Estudios Multicéntricos en Argentina.

Science.gov (United States)

Nul, D R; Doval, H C; Grancelli, H O; Varini, S D; Soifer, S; Perrone, S V; Prieto, N; Scapin, O

1997-05-01

The impact of amiodarone on mortality in patients with severe congestive heart failure (CHF) (New York Heart Association functional classes II [advanced], III and IV; left ventricular ejection fraction Insuficiencia Cardiaca en Argentina (GESICA) trial was analyzed in relation to initial mean baseline heart rate (BHR) and its change after 6 months of follow-up. Trials of amiodarone therapy in CHF have produced discordant results, suggesting that the effect is not uniform in all patient subgroups with regard to survival. The present analysis was carried out in 516 patients randomized to receive amiodarone, 300 mg/day (n = 260), or nonantiarrhythmic therapy (n = 256, control group) and followed up for 2 years. Survival was evaluated for patients with a BHR > or = 90 beats/min (control: n = 132; amiodarone: n = 122) and or = 90 beats/min, amiodarone therapy reduced mortality to 38.4% compared with 62.4% in control patients (relative risk [RR] 0.55, 95% confidence interval [CI] 0.35 to 0.95, p or = 90 beats/min, which was reduced at 6 months. Elevated rest heart rates in severe CHF identify a subgroup of patients who benefit from treatment with amiodarone. Amiodarone-induced heart rate slowing may be an important benefit for patients.

Effect of low oral doses of disopyramide and amiodarone on ventricular and atrial arrhythmias of chagasic patients with advanced myocardial damage.

Science.gov (United States)

Carrasco, H A; Vicuña, A V; Molina, C; Landaeta, A; Reynosa, J; Vicuña, N; Fuenmayor, A; López, F

1985-12-01

Low-dose (7 mg/kg per day) disopyramide administration to arrhythmic chagasic patients decreased the frequency of ventricular extrasystoles in 4 of 17 patients (24%) and suppressed most complex ventricular arrhythmias in 12 of 15 patients (80%). This assessment was made from 72-h continuous Holter monitoring recorded during the course of this double blind, placebo-controlled randomized crossover study. Seven patients (41%) complained of anticholinergic side effects, but no contractile or conduction system depression was seen. Amiodarone (200 mg) given on a single blind, placebo-controlled basis to 9 of these patients reduced the frequency of ventricular extrasystoles in 6 of 9 patients (67%) and suppressed complex ventricular ectopy in 6 of 7 patients (85%). One patient was unable to tolerate this drug (11%). Both drugs seemed less effective in controlling supraventricular arrhythmias, although disopyramide eliminated paroxysms of supraventricular tachycardia in 9 of 13 (69%) and amiodarone in all 6 patients with this arrhythmia. Amiodarone appears to be a better antiarrhythmic drug for chagasic patients, due to its greater effectiveness and lower incidence of side effects.

Effects of amiodarone therapy on thyroid iodine content as measured by x-ray fluorescence

International Nuclear Information System (INIS)

Fragu, P.; Schlumberger, M.; Davy, J.M.; Slama, M.; Berdeaux, A.

1988-01-01

Thyroid iodine content (TIC) was measured by x-ray fluorescence in 68 patients who had received amiodarone treatment for varying intervals (1 g/week for 1-120 months). Thirty-six patients were euthyroid; the mean TIC of the patients (n = 15), who had been treated for less than 12 months was 30 +/- 19 (+/- SD) mg, twice the normal mean value (14.6 +/- 5.0 mg), and it was 39 +/- 17 mg in those (n = 16) who had been treated for 12-60 months and 29 +/- 6 mg in those (n = 5) who had been treated longer (greater than 60 months). Nineteen patients were hyperthyroid and had elevated TIC values. Of them, 6 patients had a goiter; their TIC (50 +/- 19 mg) was not significantly different from that of the hyperthyroid patients with no goiter (55 +/- 29 mg), but they became hyperthyroid more rapidly. Thirteen patients were hypothyroid; none had TIC values above the normal range, and it was below 2.5 mg in 5 patients. A sequential study was undertaken in 11 euthyroid patients who had no detectable antithyroid antibodies. TIC did not increase during treatment in 2 patients; both developed hypothyroidism, which was transient in 1 despite continuation of amiodarone treatment. The TIC initially increased during amiodarone treatment in the other 9 patients, leveling off at the end of the first year. The TIC rose well above the upper limit of the normal range in 4 patients, of whom 2 became hyperthyroid during the second year of treatment. TIC remained within the normal range in the other 5 patients, of whom 3 became hypothyroid after 12-24 months of treatment (1 subclinical, 2 overt). Although the TIC was significantly higher in the patients with hyperthyroidism than in the patients who remained euthyroid, the TIC test cannot be used to predict the occurrence of hyperthyroidism

Stabilizing versus Destabilizing the Microtubules: A Double-Edge Sword for an Effective Cancer Treatment Option?

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Daniele Fanale

2015-01-01

Full Text Available Microtubules are dynamic and structural cellular components involved in several cell functions, including cell shape, motility, and intracellular trafficking. In proliferating cells, they are essential components in the division process through the formation of the mitotic spindle. As a result of these functions, tubulin and microtubules are targets for anticancer agents. Microtubule-targeting agents can be divided into two groups: microtubule-stabilizing, and microtubule-destabilizing agents. The former bind to the tubulin polymer and stabilize microtubules, while the latter bind to the tubulin dimers and destabilize microtubules. Alteration of tubulin-microtubule equilibrium determines the disruption of the mitotic spindle, halting the cell cycle at the metaphase-anaphase transition and, eventually, resulting in cell death. Clinical application of earlier microtubule inhibitors, however, unfortunately showed several limits, such as neurological and bone marrow toxicity and the emergence of drug-resistant tumor cells. Here we review several natural and synthetic microtubule-targeting agents, which showed antitumor activity and increased efficacy in comparison to traditional drugs in various preclinical and clinical studies. Cryptophycins, combretastatins, ombrabulin, soblidotin, D-24851, epothilones and discodermolide were used in clinical trials. Some of them showed antiangiogenic and antivascular activity and others showed the ability to overcome multidrug resistance, supporting their possible use in chemotherapy.

Short-term amiodarone treatment for atrial fibrillation after catheter ablation induces a transient thyroid dysfunction

DEFF Research Database (Denmark)

Diederichsen, Søren Zöga; Darkner, Stine; Chen, Xu

2016-01-01

ablation in a randomised, double-blind clinical trial. METHODS: 212 patients referred for AF ablation at two centres were randomized to 8weeks of oral amiodarone or placebo. Thyroid function tests (TSH, thyroid stimulating hormone; T4, thyroxine; T3, triiodothyronine; fT4, free T4; fT3, free T3) were...

AMIODARONE INDUCES THE SYNTHESIS OF HSPS IN SACCHAROMYCES CEREVISIAE AND ARABIDOPSIS THALIANA CELLS

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Pyatrikas D.V.

2012-08-01

Full Text Available Many biotic and abiotic stresses cause an increase of cytosolic Ca2+ level in cells. Calcium is one of the most important second messengers, regulating many various activities in the cell and was known to affect expression of stress activated genes. Mild heat shock induces the expression of heat shock proteins (Hsps which protect cell from drastic heat shock exposure. There are some literature data permitting to suggest that transient elevation of cytosolic Ca2+ level in plant cells is important for activation of Hsps expression. On the other hand mitochondria are known to regulate the intracellular calcium and reactive oxygen species signaling. It has been shown recently that mild heat shock induces hyperpolarization of inner mitochondrial membrane in plant and yeast cells and this event is critically important for activation of Hsps expression. To reveal the relationship between mitochondrial activity, intracellular calcium homeostasis and Hsps expression an antiarrhythmic drug amiodarone (AMD have been used. AMD is known to cause transient increase of cytosolic Ca2+ level in Saccharomyces cerevisiae. Obtained results have showed that AMD treatment induced the synthesis of Hsp104p in S. cerevisiae cells and Hsp101p in A. thaliana cell culture. Induction of Hsp104p synthesis leads to enhanced yeast capability to survive lethal heat shock exposure. Development of S. cerevisiae thermotolerance depended significantly on the presence of Hsp104p. Elevation of Hsp104p level in the result of AMD treatment was shown to be governed by activity of Msn2p and Msn4p transcription factors. Deletion of the MSN2 and MSN4 genes abrogated the AMD ability to induce Hsp104p synthesis. Mild heat shock and AMD treatment induced the hyperpolarization of the inner mitochondrial membrane in yeast and Arabidopsis cells which accompanied by HSP synthesis and development of thermotolerance. It was suggested that increase of cytosolic Ca2+ level after AMD treatment

Comparative Study of Nifekalant Versus Amiodarone for Shock-Resistant Ventricular Fibrillation in Out-of-Hospital Cardiopulmonary Arrest Patients

NARCIS (Netherlands)

Amino, Mari; Yoshioka, Koichiro; Opthof, Tobias; Morita, Seiji; Uemura, Shunryo; Tamura, Kozo; Fukushima, Tomokazu; Higami, Shigeo; Otsuka, Hiroyuki; Akieda, Kazuki; Shima, Makiyoshi; Fujibayashi, Daisuke; Hashida, Tadashi; Inokuchi, Sadaki; Kodama, Itsuo; Tanabe, Teruhisa

2010-01-01

Background: In Japan, intravenous nifekalant ( NIF) was often used for direct current cardioversion-resistant ventricular fibrillation (VF), until the use of intravenous amiodarone (AMD) was approved in 2007. The defibrillatory efficacy of NIF and AMD has thus far not been compared for

Amiodarone therapy in chronic heart failure and myocardial infarction: a review of the mortality trials with special attention to STAT-CHF and the GESICA trials. Grupo de Estudio de la Sobrevida en la Insuficiencia Cardiaca en Argentina.

Science.gov (United States)

Pinto, J V; Ramani, K; Neelagaru, S; Kown, M; Gheorghiade, M

1997-01-01

Amiodarone appears to reduce sudden death in patients with left ventricular dysfunction resulting from an acute MI or a primary dilated cardiomyopathy, particularly if complex ventricular arrhythmias are present. Amiodarone's beneficial effect on mortality in these patients could be unrelated to its antiarrhythmic effects. Multiple factors could account for the improvement in mortality such as the drug's antiischemic effects, neuromodulating effects, its effect on left ventricular function and on heart rate. Moreover, patients with LV dysfunction who have survived an episode of sudden death would potentially benefit from amiodarone therapy. Future trials are needed to determine the precise subsets(s) of patients who would benefit from the drug and the most efficacious dosing regimen for the drug. Based on available data, amiodarone is the only antiarrhythmic agent which has not been shown to increase mortality in patients with chronic heart failure.

Effect of amiodarone and dronedarone administration in rats on thyroid hormone-dependent gene expression in different cardiac components

NARCIS (Netherlands)

Stoykov, I.; van Beeren, H. C.; Moorman, A. F. M.; Christoffels, V. M.; Wiersinga, W. M.; Bakker, O.

2007-01-01

OBJECTIVE: In view of their different actions on thyroid hormone receptor (TR) isoforms we set out to investigate whether amiodarone (AM) and dronedarone (Dron) have different and/or component-specific effects on cardiac gene expression. DESIGN: Rats were treated with AM or Dron and the expression

Interaction between amiodarone and hepatitis-C virus nucleotide inhibitors in human induced pluripotent stem cell-derived cardiomyocytes and HEK-293 Cav{sub 1.2} over-expressing cells

Energy Technology Data Exchange (ETDEWEB)

Lagrutta, Armando, E-mail: armando_lagrutta@merck.com; Zeng, Haoyu; Imredy, John; Balasubramanian, Bharathi; Dech, Spencer; Lis, Edward; Wang, Jixin; Zhai, Jin; DeGeorge, Joseph; Sannajust, Frederick

2016-10-01

Several clinical cases of severe bradyarrhythmias have been reported upon co-administration of the Hepatitis-C NS5B Nucleotide Polymerase Inhibitor (HCV-NI) direct-acting antiviral agent, sofosbuvir (SOF), and the Class-III anti-arrhythmic amiodarone (AMIO). We model the cardiac drug-drug interaction (DDI) between AMIO and SOF, and between AMIO and a closely-related SOF analog, MNI-1 (Merck Nucleotide Inhibitor #1), in functional assays of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), to provide mechanistic insights into recently reported clinical cases. AMIO co-applied with SOF or MNI-1 increased beating rate or field potential (FP) rate and decreased impedance (IMP) and Ca{sup 2+} transient amplitudes in hiPSC-CM syncytia. This action resembled that of Ca{sup 2+} channel blockers (CCBs) in the model, but CCBs did not substitute for AMIO in the DDI. AMIO analog dronedarone (DRON) did not substitute for, but competed with AMIO in the DDI. Ryanodine and thapsigargin, decreasing intracellular Ca{sup 2+} stores, and SEA-0400, a Na{sup +}/Ca{sup 2+} exchanger-1 (NCX1) inhibitor, partially antagonized or suppressed DDI effects. Other agents affecting FP rate only exerted additive or subtractive effects, commensurate with their individual effects. We also describe an interaction between AMIO and MNI-1 on Cav{sub 1.2} ion channels in an over-expressing HEK-293 cell line. MNI-1 enhanced Cav{sub 1.2} channel inhibition by AMIO, but did not affect inhibition of Cav{sub 1.2} by DRON, verapamil, nifedipine, or diltiazem. Our data in hiPSC-CMs indicate that HCV-NI agents such as SOF and MNI-1 interact with key intracellular Ca{sup 2+}-handling mechanisms. Additional study in a Cav{sub 1.2} HEK-293 cell-line suggests that HCV-NIs potentiate the inhibitory action of AMIO on L-type Ca{sup 2+} channels. - Highlights: • Adverse clinical interaction between amiodarone and HCV-NI drugs is captured by in vitro models. • Human iPSC-derived cardiomyocyte

Efficacy of topical chamomile on the incidence of phlebitis due to an amiodarone infusion in coronary care patients: a double-blind, randomized controlled trial.

Science.gov (United States)

Sharifi-Ardani, Maryam; Yekefallah, Leili; Asefzadeh, Saeed; Nassiri-Asl, Marjan

2017-09-01

Amiodarone is a useful antiarrhythmic drug. Phlebitis, caused by intravenous amiodarone, is common in patients in coronary care units (CCUs). The aim of this study was to evaluate the effect of topical chamomile on the incidence of phlebitis due to the administration of an amiodarone infusion into the peripheral vein. This was a randomized, double-blind clinical trial, conducted on 40 patients (n = 20 per group) in two groups-an intervention group (chamomile ointment) and a control group (lanoline, as a placebo), hospitalized in the CCUs and undergoing an amiodarone infusion into the peripheral vein over 24 h. Following the cannulation and commencement of the infusion, placebo or chamomile ointment was rubbed in, up to 10 cm superior to the catheter and repeated every eight hours for three days. The cannula site was then assessed based on the phlebitis checklist. The incidence and time of occurrence of phlebitis, relative risk, severity of phlebitis were the main outcome measures. Nineteen patients (19/20) in the control group had phlebitis on the first day of the study and one patient (20/20) on the second day. In the intervention group, phlebitis occurred in 13 cases (13/20) on the first day and another two (2/7) was found on the second day. The incidence of phlebitis was significantly different between two groups (P = 0.023). The cumulative incidence of phlebitis in the intervention group (15/20) is significantly later and lower than that in the control group (20/20) during two days (P = 0.008). Two patients in the intervention group did not develop phlebitis at all during the 3-day study. Also, the relative risk of phlebitis in the two groups was 0.68 (P = 0.008 5). A significant difference was not observed with regard to phlebitis severity in both groups. It seems that phlebitis occurred to a lesser extent and at a later time frame in the intervention group compared to control group. Topical chamomile may be effective in decreasing the incidence of phlebitis

Coadministration of Atorvastatin and Amiodarone Increases the Risk of Pulmonary Fibrosis in Rats

OpenAIRE

Nasri, Hamid-Reza; Joukar, Siyavash; Kheradmand, Hamid; Poursalehi, Hamid-Reza; Dabiri, Shahriar

2015-01-01

Objective The purpose of this study was to evaluate the effect of atorvastatin administration on amiodarone-induced pulmonary fibrosis in rats. Materials and Methods Thirty-six male Wistar rats were randomly divided into 4 groups. The control group (CTL) received distilled water (0.3 ml intratracheally on days 0 and 2 and 0.5 ml orally from day 0 for 3 weeks). The atorvastatin group (AT), in addition to intratracheal distilled water, received 1 mg/kg of atorvastatin orally from day 0 for 3 we...

MTOR-Driven Metabolic Reprogramming Regulates Legionella pneumophila Intracellular Niche Homeostasis

Science.gov (United States)

Abshire, Camille F.; Roy, Craig R.

2016-01-01

Vacuolar bacterial pathogens are sheltered within unique membrane-bound organelles that expand over time to support bacterial replication. These compartments sequester bacterial molecules away from host cytosolic immunosurveillance pathways that induce antimicrobial responses. The mechanisms by which the human pulmonary pathogen Legionella pneumophila maintains niche homeostasis are poorly understood. We uncovered that the Legionella-containing vacuole (LCV) required a sustained supply of host lipids during expansion. Lipids shortage resulted in LCV rupture and initiation of a host cell death response, whereas excess of host lipids increased LCVs size and housing capacity. We found that lipids uptake from serum and de novo lipogenesis are distinct redundant supply mechanisms for membrane biogenesis in Legionella-infected macrophages. During infection, the metabolic checkpoint kinase Mechanistic Target of Rapamycin (MTOR) controlled lipogenesis through the Serum Response Element Binding Protein 1 and 2 (SREBP1/2) transcription factors. In Legionella-infected macrophages a host-driven response that required the Toll-like receptors (TLRs) adaptor protein Myeloid differentiation primary response gene 88 (Myd88) dampened MTOR signaling which in turn destabilized LCVs under serum starvation. Inactivation of the host MTOR-suppression pathway revealed that L. pneumophila sustained MTOR signaling throughout its intracellular infection cycle by a process that required the upstream regulator Phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) and one or more Dot/Icm effector proteins. Legionella-sustained MTOR signaling facilitated LCV expansion and inhibition of the PI3K-MTOR-SREPB1/2 axis through pharmacological or genetic interference or by activation of the host MTOR-suppression response destabilized expanding LCVs, which in turn triggered cell death of infected macrophages. Our work identified a host metabolic requirement for LCV homeostasis and demonstrated that L

Atherosclerotic Plaque Destabilization in Mice: A Comparative Study.

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Helene Hartwig

Full Text Available Atherosclerosis-associated diseases are the main cause of mortality and morbidity in western societies. The progression of atherosclerosis is a dynamic process evolving from early to advanced lesions that may become rupture-prone vulnerable plaques. Acute coronary syndromes are the clinical manifestation of life-threatening thrombotic events associated with high-risk vulnerable plaques. Hyperlipidemic mouse models have been extensively used in studying the mechanisms controlling initiation and progression of atherosclerosis. However, the understanding of mechanisms leading to atherosclerotic plaque destabilization has been hampered by the lack of proper animal models mimicking this process. Although various mouse models generate atherosclerotic plaques with histological features of human advanced lesions, a consensus model to study atherosclerotic plaque destabilization is still lacking. Hence, we studied the degree and features of plaque vulnerability in different mouse models of atherosclerotic plaque destabilization and find that the model based on the placement of a shear stress modifier in combination with hypercholesterolemia represent with high incidence the most human like lesions compared to the other models.

Control of locomotor stability in stabilizing and destabilizing environments.

Science.gov (United States)

Wu, Mengnan/Mary; Brown, Geoffrey; Gordon, Keith E

2017-06-01

To develop effective interventions targeting locomotor stability, it is crucial to understand how people control and modify gait in response to changes in stabilization requirements. Our purpose was to examine how individuals with and without incomplete spinal cord injury (iSCI) control lateral stability in haptic walking environments that increase or decrease stabilization demands. We hypothesized that people would adapt to walking in a predictable, stabilizing viscous force field and unpredictable destabilizing force field by increasing and decreasing feedforward control of lateral stability, respectively. Adaptations in feedforward control were measured using after-effects when fields were removed. Both groups significantly (pfeedforward adaptions to increase control of lateral stability. In contrast, in the destabilizing field, non-impaired subjects increased movement variability (p0.05). When the destabilizing field was removed, increases in movement variability persisted (pfeedforward decreases in resistance to perturbations. Published by Elsevier B.V.

Cell internalizable and intracellularly degradable cationic polyurethane micelles as a potential platform for efficient imaging and drug delivery.

Science.gov (United States)

Ding, Mingming; Zeng, Xin; He, Xueling; Li, Jiehua; Tan, Hong; Fu, Qiang

2014-08-11

A cell internalizable and intracellularly degradable micellar system, assembled from multiblock polyurethanes bearing cell-penetrating gemini quaternary ammonium pendent groups in the side chain and redox-responsive disulfide linkages throughout the backbone, was developed for potential magnetic resonance imaging (MRI) and drug delivery. The nanocarrier is featured as a typical "cleavable core-internalizable shell-protective corona" architecture, which exhibits small size, positive surface charge, high loading capacity, and reduction-triggered destabilization. Furthermore, it can rapidly enter tumor cells and release its cargo in response to an intracellular level of glutathione, resulting in enhanced drug efficacy in vitro. The magnetic micelles loaded with superparamagnetic iron oxide (SPIO) nanoparticles demonstrate excellent MRI contrast enhancement, with T2 relaxivity found to be affected by the morphology of SPIO-clustering inside the micelle core. The multifunctional carrier with good cytocompatibility and nontoxic degradation products can serve as a promising theranostic candidate for efficient intracellular delivery of anticancer drugs and real-time monitoring of therapeutic effect.

Intracellular drug delivery nanocarriers of glutathione-responsive degradable block copolymers having pendant disulfide linkages.

Science.gov (United States)

Khorsand, Behnoush; Lapointe, Gabriel; Brett, Christopher; Oh, Jung Kwon

2013-06-10

Self-assembled micelles of amphiphilic block copolymers (ABPs) with stimuli-responsive degradation (SRD) properties have a great promise as nanotherapeutics exhibiting enhanced release of encapsulated therapeutics into targeted cells. Here, thiol-responsive degradable micelles based on a new ABP consisting of a pendant disulfide-labeled methacrylate polymer block (PHMssEt) and a hydrophilic poly(ethylene oxide) (PEO) block were investigated as effective intracellular nanocarriers of anticancer drugs. In response to glutathione (GSH) as a cellular trigger, the cleavage of pendant disulfide linkages in hydrophobic PHMssEt blocks of micellar cores caused the destabilization of self-assembled micelles due to change in hydrophobic/hydrophilic balance. Such GSH-triggered micellar destabilization changed their size distribution with an appearance of large aggregates and led to enhanced release of encapsulated anticancer drugs. Cell culture results from flow cytometry and confocal laser scanning microscopy for cellular uptake as well as cell viability measurements for high anticancer efficacy suggest that new GSH-responsive degradable PEO-b-PHMssEt micelles offer versatility in multifunctional drug delivery applications.

Destabilization and recovery of a yeast prion after mild heat shock.

Science.gov (United States)

Newnam, Gary P; Birchmore, Jennifer L; Chernoff, Yury O

2011-05-06

Yeast prion [PSI(+)] is a self-perpetuating amyloid of the translational termination factor Sup35. Although [PSI(+)] propagation is modulated by heat shock proteins (Hsps), high temperature was previously reported to have little or no effect on [PSI(+)]. Our results show that short-term exposure of exponentially growing yeast culture to mild heat shock, followed by immediate resumption of growth, leads to [PSI(+)] destabilization, sometimes persisting for several cell divisions after heat shock. Prion loss occurring in the first division after heat shock is preferentially detected in a daughter cell, indicating the impairment of prion segregation that results in asymmetric prion distribution between a mother cell and a bud. Longer heat shock or prolonged incubation in the absence of nutrients after heat shock led to [PSI(+)] recovery. Both prion destabilization and recovery during heat shock depend on protein synthesis. Maximal prion destabilization coincides with maximal imbalance between Hsp104 and other Hsps such as Hsp70-Ssa. Deletions of individual SSA genes increase prion destabilization and/or counteract recovery. The dynamics of prion aggregation during destabilization and recovery are consistent with the notion that efficient prion fragmentation and segregation require a proper balance between Hsp104 and other (e.g., Hsp70-Ssa) chaperones. In contrast to heat shock, [PSI(+)] destabilization by osmotic stressors does not always depend on cell proliferation and/or protein synthesis, indicating that different stresses may impact the prion via different mechanisms. Our data demonstrate that heat stress causes asymmetric prion distribution in a cell division and confirm that the effects of Hsps on prions are physiologically relevant. Copyright © 2011 Elsevier Ltd. All rights reserved.

Pattern destabilization and emotional processing in cognitive therapy for personality disorders.

Science.gov (United States)

Hayes, Adele M; Yasinski, Carly

2015-01-01

Clinical trials of treatments for personality disorders can provide a medium for studying the process of therapeutic change with particularly entrenched and self-perpetuating systems and might reveal important principles of system transition. We examined the extent to which maladaptive personality patterns were destabilized in a trial of cognitive therapy personality disorders (CT-PD) and how destabilization was associated with emotional processing and treatment outcomes. Dynamic systems theory was used as a theoretical framework for studying change. Participants were 27 patients diagnosed with Avoidant or Obsessive Compulsive Personality Disorder (AVPD or OCPD), who completed an open trial of CT-PD. Raters coded treatment sessions using a coding system that operationalizes emotional processing, as well as cognitive, affective, behavioral, and somatic components of pathological (negative) and more adaptive (positive) patterns of functioning. Pattern destabilization (dispersion) scores during the early phase of treatment (phase 1: session 1-10) and the schema-focused phase (phase 2: session 11-34) were calculated using a program called GridWare. More pattern destabilization and emotional processing in the schema-focused phase of CT-PD predicted more improvement in personality disorder symptoms and positive pattern strength at the end of treatment, whereas these variables in phase 1 did not predict outcome. In addition to illustrating a quantitative method for studying destabilization and change of patterns of psychopathology, we present findings that are consistent with recent updates of emotional processing theory and with principles from dynamic systems theory.

Pattern destabilization and emotional processing in cognitive therapy for personality disorders

Directory of Open Access Journals (Sweden)

Adele M. Hayes

2015-02-01

Full Text Available Clinical trials of treatments for personality disorders can provide a medium for studying the process of therapeutic change with particularly entrenched and self-perpetuating systems and might reveal important principles of system transition. We examined the extent to which maladaptive personality patterns were destabilized in a trial of cognitive therapy personality disorders (CT-PD and how destabilization was associated with emotional processing and treatment outcomes. Dynamic systems theory was used as a theoretical framework for studying change. Method: Participants were 27 patients diagnosed with Avoidant or Obsessive Compulsive Personality Disorder, who completed an open trial of CT-PD. Raters coded treatment sessions using a coding system that operationalizes emotional processing, as well as cognitive, affective, behavioral, and somatic components of pathological (negative and more adaptive (positive patterns of functioning. Pattern destabilization (dispersion scores during the early phase of treatment (phase 1: session 1-10 and the schema-focused phase (phase 2: session 11-34 were calculated using a program called GridWare. Results: More pattern destabilization and emotional processing in the schema-focused phase of CT-PD predicted more improvement in personality disorder symptoms and positive pattern strength at the end of treatment, whereas these variables in phase 1 did not predict outcome. Conclusions: In addition to illustrating a quantitative method for studying destabilization and change of patterns of psychopathology, we present findings that are consistent with recent updates of emotional processing theory and with principles from dynamic systems theory.

Study with radio aerosol of DTPA technetium-99 m in individuals with pulmonary disease by amiodarone

International Nuclear Information System (INIS)

Terra Filho, M.

1989-01-01

In order to evaluate the role of the clearance of 99 m Technetium chelated to diethylenetriamine-penta-acetate (99 m Tc-DTPA) in amiodarone induced pulmonary disease, 40 individuals were studied in four groups. After spirometry, where a volume-time curve was registered, all individuals inhaled 740 MBq of 99 m Tc-DTPA diluted in 4 ml of saline, for five minutes. Pulmonary images were obtained in a computerized scintillation camera and 9 regions of interest were selected. (author)

Destabilization of TAE modes by particle anisotropy

International Nuclear Information System (INIS)

Wong, H.V.; Berk, H.L.

1998-01-01

Plasmas heated by ICRF produce energetic particle distribution functions which are sharply peaked in pitch-angle, and the authors show that at moderate toroidal mode numbers, this anisotropy is a competitive and even dominant instability drive when compared with the universal instability drive due to spatial gradient. The universal drive, acting along, destabilizes only co-propagating waves (i.e., waves propagating in the same toroidal direction as the diamagnetic flow of the energetic particles), but stabilizes counter-propagating waves (i.e., waves propagating in the opposite toroidal direction as the diamagnetic flow of the energetic particles). Nonetheless, the authors show that in a tokamak, it is possible that particle anisotropy can produce a larger linear growth rate for counter-propagating waves, and provide a mechanism for preferred destabilization of the counter-propagating TAE modes that are sometimes experimentally observed

Destabilization of low-n peeling modes by trapped energetic particles

Energy Technology Data Exchange (ETDEWEB)

Hao, G. Z.; Wang, A. K.; Mou, Z. Z.; Qiu, X. M. [Southwestern Institute of Physics, PO Box 432, Chengdu 610041 (China); Liu, Y. Q. [Euratom/CCFE Fusion Association, Culham Science Centre, Abingdon, OX14 3DB (United Kingdom); Matsunaga, G. [Japan Atomic Energy Agency, 801-1, Mukouyama, Naka, Ibaraki 311-0193 (Japan); Okabayashi, M. [Princeton Plasma Physics Laboratory, PO Box 451, Princeton, New Jersey 08543-0451 (United States)

2013-06-15

The kinetic effect of trapped energetic particles (EPs), arising from perpendicular neutral beam injection, on the stable low-n peeling modes in tokamak plasmas is investigated, through numerical solution of the mode's dispersion relation derived from an energy principle. A resistive-wall peeling mode with m/n=6/1, with m and n being the poloidal and toroidal mode numbers, respectively, is destabilized by trapped EPs as the EPs' pressure exceeds a critical value β{sub c}{sup *}, which is sensitive to the pitch angle of trapped EPs. The dependence of β{sub c}{sup *} on the particle pitch angle is eventually determined by the bounce average of the mode eigenfunction. Peeling modes with higher m and n numbers can also be destabilized by trapped EPs. Depending on the wall distance, either a resistive-wall peeling mode or an ideal-kink peeling mode can be destabilized by EPs.

Involvement of Protein Phosphatases in the Destabilization of Methamphetamine-Associated Contextual Memory

Science.gov (United States)

Yu, Yang-Jung; Huang, Chien-Hsuan; Chang, Chih-Hua; Gean, Po-Wu

2016-01-01

Destabilization refers to a memory that becomes unstable when reactivated and is susceptible to disruption by amnestic agents. Here we delineated the cellular mechanism underlying the destabilization of drug memory. Mice were conditioned with methamphetamine (MeAM) for 3 d, and drug memory was assessed with a conditioned place preference (CPP)…

The effects of nifekalant hydrochloride on the spatial dispersion of repolarization after direct current defibrillation in patients with oral amiodarone and β-blocker therapy

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Keiko Maeda

2014-06-01

Conclusions: NIF suppressed the deterioration of the SDR after ICD shock. This might be one of the mechanisms by which NIF suppresses recurrence of ventricular tachyarrhythmia just after ICD shock in patients with oral amiodarone and β-blocker therapy.

Preformed β-amyloid fibrils are destabilized by coenzyme Q10 in vitro

International Nuclear Information System (INIS)

Ono, Kenjiro; Hasegawa, Kazuhiro; Naiki, Hironobu; Yamada, Masahito

2005-01-01

Inhibition of the formation of β-amyloid fibrils (fAβ), as well as the destabilization of preformed fAβ in the CNS, would be attractive therapeutic targets for the treatment of Alzheimer's disease (AD). We reported previously that nordihydroguaiaretic acid (NDGA) and wine-related polyphenol, myricetin (Myr), inhibit fAβ formation from Aβ and destabilize preformed fAβ in vitro. Using fluorescence spectroscopic analysis with thioflavin T and electron microscopic studies, we examined the effects of coenzyme Q 10 (CoQ 10 ) on the formation, extension, and destabilization of fAβ at pH 7.5 at 37 deg C in vitro. We next compared the anti-amyloidogenic activities of CoQ 10 with NDGA and Myr. CoQ 10 dose-dependently inhibited fAβ formation from amyloid β-peptide (Aβ), as well as their extension. Moreover, it destabilized preformed fAβs. The anti-amyloidogenic effects of CoQ 10 were slightly weaker than those of NDGA and Myr. CoQ 10 could be a key molecule for the development of therapeutics for AD

Estudo prospectivo dos efeitos da amiodarona na função tiroidiana de pacientes chagásicos em área de deficiência de iodo Prospective study of amiodarone effects on thyroid function of chagasic patients in an iodine deficient area

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Maria Aparecida Enes de Barros

1994-09-01

Full Text Available Com o objetivo de avaliar a junção tiroidiana após uso crônico da amiodarona, em área de deficiência de iodo e endemia chagásica, 24 pacientes foram analisados antes e após três e nove meses de uso da droga. A avaliação constou de exame clínico, dosagem sérica de T4, T3, rT3, TSH, anticorpo antitiroglobulina e TSH 30 minutos após infusão venosa de uma ampola de 200µg de TRH. A captação do iodo radioativo 131 e a cintilografia datiróide foram realizadas antes e aos 9 meses após tratamento. Disfunção tiroidiana ocorreu em 20,8% dos pacientes sendo 12,5% de hipertiroidismo e 8,3% de hipotiroidismo, com anticorpos antitiroglobulina negativos. Captação do iodo radioativo 131 foi positiva em um paciente hipertiroideo com bócio. O diagnóstico de hipertiroidismo foi melhor evidenciado pela resposta reduzida ou bloqueada do TSH ao TRH e não pela concentração do T3 no soro e o de hipotiroidismo pela concentração elevada do TSH. O TSH elevado desde o início do tratamento pode predispor ao aparecimento de bócio. Concluímos que o uso da amiodarona em nossa região deve serjudiciosamente analisado, sendo a função tiroidiana cuidadosamente monitorizada antes e durante o tratamento.In order to evaluate the development of thyroid dysfunction during chronic amiodarone treatment in an area deficient in iodine and endemic for Chagas 'disease, a group of 24patients wasprospectively studied. Clinical examination and measurement of serum T4, T3, rT3, TSH and antithyroglobulin antibodies were performed before and at 3 and 9 months of use of amiodarone. A TSH response 30 minutes after IV injection of 200µg of TRH was also compared to TSH basal levels before and during amiodarone treatment. Thyroid radioative uptake and scan were obtained before and nine months after amiodarone was started. Elevated rT3 concentrations were unexpectedly found in two thirds of the patients before treatment. Thyroid dysfunction developed during

Submarine landslides triggered by destabilization of high-saturation hydrate anomalies

Science.gov (United States)

Handwerger, Alexander L.; Rempel, Alan W.; Skarbek, Rob M.

2017-07-01

Submarine landslides occur along continental margins at depths that often intersect the gas hydrate stability zone, prompting suggestions that slope stability may be affected by perturbations that arise from changes in hydrate stability. Here we develop a numerical model to identify the conditions under which the destabilization of hydrates results in slope failure. Specifically, we focus on high-saturation hydrate anomalies at fine-grained to coarse-grained stratigraphic boundaries that can transmit bridging stresses that decrease the effective stress at sediment contacts and disrupt normal sediment consolidation. We evaluate slope stability before and after hydrate destabilization. Hydrate anomalies act to significantly increase the overall slope stability due to large increases in effective cohesion. However, when hydrate anomalies destabilize there is a loss of cohesion and increase in effective stress that causes the sediment grains to rapidly consolidate and generate pore pressures that can either trigger immediate slope failure or weaken the surrounding sediment until the pore pressure diffuses away. In cases where failure does not occur, the sediment can remain weakened for months. In cases where failure does occur, we quantify landslide dynamics using a rate and state frictional model and find that landslides can display either slow or dynamic (i.e., catastrophic) motion depending on the rate-dependent properties, size of the stress perturbation, and the size of the slip patch relative to a critical nucleation length scale. Our results illustrate the fundamental mechanisms through which the destabilization of gas hydrates can pose a significant geohazard.

Electromagnetic waves destabilized by runaway electrons in near-critical electric fields

Energy Technology Data Exchange (ETDEWEB)

Komar, A.; Pokol, G. I. [Department of Nuclear Techniques, Budapest University of Technology and Economics, Association EURATOM, H-1111 Budapest (Hungary); Fueloep, T. [Department of Applied Physics, Nuclear Engineering, Chalmers University of Technology and Euratom-VR Association, Goeteborg (Sweden)

2013-01-15

Runaway electron distributions are strongly anisotropic in velocity space. This anisotropy is a source of free energy that may destabilize electromagnetic waves through a resonant interaction between the waves and the energetic electrons. In this work, we investigate the high-frequency electromagnetic waves that are destabilized by runaway electron beams when the electric field is close to the critical field for runaway acceleration. Using a runaway electron distribution appropriate for the near-critical case, we calculate the linear instability growth rate of these waves and conclude that the obliquely propagating whistler waves are most unstable. We show that the frequencies, wave numbers, and propagation angles of the most unstable waves depend strongly on the magnetic field. Taking into account collisional and convective damping of the waves, we determine the number density of runaways that is required to destabilize the waves and show its parametric dependences.

Mechanisms governing the reactivation-dependent destabilization of memories and their role in extinction

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Charlotte Rachael Flavell

2013-12-01

Full Text Available The extinction of learned associations has traditionally been considered to involve new learning, which competes with the original memory for control over behaviour. However, a recent resurgence of interest in reactivation-dependent amnesia has revealed that the retrieval of fear-related memory (with what is essentially a brief extinction session can result in it’s destabilization. This review discusses some of the cellular and molecular mechanisms that are involved in the destabilization of a memory following it’s reactivation and/or extinction, and investigates the evidence that extinction may involve both new learning as well as a partial destabilization-induced erasure of the original memory trace.

Vectorization efforts to increase Gram-negative intracellular drug concentration: a case study on HldE-K inhibitors.

Science.gov (United States)

Atamanyuk, Dmytro; Faivre, Fabien; Oxoby, Mayalen; Ledoussal, Benoit; Drocourt, Elodie; Moreau, François; Gerusz, Vincent

2013-03-14

In this paper, we present different strategies to vectorize HldE kinase inhibitors with the goal to improve their gram-negative intracellular concentration. Syntheses and biological effects of siderophoric, aminoglycosidic, amphoteric, and polycationic vectors are discussed. While siderophoric and amphoteric vectorization efforts proved to be disappointing in this series, aminoglycosidic and polycationic vectors were able for the first time to achieve synergistic effects of our inhibitors with erythromycin. Although these effects proved to be nonspecific, this study provides information about the required stereoelectronic arrangement of the polycationic amines and their basicity requirements to fulfill outer membrane destabilization resulting in better erythromycin synergies.

Atherosclerotic Plaque Destabilization Mechanisms, Models, and Therapeutic Strategies

NARCIS (Netherlands)

Silvestre-Roig, Carlos; de Winther, Menno P.; Weber, Christian; Daemen, Mat J.; Lutgens, Esther; Soehnlein, Oliver

2014-01-01

Understanding the pathophysiology of atherogenesis and the progression of atherosclerosis have been major goals of cardiovascular research during the previous decades. However, the complex molecular and cellular mechanisms underlying plaque destabilization remain largely obscure. Here, we review how

Energetic particle destabilization of shear Alfven waves in stellarators and tokamaks

International Nuclear Information System (INIS)

Spong, D.A.; Carreras, B.A.; Hedrick, C.L.; Leboeuf, J.N.; Weller, A.

1994-01-01

An important issue for ignited devices is the resonant destabilization of shear Alfven waves by energetic populations. These instabilities have been observed in a variety of toroidal plasma experiments in recent years, including: beam-destabilized toroidal Alfven instabilities (TAE) in low magnetic field tokamaks, ICRF destabilized TAE's in higher field tokamaks, and global Alfven instabilities (GAE) in low shear stellarators. In addition, excitation and study of these modes is a significant goal of the TFIR-DT program and a component of the ITER physics tasks. The authors have developed a gyrofluid model which includes the wave-particle resonances necessary to excite such instabilities. The TAE linear mode structure is calculated nonperturbatively, including many of the relevant damping mechanisms, such as: continuum damping, non-ideal effects (ion FLR and electron collisionality), and ion/electron Landau damping. This model has been applied to both linear and nonlinear regimes for a range of experimental cases using measured profiles

Effect of amiodarone on the conversion of thyroxine to triiodotironine in rat myocardium in vivo and in vitro

International Nuclear Information System (INIS)

Ceppi, J.A.; Gonzalez, M.R.; Zaninovich, A.A.

1988-01-01

It was studied the effect of the antiarrhythmic drug amiodarone, on the conversion of T 4 to T 3 by rat myocardium and liver. Wistar rats were injected with amiodarone (AM) or iopanoic acid (IOP), afterwards the liver and the heart were homogenized in krebs-ringer phosphate. Aliquots of 400μl of the homogenized were separated and DTT 8mM and 10 -2 μCi 125 I-T 4 or 125 I-T 3 were added (either the conversion of T 4 to T 3 or degradation of T 3 are measured). In vitro: were added AM 0,1 mM or IOP 10mM. Incubation proceeded for 2 hours at 37 deg C and aliquots were chromatographied. The conversion of T 4 to T 3 in the myocardium and liver in vivo decreased, but in vitro did not have any deiodination of T 4 and the IOP reduced the production of T 3 by heart. Neither drug altered the deiodination of T 3 by heart or liver homogenates. Serum TSH and T 3 were decreased by AM and serum T 4 increased. IOP caused similar variations in T 4 and T 3 , but serum TSH was increased. It is concluded that blocking the T 3 miocardial production by AM may not be the cause of the antiarrhythmic effect of the drug. (M.E.L.) [es

Thyroid iodine content measured by x-ray fluorescence in amiodarone-induced thyrotoxicosis: concise communication

International Nuclear Information System (INIS)

Leger, A.F.; Fragu, P.; Rougier, P.; Laurent, M.F.; Tubiana, M.; Savole, J.C.

1983-01-01

Iodine-induced thyrotoxicosis (IiT) is characterized by (a) a low radioiodine uptake, increased by exogenous TSH, and (b) a spontaneous evolution towards cure within a few months. An hypothetical pathogenesis of IiT is an initial inflation in the stores of thyroid hormones during iodine excess, followed by their sudden discharge into the circulation. Thyroid iodine content was measured by fluorescent scanning in 10 patients with amiodarone-induced thyrotoxicosis and in various control groups. Results were found to be high at the onset of the disease and to decrease during its course. The data agree with the hypothetical pathogenesis. Furthermore they may permit exclusion of a painless subacute thyroiditis, which is the main differential diagnosis of IiT

Tumor necrosis factor-alpha potentiates the cytotoxicity of amiodarone in Hepa1c1c7 cells: roles of caspase activation and oxidative stress.

Science.gov (United States)

Lu, Jingtao; Miyakawa, Kazuhisa; Roth, Robert A; Ganey, Patricia E

2013-01-01

Amiodarone (AMD), a class III antiarrhythmic drug, causes idiosyncratic hepatotoxicity in human patients. We demonstrated previously that tumor necrosis factor-alpha (TNF-α) plays an important role in a rat model of AMD-induced hepatotoxicity under inflammatory stress. In this study, we developed a model in vitro to study the roles of caspase activation and oxidative stress in TNF potentiation of AMD cytotoxicity. AMD caused cell death in Hepa1c1c7 cells, and TNF cotreatment potentiated its toxicity. Activation of caspases 9 and 3/7 was observed in AMD/TNF-cotreated cells, and caspase inhibitors provided minor protection from cytotoxicity. Intracellular reactive oxygen species (ROS) generation and lipid peroxidation were observed after treatment with AMD and were further elevated by TNF cotreatment. Adding water-soluble antioxidants (trolox, N-acetylcysteine, glutathione, or ascorbate) produced only minor attenuation of AMD/TNF-induced cytotoxicity and did not influence the effect of AMD alone. On the other hand, α-tocopherol (TOCO), which reduced lipid peroxidation and ROS generation, prevented AMD toxicity and caused pronounced reduction in cytotoxicity from AMD/TNF cotreatment. α-TOCO plus a pancaspase inhibitor completely abolished AMD/TNF-induced cytotoxicity. In summary, activation of caspases and oxidative stress were observed after AMD/TNF cotreatment, and caspase inhibitors and a lipid-soluble free-radical scavenger attenuated AMD/TNF-induced cytotoxicity.

Interaction between amiodarone and hepatitis-C virus nucleotide inhibitors in human induced pluripotent stem cell-derived cardiomyocytes and HEK-293 Cav1.2 over-expressing cells.

Science.gov (United States)

Lagrutta, Armando; Zeng, Haoyu; Imredy, John; Balasubramanian, Bharathi; Dech, Spencer; Lis, Edward; Wang, Jixin; Zhai, Jin; DeGeorge, Joseph; Sannajust, Frederick

2016-10-01

Several clinical cases of severe bradyarrhythmias have been reported upon co-administration of the Hepatitis-C NS5B Nucleotide Polymerase Inhibitor (HCV-NI) direct-acting antiviral agent, sofosbuvir (SOF), and the Class-III anti-arrhythmic amiodarone (AMIO). We model the cardiac drug-drug interaction (DDI) between AMIO and SOF, and between AMIO and a closely-related SOF analog, MNI-1 (Merck Nucleotide Inhibitor #1), in functional assays of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), to provide mechanistic insights into recently reported clinical cases. AMIO co-applied with SOF or MNI-1 increased beating rate or field potential (FP) rate and decreased impedance (IMP) and Ca(2+) transient amplitudes in hiPSC-CM syncytia. This action resembled that of Ca(2+) channel blockers (CCBs) in the model, but CCBs did not substitute for AMIO in the DDI. AMIO analog dronedarone (DRON) did not substitute for, but competed with AMIO in the DDI. Ryanodine and thapsigargin, decreasing intracellular Ca(2+) stores, and SEA-0400, a Na(+)/Ca(2+) exchanger-1 (NCX1) inhibitor, partially antagonized or suppressed DDI effects. Other agents affecting FP rate only exerted additive or subtractive effects, commensurate with their individual effects. We also describe an interaction between AMIO and MNI-1 on Cav1.2 ion channels in an over-expressing HEK-293 cell line. MNI-1 enhanced Cav1.2 channel inhibition by AMIO, but did not affect inhibition of Cav1.2 by DRON, verapamil, nifedipine, or diltiazem. Our data in hiPSC-CMs indicate that HCV-NI agents such as SOF and MNI-1 interact with key intracellular Ca(2+)-handling mechanisms. Additional study in a Cav1.2 HEK-293 cell-line suggests that HCV-NIs potentiate the inhibitory action of AMIO on L-type Ca(2+) channels. Copyright © 2016 Elsevier Inc. All rights reserved.

[Population pressure: a factor of political destabilization].

Science.gov (United States)

Tallon, F

1993-04-01

Political stability throughout the world appears to be greater in countries with slowly growing populations than in those with rapid growth. Population is not the only influence on political stability, however. The relationship between political stability and development is strong. The rich countries with the slowest growth are the most stable, while poor developing countries with rapid growth suffer from chronic instability. Demographic pressure and density are not the same thing and must be distinguished. A fragile environment like that of the Sahel will experience demographic pressure despite low density. Japan has a greater population density than Rwanda and little cultivable land, but the population has a high standard of living. demographic pressure is not comparable in Japan and Rwanda because Japan has slow population growth and stable democratic political institutions. The rate of growth seems to be a more important element in destabilization than density. Rapid growth creates enormous political tensions especially when profound ethnic divisions exist, and it complicates problems of government by encouraging rapid urbanization. The unbalanced age structures resulting from rapid growth hinder the satisfaction of employment, educational, and health care needs for the ever-increasing masses of young people. 49% of Rwanda's population is under 15 and 66% is under 25. Rwanda is already densely populated, with around 300 inhabitants/sq km, and its population is projected to double in 20 years. 95% of the population is dependent on agriculture, but by 1988 the average landholding per family was only 1.25 hectares and 58% of families did not grown sufficient food for household needs. Further reduction in the size of holdings or a growing landless population will have multiple consequences. Urban migration will inevitably increase, bringing with it all the problems so evident in other poor countries where the process is more advanced than in Rwanda. Chaotic

Amiodarone-induced secondary thyroid dysfunctions in children

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Larisa A. Balykova

2017-09-01

Full Text Available Introduction: The choice of effective and safe antiarrhytmic therapy for children and adolescents is relevant issue for public health. The difficulty in choising therpeutic tactics is caused by the variety of formation of arrhythmias and the side effects of drugs. Materials and Methods: The condition of thyroid system in 45 children (20 girls, 25 boys suffering from disturbances of a rhythm within a year after the end of treatment by Amiodaronum is analyzed. The average age of the surveyed patients was 8,26 ± 0,9 years. A comprehensive examination including an assessment of a hormonal profile (a thyroxin (T4, triodothyronine (T3, thyrotrophic hormone (TTG, antibodies to a thyroid peroxidase and a thyreoglobulin (AT to TPO and TG, ultrasound examination (US of a thyroid gland, a standard electrocardiography at rest (ECG and the Holter monitoring (HM before, in 3, 6 and 12 months of therapy was conducted. Results: It has been established that prescription of Amiodarone was followed by changes in the level the thyroid’s hormones, but in most cases within normal values. Thyroidopathya (subclinical are more often were diagnosed for three patients. In 4.4 % of cases the hypothyroid and in 2.2 % of cases the thyrotoxicosis were detected. Discussion and Conclusions: It was shown that reception of medicine resulted in changes the sizes of a thyroid gland, but rarely followed by violations of functions.

Slurry Erosion Behavior of Destabilized and Deep Cryogenically Treated Cr-Mn-Cu White Cast Irons

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S. Gupta

2016-12-01

Full Text Available The effects of destabilization treatment and destabilization followed by cryogenic treatment have been evaluated on the microstructural evolution and sand-water slurry erosion behavior of Cr-Mn-Cu white cast irons. The phase transformations after the destabilization and cryotreatment have been characterized by bulk hardness measurement, optical and scanning electron microscopy, x-ray diffraction analysis. The static corrosion rate has been measured in tap water (with pH=7 and the erosion-corrosion behavior has been studied by slurry pot tester using sand-water slurry. The test results indicate that the cryogenic treatment has a significant effect in minimizing the as-cast retained austenite content and transforming into martensitic and bainitic matrix embedded with ultra-fine M7C3 alloy carbides. In contrast, by conventional destabilization treatment retained austenite in the matrix are not fully eliminated. The slurry erosive wear resistance has been compared with reference to destabilized and cryotreated high chromium iron samples which are commonly employed for such applications. The cryotreated Cr-Mn-Cu irons have exhibited a comparable erosive wear performance to those of high chromium irons. Higher hardness combined with improved corrosion resistance result in better slurry erosion resistance.

Destabilization of a peeling-ballooning mode by a toroidal rotation in tokamaks

International Nuclear Information System (INIS)

Aiba, N.; Hirota, M.; Tokuda, S.; Furukawa, M.

2009-01-01

Full text: From the viewpoint of the heat load on the divertor, Type-I edge localized mode (ELM) needs to be suppressed or the amplitude of this ELM needs to be reduced. In JT-60U, some experimental results showed that the ELM frequency depends on the toroidal rotation, and the rapid rotation in the counter direction of the plasma current changes from Type-I ELM to Grassy ELM, whose frequency is high and the amplitude is small. Recent experimental and theoretical/numerical studies in a static system have identified that both Type-I and Grassy ELMs are considered ideal magnetohydrodynamic (MHD) modes destabilizing near the plasma surface, called peeling-ballooning modes. To investigate the mechanism of the change of ELM frequency by a toroidal rotation, theoretical and numerical analyses are important for understanding the toroidal rotation effects on the peeling-ballooning mode. Previous works about the toroidal rotation effect on the edge MHD stability have illustrated that the toroidal rotation with shear can destabilize low/intermediate-n (<50) modes but can stabilize high-n modes, where n is the toroidal mode number. The stabilization of the high-n mode can be understood qualitatively in analogy with the infinite-n ballooning mode case. However, the destabilizing mechanism of the low/intermediate-n mode is not still clarified, and to understand the stability property related to ELM suppression/mitigation, it is important to clarify this destabilizing mechanism. In this paper, we investigate numerically the destabilizing effect of a toroidal rotation on the peeling-ballooning mode with a newly developed code MINERVA, which solves the Frieman-Rotenberg equation. Particularly, we pay attention to the effect of the centrifuged force on not only equilibrium but also change of equation of motion. (author)

Antiarrhythmic Drugs for Nonshockable-Turned-Shockable Out-of-Hospital Cardiac Arrest: The ALPS Study (Amiodarone, Lidocaine, or Placebo).

Science.gov (United States)

Kudenchuk, Peter J; Leroux, Brian G; Daya, Mohamud; Rea, Thomas; Vaillancourt, Christian; Morrison, Laurie J; Callaway, Clifton W; Christenson, James; Ornato, Joseph P; Dunford, James V; Wittwer, Lynn; Weisfeldt, Myron L; Aufderheide, Tom P; Vilke, Gary M; Idris, Ahamed H; Stiell, Ian G; Colella, M Riccardo; Kayea, Tami; Egan, Debra; Desvigne-Nickens, Patrice; Gray, Pamela; Gray, Randal; Straight, Ron; Dorian, Paul

2017-11-28

Out-of-hospital cardiac arrest (OHCA) commonly presents with nonshockable rhythms (asystole and pulseless electric activity). It is unknown whether antiarrhythmic drugs are safe and effective when nonshockable rhythms evolve to shockable rhythms (ventricular fibrillation/pulseless ventricular tachycardia [VF/VT]) during resuscitation. Adults with nontraumatic OHCA, vascular access, and VF/VT anytime after ≥1 shock(s) were prospectively randomized, double-blind, to receive amiodarone, lidocaine, or placebo by paramedics. Patients presenting with initial shock-refractory VF/VT were previously reported. The current study was a prespecified analysis in a separate cohort that initially presented with nonshockable OHCA and was randomized on subsequently developing shock-refractory VF/VT. The primary outcome was survival to hospital discharge. Secondary outcomes included discharge functional status and adverse drug-related effects. Of 37 889 patients with OHCA, 3026 with initial VF/VT and 1063 with initial nonshockable-turned-shockable rhythms were treatment-eligible, were randomized, and received their assigned drug. Baseline characteristics among patients with nonshockable-turned-shockable rhythms were balanced across treatment arms, except that recipients of a placebo included fewer men and were less likely to receive bystander cardiopulmonary resuscitation. Active-drug recipients in this cohort required fewer shocks, supplemental doses of their assigned drug, and ancillary antiarrhythmic drugs than recipients of a placebo ( P discharge ( P =0.24). No significant interaction between treatment assignment and discharge survival occurred with the initiating OHCA rhythm (asystole, pulseless electric activity, or VF/VT). Survival in each of these categories was consistently higher with active drugs, although the trends were not statistically significant. Adjusted absolute differences (95% confidence interval) in survival from nonshockable-turned-shockable arrhythmias

Acquisition of a novel eleven amino acid insertion directly N-terminal to a tetrabasic cleavage site confers intracellular cleavage of an H7N7 influenza virus hemagglutinin

International Nuclear Information System (INIS)

Hamilton, Brian S.; Sun, Xiangjie; Chung, Changik; Whittaker, Gary R.

2012-01-01

A critical feature of highly pathogenic avian influenza viruses (H5N1 and H7N7) is the efficient intracellular cleavage of the hemagglutinin (HA) protein. H7N7 viruses also exist in equine species, and a unique feature of the equine H7N7 HA is the presence of an eleven amino acid insertion directly N-terminal to a tetrabasic cleavage site. Here, we show that three histidine residues within the unique insertion of the equine H7N7 HA are essential for intracellular cleavage. An asparagine residue within the insertion-derived glycosylation site was also found to be essential for intracellular cleavage. The presence of the histidine residues also appear to be involved in triggering fusion, since mutation of the histidine residues resulted in a destabilizing effect. Importantly, the addition of a tetrabasic site and the eleven amino acid insertion conferred efficient intracellular cleavage to the HA of an H7N3 low pathogenicity avian influenza virus. Our studies show that acquisition of the eleven amino acid insertion offers an alternative mechanism for intracellular cleavage of influenza HA.

Acquisition of a novel eleven amino acid insertion directly N-terminal to a tetrabasic cleavage site confers intracellular cleavage of an H7N7 influenza virus hemagglutinin

Energy Technology Data Exchange (ETDEWEB)

Hamilton, Brian S.; Sun, Xiangjie; Chung, Changik [Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca NY 14853 (United States); New York Center of Excellence for Influenza Research and Surveillance, University of Rochester Medical Center, Rochester NY 14627 (United States); Whittaker, Gary R., E-mail: grw7@cornell.edu [Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca NY 14853 (United States); New York Center of Excellence for Influenza Research and Surveillance, University of Rochester Medical Center, Rochester NY 14627 (United States)

2012-12-05

A critical feature of highly pathogenic avian influenza viruses (H5N1 and H7N7) is the efficient intracellular cleavage of the hemagglutinin (HA) protein. H7N7 viruses also exist in equine species, and a unique feature of the equine H7N7 HA is the presence of an eleven amino acid insertion directly N-terminal to a tetrabasic cleavage site. Here, we show that three histidine residues within the unique insertion of the equine H7N7 HA are essential for intracellular cleavage. An asparagine residue within the insertion-derived glycosylation site was also found to be essential for intracellular cleavage. The presence of the histidine residues also appear to be involved in triggering fusion, since mutation of the histidine residues resulted in a destabilizing effect. Importantly, the addition of a tetrabasic site and the eleven amino acid insertion conferred efficient intracellular cleavage to the HA of an H7N3 low pathogenicity avian influenza virus. Our studies show that acquisition of the eleven amino acid insertion offers an alternative mechanism for intracellular cleavage of influenza HA.

A density functional study of backbone structures of polydiacetylene: destabilization of butatriene structure

International Nuclear Information System (INIS)

Katagiri, Hideki; Shimoi, Yukihiro; Abe, Shuji

2004-01-01

Backbone structures of polydiacetylene are studied with first-principles electronic structure method using plane-waves within generalized gradient approximation (GGA) of density functional theory. In spin-restricted calculations a coarse k-point sampling gives a potential energy curve with two local minima corresponding to acetylene and butatriene structures. However, the potential barrier between the two structures rapidly decreases with increasing number of k-points, which results in destabilization of the butatriene structure. Spin polarization effects also destabilize the butatriene structure, inducing atom-centered spin-density-wave state. These potential energies were compared with those obtained by Hartree-Fock, density functional within local density approximation (LDA) and GGA, and hybrid density functional methods using a gaussian basis set. The comparison shows that the density functional methods within LDA and GGA favor the destabilization of the butatriene structure in contrast to the Hartree-Fock method

Mechanisms governing the reactivation-dependent destabilization of memories and their role in extinction

OpenAIRE

Flavell, Charlotte R.; Lambert, Elliot A.; Winters, Boyer D.; Bredy, Timothy W.

2013-01-01

The extinction of learned associations has traditionally been considered to involve new learning, which competes with the original memory for control over behaviour. However, a recent resurgence of interest in reactivation-dependent amnesia has revealed that the retrieval of fear-related memory (with what is essentially a brief extinction session) can result in it’s destabilization. This review discusses some of the cellular and molecular mechanisms that are involved in the destabilization of...

Mathematical modeling of atherosclerotic plaque destabilization: Role of neovascularization and intraplaque hemorrhage.

Science.gov (United States)

Guo, Muyi; Cai, Yan; Yao, Xinke; Li, Zhiyong

2018-08-07

Observational studies have identified angiogenesis from the adventitial vasa vasorum and intraplaque hemorrhage (IPH) as critical factors in atherosclerotic plaque progression and destabilization. Here we propose a mathematical model incorporating intraplaque neovascularization and hemodynamic calculation with plaque destabilization for the quantitative evaluation of the role of neoangiogenesis and IPH in the vulnerable atherosclerotic plaque formation. An angiogenic microvasculature is generated by two-dimensional nine-point discretization of endothelial cell proliferation and migration from the vasa vasorum. Three key cells (endothelial cells, smooth muscle cells and macrophages) and three key chemicals (vascular endothelial growth factors, extracellular matrix and matrix metalloproteinase) are involved in the plaque progression model, and described by the reaction-diffusion partial differential equations. The hemodynamic calculation of the microcirculation on the generated microvessel network is carried out by coupling the intravascular, interstitial and transvascular flow. The plasma concentration in the interstitial domain is defined as the description of IPH area according to the diffusion and convection with the interstitial fluid flow, as well as the extravascular movement across the leaky vessel wall. The simulation results demonstrate a series of pathophysiological phenomena during the vulnerable progression of an atherosclerotic plaque, including the expanding necrotic core, the exacerbated inflammation, the high microvessel density (MVD) region at the shoulder areas, the transvascular flow through the capillary wall and the IPH. The important role of IPH in the plaque destabilization is evidenced by simulations with varied model parameters. It is found that the IPH can significantly speed up the plaque vulnerability by increasing necrotic core and thinning fibrous cap. In addition, the decreased MVD and vessel permeability may slow down the process of

Intracellular Drug Uptake-A Comparison of Single Cell Measurements Using ToF-SIMS Imaging and Quantification from Cell Populations with LC/MS/MS.

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Newman, Carla F; Havelund, Rasmus; Passarelli, Melissa K; Marshall, Peter S; Francis, Ian; West, Andy; Alexander, Morgan R; Gilmore, Ian S; Dollery, Colin T

2017-11-21

ToF-SIMS is a label-free imaging method that has been shown to enable imaging of amiodarone in single rat macrophage (NR8383) cells. In this study, we show that the method extends to three other cell lines relevant to drug discovery: human embryonic kidney (HEK293), cervical cancer (HeLa), and liver cancer (HepG2). There is significant interest in the variation of drug uptake at the single cell level, and we use ToF-SIMS to show that there is great diversity between individual cells and when comparing each of the cell types. These single cell measurements are compared to quantitative measurements of cell-associated amiodarone for the population using LC/MS/MS and cell counting with flow cytometry. NR8383 and HepG2 cells uptake the greatest amount of amiodarone with an average of 2.38 and 2.60 pg per cell, respectively, and HeLa and Hek 293 have a significantly lower amount of amiodarone at 0.43 and 0.36 pg per cell, respectively. The amount of cell-associated drug for the ensemble population measurement (LC/MS/MS) is compared with the ToF-SIMS single cell data: a similar amount of drug was detected per cell for the NR8383, and HepG2 cells at a greater level than that for the HEK293 cells. However, the two techniques did not agree for the HeLa cells, and we postulate potential reasons for this.

DNA-Destabilizing Agents as an Alternative Approach for Targeting DNA: Mechanisms of Action and Cellular Consequences

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Gaëlle Lenglet

2010-01-01

Full Text Available DNA targeting drugs represent a large proportion of the actual anticancer drug pharmacopeia, both in terms of drug brands and prescription volumes. Small DNA-interacting molecules share the ability of certain proteins to change the DNA helix's overall organization and geometrical orientation via tilt, roll, twist, slip, and flip effects. In this ocean of DNA-interacting compounds, most stabilize both DNA strands and very few display helix-destabilizing properties. These types of DNA-destabilizing effect are observed with certain mono- or bis-intercalators and DNA alkylating agents (some of which have been or are being developed as cancer drugs. The formation of locally destabilized DNA portions could interfere with protein/DNA recognition and potentially affect several crucial cellular processes, such as DNA repair, replication, and transcription. The present paper describes the molecular basis of DNA destabilization, the cellular impact on protein recognition, and DNA repair processes and the latter's relationships with antitumour efficacy.

Application of X-ray microanalysis to the study of drug uptake in cell culture

International Nuclear Information System (INIS)

Reasor, M.J.; Lee, P.; Kirk, R.G.

1990-01-01

X-ray microanalysis has been used previously to study the accumulation of iodine in alveolar macrophages of rats treated with the iodinated drug, amiodarone. Due to metabolism of the drug in vivo, primarily to desethylamiodarone, it was not possible to identify the source of the iodine signal. In the present study we have utilized primary cell cultures of alveolar macrophages to study the intracellular accumulation of each of these drug species in vitro. Neither drug is metabolized by these cells in culture, permitting characterization of the accumulation of each independent of the other. Cells were incubated with equimolar concentrations of either amiodarone or desethylamiodarone for 42 hr, and X-ray microanalysis of freeze-dried cryosections of cells was used to quantify accumulation by monitoring the iodine signal associated with each drug. For both drug exposures, the highest iodine content was present in amorphous bodies and dense granules, consistent with the pattern following in vivo exposure. Higher levels of desethylamiodarone, compared to amiodarone, were measured in all compartments of the cells. The results of the in vitro investigation further demonstrate the utility of X-ray microanalysis in the study of the cellular response to amiodarone and desethylamiodarone

THE ESTIMATION OF HUMAN-OPERATOR CYBERNETIC ABILITIES DURING THE IMPACT OF DESTABILIZING FACTORS OF EXTERNAL ENVIRONMENT

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Sergii T. Polishchuk

2009-04-01

Full Text Available  The method of estimation of human-operator cybernetics abilities during of the impacting of destabilizing factors of external environment is suggested. It was proved that up-to-date biomedical approach for periodical health examination of pilots in civil aviation isn’t guarantees theirs cybernetics abilities in cases of influence of destabilizing factors.

E-cadherin destabilization accounts for the pathogenicity of missense mutations in hereditary diffuse gastric cancer.

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Joana Simões-Correia

Full Text Available E-cadherin is critical for the maintenance of tissue architecture due to its role in cell-cell adhesion. E-cadherin mutations are the genetic cause of Hereditary Diffuse Gastric Cancer (HDGC and missense mutations represent a clinical burden, due to the uncertainty of their pathogenic role. In vitro and in vivo, most mutations lead to loss-of-function, although the causal factor is unknown for the majority. We hypothesized that destabilization could account for the pathogenicity of E-cadherin missense mutations in HDGC, and tested our hypothesis using in silico and in vitro tools. FoldX algorithm was used to calculate the impact of each mutation in E-cadherin native-state stability, and the analysis was complemented with evolutionary conservation, by SIFT. Interestingly, HDGC patients harbouring germline E-cadherin destabilizing mutants present a younger age at diagnosis or death, suggesting that the loss of native-state stability of E-cadherin accounts for the disease phenotype. To elucidate the biological relevance of E-cadherin destabilization in HDGC, we investigated a group of newly identified HDGC-associated mutations (E185V, S232C and L583R, of which L583R is predicted to be destabilizing. We show that this mutation is not functional in vitro, exhibits shorter half-life and is unable to mature, due to premature proteasome-dependent degradation, a phenotype reverted by stabilization with the artificial mutation L583I (structurally tolerated. Herein we report E-cadherin structural models suitable to predict the impact of the majority of cancer-associated missense mutations and we show that E-cadherin destabilization leads to loss-of-function in vitro and increased pathogenicity in vivo.

Molecular mechanisms for the destabilization and restabilization of reactivated spatial memory in the Morris water maze

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Kim Ryang

2011-02-01

Full Text Available Abstract Background Memory retrieval is not a passive process. Recent studies have shown that reactivated memory is destabilized and then restabilized through gene expression-dependent reconsolidation. Molecular studies on the regulation of memory stability after retrieval have focused almost exclusively on fear memory, especially on the restabilization process of the reactivated fear memory. We previously showed that, similarly with fear memories, reactivated spatial memory undergoes reconsolidation in the Morris water maze. However, the underlying molecular mechanisms by which reactivated spatial memory is destabilized and restabilized remain poorly understood. In this study, we investigated the molecular mechanism that regulates the stability of the reactivated spatial memory. Results We first showed that pharmacological inactivation of the N-methyl-D-aspartate glutamate receptor (NMDAR in the hippocampus or genetic inhibition of cAMP-responsible element binding protein (CREB-mediated transcription disrupted reactivated spatial memory. Finally, we showed that pharmacological inhibition of cannabinoid receptor 1 (CB1 and L-type voltage gated calcium channels (LVGCCs in the hippocampus blocked the disruption of the reactivated spatial memory by the inhibition of protein synthesis. Conclusions Our findings indicated that the reactivated spatial memory is destabilized through the activation of CB1 and LVGCCs and then restabilized through the activation of NMDAR- and CREB-mediated transcription. We also suggest that the reactivated spatial memory undergoes destabilization and restabilization in the hippocampus, through similar molecular processes as those for reactivated contextual fear memories, which require CB1 and LVGCCs for destabilization and NMDAR and CREB for restabilization.

SEASONAL VARIATION IN LYSOSOMAL DESTABILIZATION IN OYSTERS, CRASSOSTREA VIRGINICA. (R826201)

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Lysosomal destabilization assays have been used as valuable biomarkers of pollutant exposures in a variety of bivalve and fish species. The responses of oysters, Crassostrea virginica, deployed at and native to various reference and degraded sites were evaluated for lys...

Effects of 2 ankle destabilization devices on electromyography measures during functional exercises in individuals with chronic ankle instability.

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Donovan, Luke; Hart, Joseph M; Hertel, Jay

2015-03-01

Randomized crossover laboratory study. To determine the effects of ankle destabilization devices on surface electromyography (sEMG) measures of selected lower extremity muscles during functional exercises in participants with chronic ankle instability. Ankle destabilization devices are rehabilitation tools that can be worn as a boot or sandal to increase lower extremity muscle activation during walking in healthy individuals. However, they have not been tested in a population with pathology. Fifteen adults with chronic ankle instability participated. Surface electromyography electrodes were located over the anterior tibialis, fibularis longus, lateral gastrocnemius, rectus femoris, biceps femoris, and gluteus medius. The activity level of these muscles was recorded in a single testing session during unipedal stance with eyes closed, the Star Excursion Balance Test, lateral hops, and treadmill walking. Each task was performed under 3 conditions: shod, ankle destabilization boot, and ankle destabilization sandal. Surface electromyography signal amplitudes were measured for each muscle during each exercise for all 3 conditions. Participants demonstrated a significant increase, with moderate to large effect sizes, in sEMG signal amplitude of the fibularis longus in the ankle destabilization boot and ankle destabilization sandal conditions during the unipedal eyes-closed balance test, the Star Excursion Balance Test in the anterior and posteromedial directions, lateral hops, and walking, when compared to the shod condition. Both devices also resulted in an increase in sEMG signal amplitudes, with large effect sizes of the lateral gastrocnemius, rectus femoris, biceps femoris, and gluteus medius during the unipedal-stance-with-eyes-closed test, compared to the shod condition. Wearing ankle destabilization devices caused greater muscle activation during functional exercises in individuals with chronic ankle instability. Based on the magnitude of the effect, there were

Destabilization of the electron Bernstein modes by runaway electrons

International Nuclear Information System (INIS)

Hitchcock, D.A.; Mahajan, S.M.

1982-01-01

It is shown that the electromagnetic finite k/sub parallel/ electron Bernstein mode can be destabilized by the runaway electron distribution which results from the quasilinear action of the magnetized plasma oscillation. This mechanism is shown to yield growth rates of the order of 10 8 sec -1 and is suggested as a mechanism for the enchanced cyclotron harmonic emission in the presence of runaway electrons

Core localized toroidal Alfven eigenmodes destabilized by energetic ions in the CHS heliotron/torsatron

International Nuclear Information System (INIS)

Takechi, M.; Matsunaga, G.; Takagi, S.

1999-09-01

Toroidal Alfven eigenmodes (TAE) destabilized by the pressure gradient of energetic alpha particles may expel the alpha particles before thermalization. TAE is important for tokamaks, and for helical systems (stellarators) as well. In CHS (compact helical system) TAE localized in the plasma core are destabilized when the plasma current is induced by co-injection of neutral beams. The observed TAE exhibits a ballooning nature. The internal structure of TAE was measured with a soft X-ray detector. The soft X-ray fluctuations level for TAE is too low to obtain the radial profiles of fluctuation intensities. (Tanaka, M.)

Disfunção tireoidiana e conduta dos cardiologistas em pacientes usando amiodarona Thyroid dysfunction and cardiological management in patients receiving amiodarone

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Anna Gabriela Fuks

2004-06-01

Full Text Available OBJETIVO: Determinar a prevalência de disfunção tireoidiana em pacientes usando amiodarona e os possíveis fatores associados. Verificar através de questionário aplicado a cardiologistas, a importância do fármaco causar alterações na função tireoidiana. MÉTODO: Avaliados 56 pacientes em uso crônico (> 3 meses de amiodarona com dosagens séricas de TSH, T4 livre, T3 total e Anti-TPO e definidos como portadores de disfunção tireoidiana (DT pacientes com TSH alterado. RESULTADOS: A prevalência de disfunção tireoidiana foi de 33,9%. Não houve diferença entre este grupo e os pacientes sem disfunção, exceto em relação à prevalência de anti-TPO positivo maior nos pacientes com DT (p=0,02. Hipotireoidismo subclínico foi diagnosticado em 10 (17,9% pacientes e hipotireoidismo clínico em 6 (10,7%. A prevalência de hipertireoidismo subclínico foi de 3,6% e de hipertireoidismo clínico de 1,8%. Anticorpos anti-TPO foram positivos em 5 (8% pacientes (dos quais 4 apresentavam disfunção. Quando comparados aos doentes sem anti-TPO positivo este grupo teve maior prevalência de disfunção (80% vs 29,4%; p=0,04. Verificado que apenas 49,2% dos cardiologistas faziam acompanhamento da função tireoidiana rotineiramente e a prevalência de disfunção referida na experiência da maioria era de 1 a 10%. CONCLUSÃO: A prevalência de disfunção tireoidiana na nossa população foi elevada, mostrando a necessidade de implementação de uma rotina laboratorial. Houve grande divergência entre os cardiologistas em relação ao tipo de acompanhamento utilizado nos pacientes em uso de amiodarona.OBJECTIVE: To determine the prevalence of thyroid dysfunction in patients receiving amiodarone, and the possible associated factors. The study also aimed at assessing the effect of amiodarone on thyroid function through the application of a questionnaire to cardiologists. METHOD: Fifty-six patients chronically (> 3 months receiving amiodarone were

Stabilization and destabilization effects of the electric field on stochastic precipitate pattern

NARCIS (Netherlands)

Lagzi, István; Izsak, F.

2004-01-01

Stabilization and destabilization effects of an applied electric field on the Liesegang pattern formation in low concentration gradient were studied with numerical model simulations. In the absence of an electric field pattern formation exhibits increasingly stochastic behaviour as the initial

Intracellular Environment-Responsive Stabilization of Polymer Vesicles Formed from Head-Tail Type Polycations Composed of a Polyamidoamine Dendron and Poly(L-lysine

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Kenji Kono

2013-09-01

Full Text Available For the development of effective drug carriers, nanocapsules that respond to micro-environmental changes including a decrease in pH and a reductive environment were prepared by the stabilization of polymer vesicles formed from head-tail type polycations, composed of a polyamidoamine dendron head and a poly(L-lysine tail (PAMAM dendron-PLL, through the introduction of disulfide bonds between the PLL tails. Disulfide bonds were successfully introduced through the reaction of Lys residues in the PAMAM dendron-PLL polymer vesicles with 2-iminothiolane. The stabilization of PAMAM dendron-PLL polymer vesicles was confirmed by dynamic light scattering measurements. In acid-base titration experiments, nanocapsules cross-linked by disulfide bonds had a buffering effect during the cellular uptake process. The PAMAM dendron-PLL nanocapsules were used to incorporate the fluorescent dyes rhodamine 6G and fluorescein as a drug model. Cationic rhodamine 6G was generally not released from the nanocapsules because of the electrostatic barrier of the PLL membrane. However, the nanocapsules were destabilized at high glutathione concentrations corresponding to intracellular concentrations. Rhodamine 6G was immediately released from the nanocapsules because of destabilization upon the cleavage of disulfide bonds. This release of rhodamine 6G from the nanocapsules was also observed in HeLa cells by laser confocal microscopy.

Survivin counteracts the therapeutic effect of microtubule de-stabilizers by stabilizing tubulin polymers

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Hsieh Hsing-Pang

2009-07-01

Full Text Available Abstract Background Survivin is a dual function protein. It inhibits the apoptosis of cells by inhibiting caspases, and also promotes cell growth by stabilizing microtubules during mitosis. Over-expression of survivin has been demonstrated to induce drug-resistance to various chemo-therapeutic agents such as cisplatin (DNA damaging agent and paclitaxel (microtubule stabilizer in cancers. However, survivin-induced resistance to microtubule de-stabilizers such as Vinca alkaloids and Combretastatin A-4 (CA-4-related compounds were seldom demonstrated in the past. Furthermore, the question remains as to whether survivin plays a dominant role in processing cytokinesis or inhibiting caspases activity in cells treated with anti-mitotic compounds. The purpose of this study is to evaluate the effect of survivin on the resistance and susceptibility of human cancer cells to microtubule de-stabilizer-induced cell death. Results BPR0L075 is a CA-4 analog that induces microtubule de-polymerization and subsequent caspase-dependent apoptosis. To study the relationship between the expression of survivin and the resistance to microtubule de-stabilizers, a KB-derived BPR0L075-resistant cancer cell line, KB-L30, was generated for this study. Here, we found that survivin was over-expressed in the KB-L30 cells. Down-regulation of survivin by siRNA induced hyper-sensitivity to BPR0L075 in KB cells and partially re-stored sensitivity to BPR0L075 in KB-L30 cells. Western blot analysis revealed that down-regulation of survivin induced microtubule de-stabilization in both KB and KB-L30 cells. However, the same treatment did not enhance the down-stream caspase-3/-7 activities in BPR0L075-treated KB cells. Translocation of a caspase-independent apoptosis-related molecule, apoptosis-inducing factor (AIF, from cytoplasm to the nucleus was observed in survivin-targeted KB cells under BPR0L075 treatment. Conclusion In this study, survivin plays an important role in the

Snowball Earth termination by destabilization of equatorial permafrost methane clathrate.

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Kennedy, Martin; Mrofka, David; von der Borch, Chris

2008-05-29

The start of the Ediacaran period is defined by one of the most severe climate change events recorded in Earth history--the recovery from the Marinoan 'snowball' ice age, approximately 635 Myr ago (ref. 1). Marinoan glacial-marine deposits occur at equatorial palaeolatitudes, and are sharply overlain by a thin interval of carbonate that preserves marine carbon and sulphur isotopic excursions of about -5 and +15 parts per thousand, respectively; these deposits are thought to record widespread oceanic carbonate precipitation during postglacial sea level rise. This abrupt transition records a climate system in profound disequilibrium and contrasts sharply with the cyclical stratigraphic signal imparted by the balanced feedbacks modulating Phanerozoic deglaciation. Hypotheses accounting for the abruptness of deglaciation include ice albedo feedback, deep-ocean out-gassing during post-glacial oceanic overturn or methane hydrate destabilization. Here we report the broadest range of oxygen isotope values yet measured in marine sediments (-25 per thousand to +12 per thousand) in methane seeps in Marinoan deglacial sediments underlying the cap carbonate. This range of values is likely to be the result of mixing between ice-sheet-derived meteoric waters and clathrate-derived fluids during the flushing and destabilization of a clathrate field by glacial meltwater. The equatorial palaeolatitude implies a highly volatile shelf permafrost pool that is an order of magnitude larger than that of the present day. A pool of this size could have provided a massive biogeochemical feedback capable of triggering deglaciation and accounting for the global postglacial marine carbon and sulphur isotopic excursions, abrupt unidirectional warming, cap carbonate deposition, and a marine oxygen crisis. Our findings suggest that methane released from low-latitude permafrost clathrates therefore acted as a trigger and/or strong positive feedback for deglaciation and warming. Methane hydrate

Photo-triggered destabilization of nanoscopic vehicles by dihydroindolizine for enhanced anticancer drug delivery in cervical carcinoma.

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Singh, Priya; Choudhury, Susobhan; Kulanthaivel, Senthilguru; Bagchi, Damayanti; Banerjee, Indranil; Ahmed, Saleh A; Pal, Samir Kumar

2018-02-01

The efficacy and toxicity of drugs depend not only on their potency but also on their ability to reach the target sites in preference to non-target sites. In this regards destabilization of delivery vehicles induced by light can be an effective strategy for enhancing drug delivery with spatial and temporal control. Herein we demonstrate that the photoinduced isomerization from closed (hydrophobic) to open isomeric form (hydrophilic) of a novel DHI encapsulated in liposome leads to potential light-controlled drug delivery vehicles. We have used steady state and picosecond resolved dynamics of a drug 8-anilino-1-naphthalenesulfonic acid ammonium salt (ANS) incorporated in liposome to monitor the efficacy of destabilization of liposome in absence and presence UVA irradiation. Steady state and picosecond resolved polarization gated spectroscopy including the well-known strategy of solvation dynamics and Förster resonance energy transfer; reveal the possible mechanism out of various phenomena involved in destabilization of liposome. We have also investigated the therapeutic efficacy of doxorubicin (DOX) delivery from liposome to cervical cancer cell line HeLa. The FACS, confocal fluorescence microscopic and MTT assay studies reveal an enhanced cellular uptake of DOX leading to significant reduction in cell viability (∼40%) of HeLa followed by photoresponsive destabilization of liposome. Our studies successfully demonstrate that these DHI encapsulated liposomes have potential application as a smart photosensitive drug delivery system. Copyright © 2017 Elsevier B.V. All rights reserved.

Insight into microtubule destabilization mechanism of 3,4,5-trimethoxyphenyl indanone derivatives using molecular dynamics simulation and conformational modes analysis

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Tripathi, Shubhandra; Srivastava, Gaurava; Singh, Aastha; Prakasham, A. P.; Negi, Arvind S.; Sharma, Ashok

2018-03-01

Colchicine site inhibitors are microtubule destabilizers having promising role in cancer therapeutics. In the current study, four such indanone derivatives (t1, t9, t14 and t17) with 3,4,5-trimethoxyphenyl fragment (ring A) and showing significant microtubule destabilization property have been explored. The interaction mechanism and conformational modes triggered by binding of these indanone derivatives and combretastatin at colchicine binding site (CBS) of αβ-tubulin dimer were studied using molecular dynamics (MD) simulation, principle component analysis and free energy landscape analysis. In the MD results, t1 showed binding similar to colchicine interacting in the deep hydrophobic core at the CBS. While t9, t14 and t17 showed binding conformation similar to combretastatin, with ring A superficially binding at the CBS. Results demonstrated that ring A played a vital role in binding via hydrophobic interactions and got anchored between the S8 and S9 sheets, H8 helix and T7 loop at the CBS. Conformational modes study revealed that twisting and bending conformational motions (as found in the apo system) were nearly absent in the ligand bound systems. Absence of twisting motion might causes loss of lateral contacts in microtubule, thus promoting microtubule destabilization. This study provides detailed account of microtubule destabilization mechanism by indanone ligands and combretastatin, and would be helpful for designing microtubule destabilizers with higher activity.

Combined etiology of anaphylactic cardiogenic shock: Amiodarone, epinephrine, cardioverter defibrillator, left ventricular assist devices and the Kounis syndrome

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Nicholas G Kounis

2015-01-01

Full Text Available Anaphylactic shock is a life-threatening condition which needs detailed and mediculous clinical assessment and thoughtful treatment. Several causes can join forces in order to degranulate mast cells. Amiodarone which is an iodine-containing highly lipophilic benzofuran can induce allergic reactions and anaphylactic shock in sensitized patients. Epinephrine is a life saving drug, but in sulfite allergic patients it should be given with caution due its metabisulfite preservative. Metals covering cardiac defibrillators and pacemakers can act as antigens attached to serum proteins and induce allergic reactions. In anaphylactic shock, myocardial involvement due to vasospasm-induced coronary blood flow reduction manifesting as Kounis syndrome should be always considered. Clinically, combined treatment targeting the primary cause of anaphylaxis together with protection of cardiac tissue seems to be of paramount importance.

Lysophosphatidic acid triggers mast cell-driven atherosclerotic plaque destabilization by increasing vascular inflammation.

NARCIS (Netherlands)

Bot, M.; , van, Berkel T.J.C.

2013-01-01

Lysophosphatidic acid (LPA), a bioactive lysophospholipid, accumulates in the atherosclerotic plaque. It has the capacity to activate mast cells, which potentially exacerbates plaque progression. In this study, we thus aimed to investigate whether LPA contributes to plaque destabilization by

A tomografia computadorizada de alta resolução na avaliação da toxicidade pulmonar por amiodarona High-resolution computed tomography of amiodarone pulmonary toxicity

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Daniela Peixoto Consídera

2006-04-01

Full Text Available OBJETIVO: Avaliar as principais alterações identificadas na tomografia computadorizada de alta resolução do tórax em pacientes com toxicidade pulmonar pela amiodarona. MATERIAIS E MÉTODOS: Foram avaliadas dez tomografias computadorizadas de alta resolução de tórax de pacientes com pneumonite pela amiodarona, seis desses pacientes do sexo masculino e quatro do sexo feminino, com idade média de 73,5 anos. RESULTADOS: Os achados tomográficos mais relevantes foram opacidades lineares ou reticulares em seis casos (60%, pequenos nódulos com densidade elevada em seis casos (60%, consolidações densas em três casos (30% e aumento da densidade do parênquima hepático em cinco de oito casos em que havia estudo tomográfico do abdome superior (62,5%. CONCLUSÃO: A tomografia computadorizada de alta resolução é um exame importante na avaliação de pacientes com toxicidade pulmonar pela amiodarona, devendo ser realizada sempre que houver suspeita deste diagnóstico. O achado de espessamento de septos interlobulares associado a lesões com aumento de densidade é altamente sugestivo deste diagnóstico.OBJECTIVE: To evaluate the main findings of chest high-resolution computed tomography in patients with amiodarone pulmonary toxicity. MATERIALS AND METHODS: Ten patients - six male and four female, average age of 73.5 years - with amiodarone-induced pneumonitis have undergone chest high-resolution computed tomography. RESULTS: The most relevant tomographic findings were linear or reticular opacities in six cases (60%, small high density nodules in six cases (60%, dense consolidations in three cases (30% and increased density in the hepatic parenchyma in five of eight cases in which there was a superior abdomen CT scan (62.5%. CONCLUSION: The high-resolution computed tomography is a valuable non-invasive test for evaluating patients with amiodarone pulmonary toxicity and should always be performed when one suspects of the presence of this

Emprego de altas doses de amiodarona via oral na reversão da fibrilação atrial no pós-operatório de cirurgia cardíaca High dose amiodarone for the reversion of atrial fibrillation during the postoperative period of cardiac surgery

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João Carlos Vieira da Costa Guaragna

1997-12-01

Full Text Available OBJETIVO: Relatar a experiência no emprego de altas doses de amiodarona via oral (1800mg/d na reversão da fibrilação atrial (FA em pacientes submetidos à cirurgia cardíaca. MÉTODOS: Analisados, retrospectivamente, 80 pacientes que apresentaram FA no pós operatório de cirurgia cardíaca, constituindo 2 grupos: grupo A com 28 pacientes em uso de amiodarona e grupo B recebendo digital, sendo que este grupo foi subdividido no grupo C com 21 pacientes onde foi associada amiodarona, quando não houvesse reversão da arritmia em 48h. As diferenças foram consideradas significativas para um valor de PPURPOSE: To report our experience using high dose oral amiodarone (1,800mg/day for the reversion of atrial fibrillation to sinus rhythm in patients submitted to cardiac surgery. METHODS: We retrospectively analyzed the records of 80 patients who had atrial fibrillation during the postoperative period after cardiac surgery, initially divided in two groups: group A, 28 patients that used amiodarone, and group B composed of patients receiving digoxin. The latter group was divided further in a third group (C, with 21 patients in which amiodarone was associated with digoxin if there was no reversion of the arrhythmia after 48 hours of treatment. The observed differences were considered significant at P<0.05. RESULTS: Atrial fibrillation occurred in 19.4% of the patients submitted to surgery, predominating in males, 60 to 69 years-old. In group A there was reversion to sinus rhythm in 78.6% of the cases. In group B digoxin succeeded in 60%, and in group C 90% of the patients reverted to sinus rhythm. CONCLUSION: High dose oral amiodarone, alone or combined to digoxin, can be safe and effective for the treatment of atrial fibrillation after cardiac surgery.

Preparation of liposomal amiodarone and investigation of its cardiomyocyte-targeting ability in cardiac radiofrequency ablation rat model

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Zhuge Y

2016-05-01

Full Text Available Ying Zhuge,1,* Zhi-Feng Zheng,1,* Mu-Qing Xie,2 Lin Li,2 Fang Wang,1 Feng Gao2,3 1Department of Cardiology, Shanghai First People’s Hospital of Nanjing Medical University, 2Department of Pharmaceutics, School of Pharmacy, 3Shanghai Key Laboratory of Functional Materials Chemistry, East China University of Science and Technology, Shanghai, People’s Republic of China*These authors contributed equally to this workAbstract: The objective of this study was to develop an amiodarone hydrochloride (ADHC-loaded liposome (ADHC-L formulation and investigate its potential for cardiomyocyte targeting after cardiac radiofrequency ablation (CA in vivo. The ADHC-L was prepared by thin-film method combined with ultrasonication and extrusion. The preparation process was optimized by Box–Behnken design with encapsulation efficiency as the main evaluation index. The optimum formulation was quantitatively obtained with a diameter of 99.9±0.4 nm, a zeta potential of 35.1±10.9 mV, and an encapsulation efficiency of 99.5%±13.3%. Transmission electron microscopy showed that the liposomes were spherical particles with integrated bilayers and well dispersed with high colloidal stability. Pharmacokinetic studies were investigated in rats after intravenous administration, which revealed that compared with free ADHC treatment, ADHC-L treatment showed a 5.1-fold increase in the area under the plasma drug concentration–time curve over a period of 24 hours (AUC0–24 h and an 8.5-fold increase in mean residence time, suggesting that ADHC-L could facilitate drug release in a more stable and sustained manner while increasing the circulation time of ADHC, especially in the blood. Biodistribution studies of ADHC-L demonstrated that ADHC concentration in the heart was 4.1 times higher after ADHC-L treatment in CA rat model compared with ADHC-L sham-operated treatment at 20 minutes postinjection. Fluorescence imaging studies further proved that the heart

Medical expert system for assessment of coronary heart disease destabilization based on the analysis of the level of soluble vascular adhesion molecules

Science.gov (United States)

Serkova, Valentina K.; Pavlov, Sergey V.; Romanava, Valentina A.; Monastyrskiy, Yuriy I.; Ziepko, Sergey M.; Kuzminova, Nanaliya V.; Wójcik, Waldemar; DzierŻak, RóŻa; Kalizhanova, Aliya; Kashaganova, Gulzhan

2017-08-01

Theoretical and practical substantiation of the possibility of the using the level of soluble vascular adhesion molecules (sVCAM) is performed. Expert system for the assessment of coronary heart disease (CHD) destabilization on the base of the analysis of soluble vascular adhesion molecules level is developed. Correlation between the increase of VCAM level and C-reactive protein (CRP) in patients with different variants of CHD progression is established. Association of chronic nonspecific vascular inflammation activation and CHD destabilization is shown. The expedience of parallel determination of sVCAM and CRP levels for diagnostics of CHD destabilization and forecast elaboration is noted.

Destabilizing geometrical and bimaterial effects in frictional sliding

Science.gov (United States)

Aldam, M.; Bar Sinai, Y.; Svetlizky, I.; Fineberg, J.; Brener, E.; Xu, S.; Ben-Zion, Y.; Bouchbinder, E.

2017-12-01

Asymmetry of the two blocks forming a fault plane, i.e. the lack of reflection symmetry with respect to the fault plane, either geometrical or material, gives rise to generic destabilizing effects associated with the elastodynamic coupling between slip and normal stress variations. While geometric asymmetry exists in various geophysical contexts, such as thrust faults and landslide systems, its effect on fault dynamics is often overlooked. In the first part of the talk, I will show that geometrical asymmetry alone can destabilize velocity-strengthening faults, which are otherwise stable. I will further show that geometrical asymmetry accounts for a significant weakening effect observed in rupture propagation and present laboratory data that support the theory. In the second part of the talk, I will focus on material asymmetry and discuss an unexpected property of the well-studied frictional bimaterial effect. I will show that while the bimaterial coupling between slip and normal stress variations is a monotonically increasing function of the bimaterial contrast, when it is coupled to interfacial shear stress perturbations through a friction law, various physical quantities exhibit a non-monotonic dependence on the bimaterial contrast. This non-monotonicity is demonstrated for the stability of steady-sliding and for unsteady rupture propagation in faults described by various friction laws (regularized Coulomb, slip-weakening, rate-and-state friction), using analytic and numerical tools. All in all, the importance of bulk asymmetry to interfacial fault dynamics is highlighted. [1] Michael Aldam, Yohai Bar-Sinai, Ilya Svetlizky, Efim A. Brener, Jay Fineberg, and Eran Bouchbinder. Frictional Sliding without Geometrical Reflection Symmetry. Phys. Rev. X, 6(4):041023, 2016. [2] Michael Aldam, Shiqing Xu, Efim A. Brener, Yehuda Ben-Zion, and Eran Bouchbinder. Non-monotonicity of the frictional bimaterial effect. arXiv:1707.01132, 2017.

Use of mixtures containing nonionic surfactants in the destabilization of petroleum emulsions

Energy Technology Data Exchange (ETDEWEB)

Mansur, Claudia R.E.; Mauro, Aparecida C.; Aquino, Aline S.; Lechuga, Fernanda C.; Gonzalez, Gaspar; Lucas, Elizabete F. [Universidade Federal do Rio de Janeiro (UFRJ), RJ (Brazil). Inst. de Macromoleculas; Gonzalez, Gaspar [PETROBRAS, Rio de Janeiro, RJ (Brazil). Centro de Pesquisas (CENPES)

2004-07-01

During the petroleum dehydration process, it is necessary to used chemical demulsifiers in order to break the w/o emulsions that are formed in the oil field. These demulsifiers products are, in many cases, surfactants based poly(ethylene oxide-propylene oxide) block copolymers (PEO-PPO), with different EO/PO molar ratio. In this work were correlate the structure and the properties of PEO-PPO block copolymers with their performance as petroleum emulsion-destabilizing agent. Moreover, it was used an additive in the formulations, known as hydrotrope, in order to increase the solubility of these copolymers in aqueous solution. The results showed that the copolymer branched, whose hydrophilic segments (PEO and OH) are in an external adjacent position, present the higher solubility, in spite of to own EO/PO ratio similar to the others copolymers and the highest molar mass. Moreover, this copolymer presented the best efficiency in the emulsion destabilization. The addition of the hydrotrope NaBMGS to the PEO-PPO copolymers aqueous solutions caused the solubility increasing of these compounds in water. Such additive being used in the demulsifier formulation provoked an efficiency improving on the emulsion breaking process. (author)

Effects of Amiodarone and N-Desethylamiodarone on Cardiac Voltage-gated Sodium Channels

Directory of Open Access Journals (Sweden)

Mohammad-Reza eGhovanloo

2016-03-01

Full Text Available Amiodarone (AMD is a potent antiarrhythmic drug with high efficacy for treating atrial fibrillation and tachycardia. The pharmacologic profile of AMD is complex. AMD possesses biophysical characteristics of all of class I, II, III, and IV agents. Despite its adverse side effects, AMD remains the most commonly prescribed antiarrhythmic drug. AMD was described to prolong the QT interval and can lead to torsades de pointes. Our goal was to study the effects of AMD on peak and late sodium currents (INa,P and INa,L and determine whether these effects change as AMD is metabolized into N-Desethylamiodarone (DES. We hypothesized that AMD and DES block both INa,P and INa,L with similar profiles due to structural similarities. Given the inherent small amounts of INa,L in NaV1.5, we screened AMD and DES against the Long QT-3-causing mutation, ∆KPQ, to better detect any drug-mediated effect on INa,L. Our results show that AMD and DES do not affect WT or ∆KPQ activation; however, both drugs altered the apparent valence of steady-state fast-inactivation. In addition, AMD and DES preferentially block ∆KPQ peak conductance compared to WT. Both compounds significantly increase INa,L and window currents. We conclude that both compounds have pro-arrhythmic effects on NaV1.5, especially ∆KPQ; however, DES seems to have a greater pro-arrhythmic effect than AMD.

Site-selective probing of cTAR destabilization highlights the necessary plasticity of the HIV-1 nucleocapsid protein to chaperone the first strand transfer

Science.gov (United States)

Godet, Julien; Kenfack, Cyril; Przybilla, Frédéric; Richert, Ludovic; Duportail, Guy; Mély, Yves

2013-01-01

The HIV-1 nucleocapsid protein (NCp7) is a nucleic acid chaperone required during reverse transcription. During the first strand transfer, NCp7 is thought to destabilize cTAR, the (−)DNA copy of the TAR RNA hairpin, and subsequently direct the TAR/cTAR annealing through the zipping of their destabilized stem ends. To further characterize the destabilizing activity of NCp7, we locally probe the structure and dynamics of cTAR by steady-state and time resolved fluorescence spectroscopy. NC(11–55), a truncated NCp7 version corresponding to its zinc-finger domain, was found to bind all over the sequence and to preferentially destabilize the penultimate double-stranded segment in the lower part of the cTAR stem. This destabilization is achieved through zinc-finger–dependent binding of NC to the G10 and G50 residues. Sequence comparison further revealed that C•A mismatches close to the two G residues were critical for fine tuning the stability of the lower part of the cTAR stem and conferring to G10 and G50 the appropriate mobility and accessibility for specific recognition by NC. Our data also highlight the necessary plasticity of NCp7 to adapt to the sequence and structure variability of cTAR to chaperone its annealing with TAR through a specific pathway. PMID:23511968

Blocking Blood Flow to Solid Tumors by Destabilizing Tubulin: An Approach to Targeting Tumor Growth.

Science.gov (United States)

Pérez-Pérez, María-Jesús; Priego, Eva-María; Bueno, Oskía; Martins, Maria Solange; Canela, María-Dolores; Liekens, Sandra

2016-10-13

The unique characteristics of the tumor vasculature offer the possibility to selectively target tumor growth and vascularization using tubulin-destabilizing agents. Evidence accumulated with combretastatin A-4 (CA-4) and its prodrug CA-4P support the therapeutic value of compounds sharing this mechanism of action. However, the chemical instability and poor solubility of CA-4 demand alternative compounds that are able to surmount these limitations. This Perspective illustrates the different classes of compounds that behave similar to CA-4, analyzes their binding mode to αβ-tubulin according to recently available structural complexes, and includes described approaches to improve their delivery. In addition, dissecting the mechanism of action of CA-4 and analogues allows a closer insight into the advantages and drawbacks associated with these tubulin-destabilizing agents that behave as vascular disrupting agents (VDAs).

Protein tyrosine phosphatase receptor delta acts as a neuroblastoma tumor suppressor by destabilizing the aurora kinase a oncogene

LENUS (Irish Health Repository)

Meehan, Maria

2012-02-05

Abstract Background Protein tyrosine phosphatase receptor delta (PTPRD) is a member of a large family of protein tyrosine phosphatases which negatively regulate tyrosine phosphorylation. Neuroblastoma is a major childhood cancer arising from precursor cells of the sympathetic nervous system which is known to acquire deletions and alterations in the expression patterns of PTPRD, indicating a potential tumor suppressor function for this gene. The molecular mechanism, however, by which PTPRD renders a tumor suppressor effect in neuroblastoma is unknown. Results As a molecular mechanism, we demonstrate that PTPRD interacts with aurora kinase A (AURKA), an oncogenic protein that is over-expressed in multiple forms of cancer, including neuroblastoma. Ectopic up-regulation of PTPRD in neuroblastoma dephosphorylates tyrosine residues in AURKA resulting in a destabilization of this protein culminating in interfering with one of AURKA\\'s primary functions in neuroblastoma, the stabilization of MYCN protein, the gene of which is amplified in approximately 25 to 30% of high risk neuroblastoma. Conclusions PTPRD has a tumor suppressor function in neuroblastoma through AURKA dephosphorylation and destabilization and a downstream destabilization of MYCN protein, representing a novel mechanism for the function of PTPRD in neuroblastoma.

Thermodynamics of protein destabilization in live cells.

Science.gov (United States)

Danielsson, Jens; Mu, Xin; Lang, Lisa; Wang, Huabing; Binolfi, Andres; Theillet, François-Xavier; Bekei, Beata; Logan, Derek T; Selenko, Philipp; Wennerström, Håkan; Oliveberg, Mikael

2015-10-06

Although protein folding and stability have been well explored under simplified conditions in vitro, it is yet unclear how these basic self-organization events are modulated by the crowded interior of live cells. To find out, we use here in-cell NMR to follow at atomic resolution the thermal unfolding of a β-barrel protein inside mammalian and bacterial cells. Challenging the view from in vitro crowding effects, we find that the cells destabilize the protein at 37 °C but with a conspicuous twist: While the melting temperature goes down the cold unfolding moves into the physiological regime, coupled to an augmented heat-capacity change. The effect seems induced by transient, sequence-specific, interactions with the cellular components, acting preferentially on the unfolded ensemble. This points to a model where the in vivo influence on protein behavior is case specific, determined by the individual protein's interplay with the functionally optimized "interaction landscape" of the cellular interior.

Rational destabilizing speculation, positive feedback trading, and the oil bubble of 2008

International Nuclear Information System (INIS)

Tokic, Damir

2011-01-01

This article examines how the interaction of different participants in the crude oil futures markets affects the crude oil price efficiency. Normally, the commercial market participants, such as oil producers and oil consumers, act as arbitrageurs and ensure that the price of crude oil remains within the fundamental value range. However, institutional investors that invest in crude oil to diversify their portfolios and/or hedge inflation can destabilize the interaction among commercial participants and liquidity-providing speculators. We argue that institutional investors can impose limits to arbitrage, particularly during the financial crisis when the investment demand for commodities is particularly strong. In support, we show that commercials hedgers had significantly reduced their short positions leading to the 2008 oil bubble-they were potentially aggressively offsetting their short hedges. As a result, by essentially engaging in a positive feedback trading, commercial hedgers at least contributed to 'the 2008 oil bubble'. These findings have been mainly overlooked by the existing research. - Research Highlights: → This article finds that commercial hedgers at least contributed to the 2008 oil bubble. → Commercial hedgers were aggressively offsetting their short hedges leading to the oil bubble peak. → Commercial hedgers, thus, unwillingly engaged in positive feedback trading. → Institutional investors potentially destabilized the oil markets in 2008.

Modeling of sawtooth destabilization during radio-frequency heating experiments in tokamak plasmas

International Nuclear Information System (INIS)

McClements, K.G.; Dendy, R.O.; Hastie, R.J.; Martin, T.J.

1996-01-01

Sawtooth oscillations in tokamaks have been stabilized using ion cyclotron resonance heating (ICRH), but often reappear while ICRH continues. It is shown that the reappearance of sawteeth during one particular ICRH discharge in the Joint European Torus (JET) [Campbell et al., Phys. Rev. Lett. 60, 2148 (1988)] was correlated with a change of sign in the energy δW associated with m=1 internal kink displacements. To compute δW, a new analytical model is used for the distribution function of heated minority ions, which is consistent with Fokker endash Planck simulations of ICRH. Minority ions have a stabilizing influence, arising from third adiabatic invariant conservation, but also contribute to a destabilizing shift of magnetic flux surfaces. As the minor radius of the q=1 surface rises, the stabilizing influence of minority ions diminishes, and the shape of the plasma cross section becomes increasingly important. It is shown that an increase in ICRH power can destabilize the kink mode: this is consistent with observations of sawteeth in JET discharges with varying levels of ICRH. It is suggested that the sawtooth-free period could be prolonged by minimizing the vertical extent of the ICRH power deposition profile.1996 American Institute of Physics

Distance-dependent duplex DNA destabilization proximal to G-quadruplex/i-motif sequences

Science.gov (United States)

König, Sebastian L. B.; Huppert, Julian L.; Sigel, Roland K. O.; Evans, Amanda C.

2013-01-01

G-quadruplexes and i-motifs are complementary examples of non-canonical nucleic acid substructure conformations. G-quadruplex thermodynamic stability has been extensively studied for a variety of base sequences, but the degree of duplex destabilization that adjacent quadruplex structure formation can cause has yet to be fully addressed. Stable in vivo formation of these alternative nucleic acid structures is likely to be highly dependent on whether sufficient spacing exists between neighbouring duplex- and quadruplex-/i-motif-forming regions to accommodate quadruplexes or i-motifs without disrupting duplex stability. Prediction of putative G-quadruplex-forming regions is likely to be assisted by further understanding of what distance (number of base pairs) is required for duplexes to remain stable as quadruplexes or i-motifs form. Using oligonucleotide constructs derived from precedented G-quadruplexes and i-motif-forming bcl-2 P1 promoter region, initial biophysical stability studies indicate that the formation of G-quadruplex and i-motif conformations do destabilize proximal duplex regions. The undermining effect that quadruplex formation can have on duplex stability is mitigated with increased distance from the duplex region: a spacing of five base pairs or more is sufficient to maintain duplex stability proximal to predicted quadruplex/i-motif-forming regions. PMID:23771141

Rational destabilizing speculation, positive feedback trading, and the oil bubble of 2008

Energy Technology Data Exchange (ETDEWEB)

Tokic, Damir, E-mail: Damir.tokic@esc-rennes.f [ESC Rennes - International School of Business, 2 Rue Robert d' Arbissel, 35065 Rennes cedex (France)

2011-04-15

This article examines how the interaction of different participants in the crude oil futures markets affects the crude oil price efficiency. Normally, the commercial market participants, such as oil producers and oil consumers, act as arbitrageurs and ensure that the price of crude oil remains within the fundamental value range. However, institutional investors that invest in crude oil to diversify their portfolios and/or hedge inflation can destabilize the interaction among commercial participants and liquidity-providing speculators. We argue that institutional investors can impose limits to arbitrage, particularly during the financial crisis when the investment demand for commodities is particularly strong. In support, we show that commercials hedgers had significantly reduced their short positions leading to the 2008 oil bubble-they were potentially aggressively offsetting their short hedges. As a result, by essentially engaging in a positive feedback trading, commercial hedgers at least contributed to 'the 2008 oil bubble'. These findings have been mainly overlooked by the existing research. - Research Highlights: {yields} This article finds that commercial hedgers at least contributed to the 2008 oil bubble. {yields} Commercial hedgers were aggressively offsetting their short hedges leading to the oil bubble peak. {yields} Commercial hedgers, thus, unwillingly engaged in positive feedback trading. {yields} Institutional investors potentially destabilized the oil markets in 2008.

Results of preventive radioiodine therapy in euthyroid patients with history of hyperthyroidism prior to administration of amiodarone with permanent atrial fibrillation--a preliminary study.

Science.gov (United States)

Czarnywojtek, Agata; Zgorzalewicz-Stachowiak, Małgorzata; Woliński, Kosma; Płazińska, Maria Teresa; Miechowicz, Izabela; Kwiecińska, Barbara; Czepczyński, Rafał; Królicki, Leszek; Ruchała, Marek

2014-01-01

Radioiodine (RAI) therapy is a standard procedure in the treatment of hyperthyroidism. However, the use of RAI in euthyroid patients requiring chronic administration of amiodarone (AM) where other antiarrhythmic drugs may lack efficacy is still controversial. The aim of this study was to assess the safety and efficacy of an AM therapy prior to treatment with radioiodine therapy in euthyroid patients with permanent atrial fibrillation (PAF), who had been treated for hyperthyroidism in the past. This was a retrospective observational study. Patients were assessed at baseline and two, six, eight, and 12 months after RAI therapy. 17 euthyroid patients with PAF were qualified to the RAI (female/male 3/14; age range 65 to 87, median 71). The patients required chronic administration of AM as a prophylaxis against sudden death. Each patient received an ablative dose of 800 MBq (22 mCi) of 131I. At baseline and during follow-up, no side effects of the therapy and no signs of drug intolerance were observed. Subclinical hyperthyroidism occurred in two (11.8%) cases after two months of RAI and five weeks of AM administration. In this situation, RAI therapy was repeated. Three patients (17.6%) after six months, and another two (11.8%) after eight months, required an additional dose of 131I due to amiodarone-induced thyrotoxicosis (AIT). Twelve patients (70.6%) returned to spontaneous sinus rhythm within two months. Fourteen patients (82.4%) had sinus rhythm during follow-up after six and 12 months of treatment. Preventive RAI in euthyroid (but previously hyperthyroid) patients with PAF before administration of AM may be the method of choice. This is particularly important for patients who will require permanent AM administration as a life-saving drug.

Insecticide resistance and intracellular proteases.

Science.gov (United States)

Wilkins, Richard M

2017-12-01

Pesticide resistance is an example of evolution in action with mechanisms of resistance arising from mutations or increased expression of intrinsic genes. Intracellular proteases have a key role in maintaining healthy cells and in responding to stressors such as pesticides. Insecticide-resistant insects have constitutively elevated intracellular protease activity compared to corresponding susceptible strains. This increase was shown for some cases originally through biochemical enzyme studies and subsequently putatively by transcriptomics and proteomics methods. Upregulation and expression of proteases have been characterised in resistant strains of some insect species, including mosquitoes. This increase in proteolysis results in more degradation products (amino acids) of intracellular proteins. These may be utilised in the resistant strain to better protect the cell from stress. There are changes in insect intracellular proteases shortly after insecticide exposure, suggesting a role in stress response. The use of protease and proteasome inhibitors or peptide mimetics as synergists with improved application techniques and through protease gene knockdown using RNA interference (possibly expressed in crop plants) may be potential pest management strategies, in situations where elevated intracellular proteases are relevant. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Poliovirus-associated protein kinase: Destabilization of the virus capsid and stimulation of the phosphorylation reaction by Zn2+

International Nuclear Information System (INIS)

Ratka, M.; Lackmann, M.; Ueckermann, C.; Karlins, U.; Koch, G.

1989-01-01

The previously described poliovirus-associated protein kinase activity phosphorylates viral proteins VP0 and VP2 as well as exogenous proteins in the presence of Mg 2+ . In this paper, the effect of Zn 2+ on the phosphorylation reaction and the stability of the poliovirus capsid has been studied in detail and compared to that of Mg 2+ . In the presence of Zn 2+ , phosphorylation of capsid proteins VP2 and VP4 is significantly higher while phosphorylation of VP0 and exogenous phosphate acceptor proteins is not detected. The results indicate the activation of more than one virus-associated protein kinase by Zn 2+ . The ion-dependent behavior of the enzyme activities is observed independently of whether the virus was obtained from HeLa or green monkey kidney cells. The poliovirus capsid is destabilized by Zn 2+ . This alteration of the poliovirus capsid structure is a prerequisite for effective phosphorylation of viral capsid proteins. The increased level of phosphorylation of viral capsid proteins results in further destabilization of the viral capsid. As a result of the conformational changes, poliovirus-associated protein kinase activities dissociate from the virus particle. The authors suggest that the destabilizing effect of phosphorylation on the viral capsid plays a role in uncoating of poliovirus

Improved exogenous DNA uptake in bovine spermatozoa and gene expression in embryos using membrane destabilizing agents in ICSI-SMGT.

Science.gov (United States)

Sánchez-Villalba, Esther; Arias, María Elena; Zambrano, Fabiola; Loren, Pía; Felmer, Ricardo

2018-02-01

Sperm-mediated gene transfer (SMGT) is a simple, fast, and economical biotechnological tool for producing transgenic animals. However, transgene expression with this technique in bovine embryos is still inefficient due to low uptake and binding of exogenous DNA in spermatozoa. The present study evaluated the effects of sperm membrane destabilization on the binding capacity, location and quantity of bound exogenous DNA in cryopreserved bovine spermatozoa using Triton X-100 (TX-100), lysolecithin (LL) and sodium hydroxide (NaOH). Effects of these treatments were also evaluated by intracytoplasmic sperm injection (ICSI)-SMGT. Results showed that all treatments bound exogenous DNA to spermatozoa including the control. Spermatozoa treated with different membrane destabilizing agents bound the exogenous DNA throughout the head and tail of spermatozoa, compared with the control, in which binding occurred mainly in the post-acrosomal region and tail. The amount of exogenous DNA bound to spermatozoa was much higher for the different sperm treatments than the control (P Exogenous gene expression in embryos was also improved by these treatments. These results demonstrated that sperm membrane destabilization could be a novel strategy in bovine SMGT protocols for the generation of transgenic embryos by ICSI.

Gravity-induced encapsulation of liquids by destabilization of granular rafts

Science.gov (United States)

Abkarian, Manouk; Protière, Suzie; Aristoff, Jeffrey M.; Stone, Howard A.

2013-05-01

Droplets and bubbles coated by a protective armour of particles find numerous applications in encapsulation, stabilization of emulsions and foams, and flotation techniques. Here we study the role of a body force, such as in flotation, as a means of continuous encapsulation by particles. We use dense particles, which self-assemble into rafts, at oil-water interfaces. We show that these rafts can be spontaneously or controllably destabilized into armoured oil-in-water droplets, which highlights a possible role for common granular materials in environmental remediation. We further present a method for continuous production and discuss the generalization of our approach towards colloidal scales.

Structural determinants of HIV-1 nucleocapsid protein for cTAR DNA binding and destabilization, and correlation with inhibition of self-primed DNA synthesis.

Science.gov (United States)

Beltz, Hervé; Clauss, Céline; Piémont, Etienne; Ficheux, Damien; Gorelick, Robert J; Roques, Bernard; Gabus, Caroline; Darlix, Jean-Luc; de Rocquigny, Hugues; Mély, Yves

2005-05-20

The nucleocapsid protein (NC) of human immunodeficiency virus type 1 (HIV-1) is formed of two highly conserved CCHC zinc fingers flanked by small basic domains. NC is required for the two obligatory strand transfers in viral DNA synthesis through its nucleic acid chaperoning properties. The first DNA strand transfer relies on NC's ability to bind and destabilize the secondary structure of complementary transactivation response region (cTAR) DNA, to inhibit self-priming, and to promote the annealing of cTAR to TAR RNA. To further investigate NC chaperone properties, our aim was to identify by fluorescence spectroscopy and gel electrophoresis, the NC structural determinants for cTAR binding and destabilization, and for the inhibition of self-primed DNA synthesis on a model system using a series of NC mutants and HIV-1 reverse transcriptase. NC destabilization and self-priming inhibition properties were found to be supported by the two fingers in their proper context and the basic (29)RAPRKKG(35) linker. The strict requirement of the native proximal finger suggests that its hydrophobic platform (Val13, Phe16, Thr24 and Ala25) is crucial for binding, destabilization and inhibition of self-priming. In contrast, only partial folding of the distal finger is required, probably for presenting the Trp37 residue in an appropriate orientation. Also, Trp37 and the hydrophobic residues of the proximal finger appear to be essential for the propagation of the melting from the cTAR ends up to the middle of the stem. Finally, both N-terminal and C-terminal basic domains contribute to cTAR binding but not to its destabilization.

Endoplasmic reticulum stress as a novel mechanism in amiodarone-induced destructive thyroiditis.

Science.gov (United States)

Lombardi, Angela; Inabnet, William Barlow; Owen, Randall; Farenholtz, Kaitlyn Ellen; Tomer, Yaron

2015-01-01

Amiodarone (AMIO) is one of the most effective antiarrhythmic drugs available; however, its use is limited by a serious side effect profile, including thyroiditis. The mechanisms underlying AMIO thyroid toxicity have been elusive; thus, identification of novel approaches in order to prevent thyroiditis is essential in patients treated with AMIO. Our aim was to evaluate whether AMIO treatment could induce endoplasmic reticulum (ER) stress in human thyroid cells and the possible implications of this effect in AMIO-induced destructive thyroiditis. Here we report that AMIO, but not iodine, significantly induced the expression of ER stress markers including Ig heavy chain-binding protein (BiP), phosphoeukaryotic translation initiation factor 2α (eIF2α), CCAAT/enhancer-binding protein homologous protein (CHOP) and spliced X-box binding protein-1 (XBP-1) in human thyroid ML-1 cells and human primary thyrocytes. In both experimental systems AMIO down-regulated thyroglobulin (Tg) protein but had little effect on Tg mRNA levels, suggesting a mechanism involving Tg protein degradation. Indeed, pretreatment with the specific proteasome inhibitor MG132 reversed AMIO-induced down-regulation of Tg protein levels, confirming a proteasome-dependent degradation of Tg protein. Corroborating our findings, pretreatment of ML-1 cells and human primary thyrocytes with the chemical chaperone 4-phenylbutyric acid completely prevented the effect of AMIO on both ER stress induction and Tg down-regulation. We identified ER stress as a novel mechanism contributing to AMIO-induced destructive thyroiditis. Our data establish that AMIO-induced ER stress impairs Tg expression via proteasome activation, providing a valuable therapeutic avenue for the treatment of AMIO-induced destructive thyroiditis.

Imaging and controlling intracellular reactions: Lysosome transport as a function of diameter and the intracellular synthesis of conducting polymers

Science.gov (United States)

Payne, Christine

2014-03-01

Eukaryotic cells are the ultimate complex environment with intracellular chemical reactions regulated by the local cellular environment. For example, reactants are sequestered into specific organelles to control local concentration and pH, motor proteins transport reactants within the cell, and intracellular vesicles undergo fusion to bring reactants together. Current research in the Payne Lab in the School of Chemistry and Biochemistry at Georgia Tech is aimed at understanding and utilizing this complex environment to control intracellular chemical reactions. This will be illustrated using two examples, intracellular transport as a function of organelle diameter and the intracellular synthesis of conducting polymers. Using single particle tracking fluorescence microscopy, we measured the intracellular transport of lysosomes, membrane-bound organelles, as a function of diameter as they underwent transport in living cells. Both ATP-dependent active transport and diffusion were examined. As expected, diffusion scales with the diameter of the lysosome. However, active transport is unaffected suggesting that motor proteins are insensitive to cytosolic drag. In a second example, we utilize intracellular complexity, specifically the distinct micro-environments of different organelles, to carry out chemical reactions. We show that catalase, found in the peroxisomes of cells, can be used to catalyze the polymerization of the conducting polymer PEDOT:PSS. More importantly, we have found that a range of iron-containing biomolecules are suitable catalysts with different iron-containing biomolecules leading to different polymer properties. These experiments illustrate the advantage of intracellular complexity for the synthesis of novel materials.

Critical thinking of destabilizing interpretations of events and phenomena: the role of economic sciences

Directory of Open Access Journals (Sweden)

Тетяна Андріївна Непокупна

2015-03-01

Full Text Available This article analyzes the global transformations and their impact on the main society life; the specifics of modern interpretations of events and phenomena, their destabilizing effects on behavior, health and life of humans; the role of economic sciences in the formation of critical thinking as a means of combating ignorance and propaganda, formation of an objective world view that grounded on knowledge

Nanoparticles for intracellular-targeted drug delivery

International Nuclear Information System (INIS)

Paulo, Cristiana S O; Pires das Neves, Ricardo; Ferreira, Lino S

2011-01-01

Nanoparticles (NPs) are very promising for the intracellular delivery of anticancer and immunomodulatory drugs, stem cell differentiation biomolecules and cell activity modulators. Although initial studies in the area of intracellular drug delivery have been performed in the delivery of DNA, there is an increasing interest in the use of other molecules to modulate cell activity. Herein, we review the latest advances in the intracellular-targeted delivery of short interference RNA, proteins and small molecules using NPs. In most cases, the drugs act at different cellular organelles and therefore the drug-containing NPs should be directed to precise locations within the cell. This will lead to the desired magnitude and duration of the drug effects. The spatial control in the intracellular delivery might open new avenues to modulate cell activity while avoiding side-effects.

Efficient intracellular delivery and improved biocompatibility of colloidal silver nanoparticles towards intracellular SERS immuno-sensing.

Science.gov (United States)

Bhardwaj, Vinay; Srinivasan, Supriya; McGoron, Anthony J

2015-06-21

High throughput intracellular delivery strategies, electroporation, passive and TATHA2 facilitated diffusion of colloidal silver nanoparticles (AgNPs) are investigated for cellular toxicity and uptake using state-of-art analytical techniques. The TATHA2 facilitated approach efficiently delivered high payload with no toxicity, pre-requisites for intracellular applications of plasmonic metal nanoparticles (PMNPs) in sensing and therapeutics.

Destabilization of counter-propagating TAEs by off-axis, co-current Neutral Beam Injection

Science.gov (United States)

Podesta', M.; Fredrickson, E.; Gorelenkova, M.

2017-10-01

Neutral Beam injection (NBI) is a common tool to heat the plasma and drive current non-inductively in fusion devices. Energetic particles (EP) resulting from NBI can drive instabilities that are detrimental for the performance and the predictability of plasma discharges. A broad NBI deposition profile, e.g. by off-axis injection aiming near the plasma mid-radius, is often assumed to limit those undesired effects by reducing the radial gradient of the EP density, thus reducing the ``universal'' drive for instabilities. However, this work presents new evidence that off-axis NBI can also lead to undesired effects such as the destabilization of Alfvénic instabilities, as observed in NSTX-U plasmas. Experimental observations indicate that counter propagating toroidal AEs are destabilized as the radial EP density profile becomes hollow as a result of off-axis NBI. Time-dependent analysis with the TRANSP code, augmented by a reduced fast ion transport model (known as kick model), indicates that instabilities are driven by a combination of radial and energy gradients in the EP distribution. Understanding the mechanisms for wave-particle interaction, revealed by the phase space resolved analysis, is the basis to identify strategies to mitigate or suppress the observed instabilities. Work supported by the U.S. Department of Energy, Office of Science, Office of Fusion Energy Sciences under Contract Number DE-AC02-09CH11466.

MR imaging of intracellular and extracellular deoxyhemoglobin

International Nuclear Information System (INIS)

Janick, P.A.; Grossman, R.I.; Asakura, T.

1989-01-01

MR imaging was performed on varying concentrations of intracellular and extracellular deoxyhemoglobin as well as varying proportions of deoxyhemoglobin and oxyhemoglobin in vitro at 1.5T with use of standard spin-echo and gradient-refocused spin sequences. This study indicates that susceptibility-induced T2 shortening occurs over a broad range of intracellular deoxyhemoglobin concentrations (maximal at hematocrits between 20% and 45%), reflecting diffusional effects at the cellular level. T2* gradient-echo imaging enhances the observed hypointensity in images of intracellular deoxyhemoglobin. The characteristic MR appearance of acute hemotomas can be modeled by the behavior of intracellular and extracellular deoxyhemoglobin and oxyhemoglobin

Influence of molybdenum on hardenability of destabilized high chromium cast irons. Ko kuromu chutetsu no netsushori tokusei ni oyobosu moribuden no eikyo

Energy Technology Data Exchange (ETDEWEB)

Kuwano, M. (Ube, Collete of Technology, Yamaguchi (Japan)); Ogi, K. (Kyushu University, Fukuoka (Japan). Faculty of Engineering); Sawamoto, A. (Yamaguchi University, Yamaguchi (Japan))

1991-07-25

Alloy elements are added to compensate for reduction in hardenability of high chromium cast iron as a result of destabilization heat treatment. With the purpose of investigating the influence of the added elements, this paper describes investigations made on effects of Mo and destabilization heat treatment conditions on martensite transformation , eutectoid transformation and base hardness, using Fe-Cr-C-based alloy containing C at 1.6-3.6%, and Cr at 6.6-26%. The main results obtained are as follows: Mo and Cr had the distribution coefficient for initial crystal austenite is smaller than one and were microscopically segregated in the dendrite base; the solute element distribution in the base is homogenized as a result of the long-time destabilization heat treatment while the Cr and C concentrations reduce largely, and the Mo concentration increases slightly; Mo has little influence on the Ms point, and reduces with the smaller the Cr/C ratio and the higher the retention temperature; and the base hardness corresponds well to a CCT diagram. 14 refs., 11 figs., 1 tab.

Destabilization of Titania Nanosheet Suspensions by Inorganic Salts: Hofmeister Series and Schulze-Hardy Rule.

Science.gov (United States)

Rouster, Paul; Pavlovic, Marko; Szilagyi, Istvan

2017-07-13

Ion specific effects on colloidal stability of titania nanosheets (TNS) were investigated in aqueous suspensions. The charge of the particles was varied by the pH of the solutions, therefore, the influence of mono- and multivalent anions on the charging and aggregation behavior could be studied when they were present either as counter or co-ions in the systems. The aggregation processes in the presence of inorganic salts were mainly driven by interparticle forces of electrostatic origin, however, chemical interactions between more complex ions and the surface led to additional attractive forces. The adsorption of anions significantly changed the surface charge properties and hence, the resistance of the TNS against salt-induced aggregation. On the basis of their ability in destabilization of the dispersions, the monovalent ions could be ordered according to the Hofmeister series in acidic solutions, where they act as counterions. However, the behavior of the biphosphate anion was atypical and its adsorption induced charge reversal of the particles. The multivalent anions destabilized the oppositely charged TNS more effectively and the aggregation processes followed the Schulze-Hardy rule. Only weak or negligible interactions were observed between the anions and the particles in alkaline suspensions, where the TNS possessed negative charge.

The Destabilization of Protected Soil Organic Carbon Following Experimental Drought at the Pore and Core scale

Science.gov (United States)

Smith, A. P.; Bond-Lamberty, B. P.; Tfaily, M. M.; Todd-Brown, K. E.; Bailey, V. L.

2015-12-01

The movement of water and solutes through the pore matrix controls the distribution and transformation of carbon (C) in soils. Thus, a change in the hydrologic connectivity, such as increased saturation, disturbance or drought, may alter C mineralization and greenhouse gas (GHG) fluxes to the atmosphere. While these processes occur at the pore scale, they are often investigated at coarser scale. This project investigates pore- and core-scale soil C dynamics with varying hydrologic factors (simulated precipitation, groundwater-led saturation, and drought) to assess how climate-change induced shifts in hydrologic connectivity influences the destabilization of protected C in soils. Surface soil cores (0-15 cm depth) were collected from the Disney Wilderness Preserve, Florida, USA where water dynamics, particularly water table rise and fall, appear to exert a strong control on the emissions of GHGs and the persistence of soil organic matter in these soils. We measured CO2 and CH4 from soils allowed to freely imbibe water from below to a steady state starting from either field moist conditions or following experimental drought. Parallel treatments included the addition of similar quantities of water from above to simulate precipitation. Overall respiration increased in soil cores subjected to drought compared to field moist cores independent of wetting type. Cumulative CH4 production was higher in drought-induced soils, especially in the soils subjected to experimental groundwater-led saturation. Overall, the more C (from CO2 and CH4) was lost in drought-induced soils compared to field moist cores. Our results indicate that future drought events could have profound effects on the destabilization of protected C, especially in groundwater-fed soils. Our next steps focus on how to accurately capture drought-induced C destabilization mechanisms in earth system models.

Sediment-stabilizing and Destabilizing Ecoengineering Species from River to Estuary: the Case of the Scheldt System

Science.gov (United States)

Selakovic, S.; Cozzoli, F.; Leuven, J.; Van Braeckel, A.; Speybroeck, J.; Kleinhans, M. G.; Bouma, T.

2017-12-01

Interactions between organisms and landscape forming processes play an important role in evolution of coastal landscapes. In particular, biota has a strong potential to interact with important geomorphological processes such as sediment dynamics. Although many studies worked towards quantifying the impact of different species groups on sediment dynamics, information has been gathered on an ad hoc base. Depending on species' traits and distribution, functional groups of ecoengineering species may have differential effects on sediment deposition and erosion. We hypothesize that the spatial distributions of sediment-stabilizing and destabilizing species across the channel and along the whole salinity gradient of an estuary partly determine the planform shape and channel-shoal morphology of estuaries. To test this hypothesis, we analyze vegetation and macrobenthic data taking the Scheldt river-estuarine continuum as model ecosystem. We identify species traits with important effects on sediment dynamics and use them to form functional groups. By using linearized mixed modelling, we are able to accurately describe the distributions of the different functional groups. We observe a clear distinction of dominant ecosystem engineering functional groups and their potential effects on the sediment in the river-estuarine continuum. The first results of longitudinal cross section show the highest effects of stabilizing plant species in riverine and sediment bioturbators in weak polyhaline part of continuum. The distribution of functional groups in transverse cross sections shows dominant stabilizing effect in supratidal zone compared to dominant destabilizing effect in the lower intertidal zone. This analysis offers a new and more general conceptualization of distributions of sediment stabilizing and destabilizing functional groups and their potential impacts on sediment dynamics, shoal patterns, and planform shapes in river-estuarine continuum. We intend to test this in future

Experimental study of smectite interaction with metal iron at low temperature: 1. Smectite destabilization.

OpenAIRE

Lantenois , Sébastien; Lanson , Bruno; Muller , Fabrice; Bauer , Andreas; Jullien , Michel; Plançon , Alain

2005-01-01

Interaction between metal Fe and a variety of natural and synthetic smectite samples with contrasting crystal chemistry was studied by scanning electron microscopy and X-ray diffraction from experiments conducted at 80°C. These experiments demonstrate an important reactivity contrast as a function of smectite crystal chemistry. An XRD method involving the use of an internal standard allowed quantification of the relative proportion of smectite destabilized as a function of initial pH conditio...

Single-cell intracellular nano-pH probes†

OpenAIRE

Özel, Rıfat Emrah; Lohith, Akshar; Mak, Wai Han; Pourmand, Nader

2015-01-01

Within a large clonal population, such as cancerous tumor entities, cells are not identical, and the differences between intracellular pH levels of individual cells may be important indicators of heterogeneity that could be relevant in clinical practice, especially in personalized medicine. Therefore, the detection of the intracellular pH at the single-cell level is of great importance to identify and study outlier cells. However, quantitative and real-time measurements of the intracellular p...

NMR studies on the mechanism of structural destabilization of the globular proteins and DNA by aliphatic alcohols

International Nuclear Information System (INIS)

Lubas, B.; Witman, B.; Wieniewska, T.; Soltysik, M.

1977-01-01

The concept that the mechanism of structural destabilization of the biologically active macromolecules by typical denaturing agents should find a reflection in the NMR spectra of the denaturants themselves has been followed by proton NMR for some aliphatic alcohols in the system containing the serum albumin of DNA. (author)

The effects of cognitive reappraisal following retrieval-procedures designed to destabilize alcohol memories in high-risk drinkers.

Science.gov (United States)

Hon, Tiffany; Das, Ravi K; Kamboj, Sunjeev K

2016-03-01

Addiction is a disorder of motivational learning and memory. Maladaptive motivational memories linking drug-associated stimuli to drug seeking are formed over hundreds of reinforcement trials and accompanied by aberrant neuroadaptation in the mesocorticolimbic reward system. Such memories are resistant to extinction. However, the discovery of retrieval-dependent memory plasticity has opened up the possibility of permanent modification of established (long-term) memories during 'reconsolidation'. Here, we investigate whether reappraisal of maladaptive alcohol cognitions performed after procedures designed to destabilize alcohol memory networks affected subsequent alcohol memory, craving, drinking and attentional bias. Forty-seven at-risk drinkers attended two sessions. On the first lab session, participants underwent one of two prediction error-generating procedures in which outcome expectancies were violated while retrieving alcohol memories (omission and value prediction error groups). Participants in a control group retrieved non-alcohol memories. Participants then reappraised personally relevant maladaptive alcohol memories and completed measures of reappraisal recall, alcohol verbal fluency and craving. Seven days later, they repeated these measures along with attentional bias assessment. Omission prediction error (being unexpectedly prevented from drinking beer), but not a value prediction error (drinking unexpectedly bitter-tasting beer) or control procedure (drinking unexpectedly bitter orange juice), was associated with significant reductions in verbal fluency for positive alcohol-related words. No other statistically robust outcomes were detected. This study provides partial preliminary support for the idea that a common psychotherapeutic strategy used in the context of putative memory retrieval-destabilization can alter accessibility of alcohol semantic networks. Further research delineating the necessary and sufficient requirements for producing

CRITICAL HEIGHT FOR THE DESTABILIZATION OF SOLAR PROMINENCES: STATISTICAL RESULTS FROM STEREO OBSERVATIONS

Energy Technology Data Exchange (ETDEWEB)

Liu Kai; Wang Yuming; Wang Shui; Shen Chenglong, E-mail: ymwang@ustc.edu.cn [CAS Key Laboratory of Geospace Environment, Department of Geophysics and Planetary Sciences, University of Science and Technology of China, Hefei, Anhui 230026 (China)

2012-01-10

At which height is a prominence inclined to be unstable, or where is the most probable critical height for the prominence destabilization? This question was statistically studied based on 362 solar limb prominences well recognized by Solar Limb Prominence Catcher and Tracker from 2007 April to the end of 2009. We found that there are about 71% disrupted prominences (DPs), among which about 42% of them did not erupt successfully and about 89% of them experienced a sudden destabilization process. After a comprehensive analysis of the DPs, we discovered the following: (1) Most DPs become unstable at a height of 0.06-0.14 R{sub Sun} from the solar surface, and there are two most probable critical heights at which a prominence is very likely to become unstable, the first one is 0.13 R{sub Sun} and the second one is 0.19 R{sub Sun }. (2) An upper limit for the erupting velocity of eruptive prominences (EPs) exists, which decreases following a power law with increasing height and mass; accordingly, the kinetic energy of EPs has an upper limit too, which decreases as the critical height increases. (3) Stable prominences are generally longer and heavier than DPs, and not higher than 0.4 R{sub Sun }. (4) About 62% of the EPs were associated with coronal mass ejections (CMEs); but there is no difference in apparent properties between EPs associated with CMEs and those that are not.

CRITICAL HEIGHT FOR THE DESTABILIZATION OF SOLAR PROMINENCES: STATISTICAL RESULTS FROM STEREO OBSERVATIONS

International Nuclear Information System (INIS)

Liu Kai; Wang Yuming; Wang Shui; Shen Chenglong

2012-01-01

At which height is a prominence inclined to be unstable, or where is the most probable critical height for the prominence destabilization? This question was statistically studied based on 362 solar limb prominences well recognized by Solar Limb Prominence Catcher and Tracker from 2007 April to the end of 2009. We found that there are about 71% disrupted prominences (DPs), among which about 42% of them did not erupt successfully and about 89% of them experienced a sudden destabilization process. After a comprehensive analysis of the DPs, we discovered the following: (1) Most DPs become unstable at a height of 0.06-0.14 R ☉ from the solar surface, and there are two most probable critical heights at which a prominence is very likely to become unstable, the first one is 0.13 R ☉ and the second one is 0.19 R ☉ . (2) An upper limit for the erupting velocity of eruptive prominences (EPs) exists, which decreases following a power law with increasing height and mass; accordingly, the kinetic energy of EPs has an upper limit too, which decreases as the critical height increases. (3) Stable prominences are generally longer and heavier than DPs, and not higher than 0.4 R ☉ . (4) About 62% of the EPs were associated with coronal mass ejections (CMEs); but there is no difference in apparent properties between EPs associated with CMEs and those that are not.

Amiodarone increases the accumulation of DEA in a human alveolar epithelium-derived cell line.

Science.gov (United States)

Seki, Satoru; Itagaki, Shirou; Kobayashi, Masaki; Hirano, Takeshi; Iseki, Ken

2008-07-01

Amiodarone (AMD)-induced pulmonary toxicity (AIPT) is the most life-threatening side-effect of AMD treatment. N-Monodesethylamiodarone (DEA), an active metabolite of AMD, also exhibits cytotoxicity and tends to accumulate in the lung more intensively than AMD. In this study, we characterized the mechanism of DEA accumulation using A549 cells as a model of the alveolar epithelium. Typical ATP-depletion compounds caused an approximately 30% increase in the accumulation of DEA in A549 cells, although these effects were less than those in Caco-2 cells. Triiodothyronine (T(3)), which exhibited an inhibitory effect on DEA efflux in Caco-2 cells, did not affect the accumulation of DEA in A549 cells. On the other hand, 100 microM AMD caused an approximately 200% increase in DEA content in A549 cells, although AMD accumulation was not affected by 100 microM DEA. Since the reducing effect of AMD on cellular ATP levels and that of FCCP were similar, the mechanism by which DEA accumulation is increased by AMD might be different from the ATP-dependent DEA efflux mechanism. The decrease in cell viability by DEA in the presence of AMD (IC(50) value of DEA for A549 cell viability: 25.4+/-2.4 microM) was more pronounced than that by DEA alone (IC(50) value: 11.5+/-3.0 microM). This further DEA accumulation by AMD might be a factor responsible for the greater accumulation of DEA than that of AMD in the lung in long-term AMD-treated patients.

Morphometric Characterization of Rat and Human Alveolar Macrophage Cell Models and their Response to Amiodarone using High Content Image Analysis.

Science.gov (United States)

Hoffman, Ewelina; Patel, Aateka; Ball, Doug; Klapwijk, Jan; Millar, Val; Kumar, Abhinav; Martin, Abigail; Mahendran, Rhamiya; Dailey, Lea Ann; Forbes, Ben; Hutter, Victoria

2017-12-01

Progress to the clinic may be delayed or prevented when vacuolated or "foamy" alveolar macrophages are observed during non-clinical inhalation toxicology assessment. The first step in developing methods to study this response in vitro is to characterize macrophage cell lines and their response to drug exposures. Human (U937) and rat (NR8383) cell lines and primary rat alveolar macrophages obtained by bronchoalveolar lavage were characterized using high content fluorescence imaging analysis quantification of cell viability, morphometry, and phospholipid and neutral lipid accumulation. Cell health, morphology and lipid content were comparable (p content. Responses to amiodarone, a known inducer of phospholipidosis, required analysis of shifts in cell population profiles (the proportion of cells with elevated vacuolation or lipid content) rather than average population data which was insensitive to the changes observed. A high content image analysis assay was developed and used to provide detailed morphological characterization of rat and human alveolar-like macrophages and their response to a phospholipidosis-inducing agent. This provides a basis for development of assays to predict or understand macrophage vacuolation following inhaled drug exposure.

Intracellular Drug Bioavailability: Effect of Neutral Lipids and Phospholipids.

Science.gov (United States)

Treyer, Andrea; Mateus, André; Wiśniewski, Jacek R; Boriss, Hinnerk; Matsson, Pär; Artursson, Per

2018-06-04

Intracellular unbound drug concentrations are the pharmacologically relevant concentrations for targets inside cells. Intracellular drug concentrations are determined by multiple processes, including the extent of drug binding to intracellular structures. The aim of this study was to evaluate the effect of neutral lipid (NL) and phospholipid (PL) levels on intracellular drug disposition. The NL and/or PL content of 3T3-L1 cells were enhanced, resulting in phenotypes (in terms of morphology and proteome) reminiscent of adipocytes (high NL and PL) or mild phospholipidosis (only high PL). Intracellular bioavailability ( F ic ) was then determined for 23 drugs in these cellular models and in untreated wild-type cells. A higher PL content led to higher intracellular drug binding and a lower F ic . The induction of NL did not further increase drug binding but led to altered F ic due to increased lysosomal pH. Further, there was a good correlation between binding to beads coated with pure PL and intracellular drug binding. In conclusion, our results suggest that PL content is a major determinant of drug binding in cells and that PL beads may constitute a simple alternative to estimating this parameter. Further, the presence of massive amounts of intracellular NLs did not influence drug binding significantly.

Electrical storm: case series and review of management.

Science.gov (United States)

Srivatsa, Uma N; Ebrahimi, Ramin; El-Bialy, Adel; Wachsner, Robin Y

2003-09-01

Electrical storm is defined as a recurrent episode of hemodynamically destabilizing ventricular tachyarrhythmia that usually requires electrical cardioversion or defibrillation. We describe three cases presenting with electrical storm under differing circumstances: (1) a 57-year-old man with ST-elevation myocardial infarction within 1 week of a posterior circulation stroke who developed refractory sustained ventricular tachycardia 10 days after an acute myocardial infarction; (2) a 65-year-old man who developed polymorphic ventricular tachycardia and ventricular fibrillation following dobutamine echocardiography; and (3) a 20-year-old woman who developed intractable ventricular fibrillation following an overdose of a weight-reduction pill. The management of electrical storm is discussed, and evolving literature supporting the routine use of intravenous amiodarone and beta-blockers in place of intravenous lidocaine is critically examined.

Enhanced intracellular delivery and antibacterial efficacy of enrofloxacin-loaded docosanoic acid solid lipid nanoparticles against intracellular Salmonella.

Science.gov (United States)

Xie, Shuyu; Yang, Fei; Tao, Yanfei; Chen, Dongmei; Qu, Wei; Huang, Lingli; Liu, Zhenli; Pan, Yuanhu; Yuan, Zonghui

2017-01-23

Enrofloxacin-loaded docosanoic acid solid lipid nanoparticles (SLNs) with different physicochemical properties were developed to enhance activity against intracellular Salmonella. Their cellular uptake, intracellular elimination and antibacterial activity were studied in RAW 264.7 cells. During the experimental period, SLN-encapsulated enrofloxacin accumulated in the cells approximately 27.06-37.71 times more efficiently than free drugs at the same extracellular concentration. After incubation for 0.5 h, the intracellular enrofloxacin was enhanced from 0.336 to 1.147 μg/mg of protein as the sizes of nanoparticles were increased from 150 to 605 nm, and from 0.960 to 1.147 μg/mg of protein when the charge was improved from -8.1 to -24.9 mv. The cellular uptake was more significantly influenced by the size than it was by the charge, and was not affected by whether the charge was positive or negative. The elimination of optimal SLN-encapsulated enrofloxacin from the cells was significantly slower than that of free enrofloxacin after removing extracellular drug. The inhibition effect against intracellular Salmonella CVCC541 of 0.24 and 0.06 μg/mL encapsulated enrofloxacin was stronger than 0.6 μg/mL free drug after all of the incubation periods and at 48 h, respectively. Docosanoic acid SLNs are thus considered as a promising carrier for intracellular bacterial treatment.

Asymmetric membranes for destabilization of oil droplets in produced water from alkaline-surfactant-polymer (ASP) flooding

Science.gov (United States)

Ramlee, Azierah; Chiam, Chel-Ken; Sarbatly, Rosalam

2018-05-01

This work presents a study of destabilization of oil droplets in the produced water from alkaline-surfactant-polymer (ASP) flooding by using four types of laboratory-fabricated polyvinylidene fluoride (PVDF) membranes. The PVDF membranes were fabricated via immersion precipitation method with ethanol (0 - 30 %, v/v) as the coagulant. The membranes with the effective area of 17.35 cm2 were tested with synthesized ASP solution as the feed in cross-flow microfiltration process. The ASP feed solution initially contained the oil droplets with radius ranged from 40 to 100 nm and the mean radius was 61 nm. Results have shown that the concentration of the ethanol in the coagulation bath affects the formation of the membrane structure and the corresponding porosity, while no significance influence on the membrane thickness. Coalescence of the oil droplets was occurred when the ASP solution permeated through the asymmetric PVDF membranes. Through the coalescence process, the oil droplets were destabilized where the radius of the oil droplets in the permeates increased to 1.5-4 µm with the corresponding mean radius ranged from 2.4 to 2.7 µm.

Self-organization of intracellular gradients during mitosis

Directory of Open Access Journals (Sweden)

Fuller Brian G

2010-01-01

Full Text Available Abstract Gradients are used in a number of biological systems to transmit spatial information over a range of distances. The best studied are morphogen gradients where information is transmitted over many cell lengths. Smaller mitotic gradients reflect the need to organize several distinct events along the length of the mitotic spindle. The intracellular gradients that characterize mitosis are emerging as important regulatory paradigms. Intracellular gradients utilize intrinsic auto-regulatory feedback loops and diffusion to establish stable regions of activity within the mitotic cytosol. We review three recently described intracellular mitotic gradients. The Ran GTP gradient with its elaborate cascade of nuclear transport receptors and cargoes is the best characterized, yet the dynamics underlying the robust gradient of Ran-GTP have received little attention. Gradients of phosphorylation have been observed on Aurora B kinase substrates both before and after anaphase onset. In both instances the phosphorylation gradient appears to result from a soluble gradient of Aurora B kinase activity. Regulatory properties that support gradient formation are highlighted. Intracellular activity gradients that regulate localized mitotic events bare several hallmarks of self-organizing biologic systems that designate spatial information during pattern formation. Intracellular pattern formation represents a new paradigm in mitotic regulation.

S-Nitrosation destabilizes glutathione transferase P1-1.

Science.gov (United States)

Balchin, David; Stoychev, Stoyan H; Dirr, Heini W

2013-12-23

Protein S-nitrosation is a post-translational modification that regulates the function of more than 500 human proteins. Despite its apparent physiological significance, S-nitrosation is poorly understood at a molecular level. Here, we investigated the effect of S-nitrosation on the activity, structure, stability, and dynamics of human glutathione transferase P1-1 (GSTP1-1), an important detoxification enzyme ubiquitous in aerobes. S-Nitrosation at Cys47 and Cys101 reduces the activity of the enzyme by 94%. Circular dichroism spectroscopy, acrylamide quenching, and amide hydrogen-deuterium exchange mass spectrometry experiments indicate that the loss of activity is caused by the introduction of local disorder at the active site of GSTP1-1. Furthermore, the modification destabilizes domain 1 of GSTP1-1 against denaturation, smoothing the unfolding energy landscape of the protein and introducing a refolding defect. In contrast, S-nitrosation at Cys101 alone introduces a refolding defect in domain 1 but compensates by stabilizing the domain kinetically. These data elucidate the physical basis for the regulation of GSTP1-1 by S-nitrosation and provide general insight into the consequences of S-nitrosation on protein stability and dynamics.

Quasi-linear analysis of the extraordinary electron wave destabilized by runaway electrons

Energy Technology Data Exchange (ETDEWEB)

Pokol, G. I.; Kómár, A.; Budai, A. [Department of Nuclear Techniques, Budapest University of Technology and Economics, Budapest (Hungary); Stahl, A.; Fülöp, T. [Department of Applied Physics, Chalmers University of Technology, Göteborg (Sweden)

2014-10-15

Runaway electrons with strongly anisotropic distributions present in post-disruption tokamak plasmas can destabilize the extraordinary electron (EXEL) wave. The present work investigates the dynamics of the quasi-linear evolution of the EXEL instability for a range of different plasma parameters using a model runaway distribution function valid for highly relativistic runaway electron beams produced primarily by the avalanche process. Simulations show a rapid pitch-angle scattering of the runaway electrons in the high energy tail on the 100–1000 μs time scale. Due to the wave-particle interaction, a modification to the synchrotron radiation spectrum emitted by the runaway electron population is foreseen, exposing a possible experimental detection method for such an interaction.

Epithelial Cell Gene Expression Induced by Intracellular Staphylococcus aureus

Directory of Open Access Journals (Sweden)

Xianglu Li

2009-01-01

Full Text Available HEp-2 cell monolayers were cocultured with intracellular Staphylococcus aureus, and changes in gene expression were profiled using DNA microarrays. Intracellular S. aureus affected genes involved in cellular stress responses, signal transduction, inflammation, apoptosis, fibrosis, and cholesterol biosynthesis. Transcription of stress response and signal transduction-related genes including atf3, sgk, map2k1, map2k3, arhb, and arhe was increased. In addition, elevated transcription of proinflammatory genes was observed for tnfa, il1b, il6, il8, cxcl1, ccl20, cox2, and pai1. Genes involved in proapoptosis and fibrosis were also affected at transcriptional level by intracellular S. aureus. Notably, intracellular S. aureus induced strong transcriptional down-regulation of several cholesterol biosynthesis genes. These results suggest that epithelial cells respond to intracellular S. aureus by inducing genes affecting immunity and in repairing damage caused by the organism, and are consistent with the possibility that the organism exploits an intracellular environment to subvert host immunity and promote colonization.

The mass media destabilizes the cultural homogenous regime in Axelrod's model

Science.gov (United States)

Peres, Lucas R.; Fontanari, José F.

2010-02-01

An important feature of Axelrod's model for culture dissemination or social influence is the emergence of many multicultural absorbing states, despite the fact that the local rules that specify the agents interactions are explicitly designed to decrease the cultural differences between agents. Here we re-examine the problem of introducing an external, global interaction—the mass media—in the rules of Axelrod's model: in addition to their nearest neighbors, each agent has a certain probability p to interact with a virtual neighbor whose cultural features are fixed from the outset. Most surprisingly, this apparently homogenizing effect actually increases the cultural diversity of the population. We show that, contrary to previous claims in the literature, even a vanishingly small value of p is sufficient to destabilize the homogeneous regime for very large lattice sizes.

The mass media destabilizes the cultural homogenous regime in Axelrod's model

International Nuclear Information System (INIS)

Peres, Lucas R; Fontanari, Jose F

2010-01-01

An important feature of Axelrod's model for culture dissemination or social influence is the emergence of many multicultural absorbing states, despite the fact that the local rules that specify the agents interactions are explicitly designed to decrease the cultural differences between agents. Here we re-examine the problem of introducing an external, global interaction-the mass media-in the rules of Axelrod's model: in addition to their nearest neighbors, each agent has a certain probability p to interact with a virtual neighbor whose cultural features are fixed from the outset. Most surprisingly, this apparently homogenizing effect actually increases the cultural diversity of the population. We show that, contrary to previous claims in the literature, even a vanishingly small value of p is sufficient to destabilize the homogeneous regime for very large lattice sizes.

Destabilizing effect of alpha particles in a Maxwellian plasma

International Nuclear Information System (INIS)

Wang, M.Y.

1976-01-01

Various plasma waves which are possibly excited by MeV alphas have been investigated. For a delta birth distribution it is found that: a) The right-circularly polarized Alfven wave can be excited. Its growth rate is linearly proportional to the α-particle density. b) The drift Alfven wave is stable against α-particles. c) For a uniform temperature, the plasma wave spectrum changes from three branches with n/sub α/ = 0 to four branches for n/sub α/ not equal to 0 case. d) α-particles can destabilize the ion drift acoustic wave even with uniform temperature. However, the ion acoustic wave appears to be stable against fusion products in a fusion grade plasma. e) If their effect on the background plasma spectrum is neglected, α-particles can excite the electromagnetic cyclotron wave in a range of harmonics (band structure). The growth rate is proportional to the square root of α-particle density. f) If the effect of α-particle on the plasma spectrum is included, we find that electromagnetic cyclotron wave is stable

HYPERTHERMIA, INTRACELLULAR FREE CALCIUM AND CALCIUM IONOPHORES

NARCIS (Netherlands)

STEGE, GJJ; WIERENGA, PK; KAMPINGA, HH; KONINGS, AWT

1993-01-01

It is shown that heat-induced increase of intracellular calcium does not correlate with hyperthermic cell killing. Six different cell lines were investigated; in four (EAT, HeLa S3, L5178Y-R and L5178Y-S) heat treatments killing 90% of the cells did not affect the levels of intracellular free

Characterization of Leptin Intracellular Trafficking

Directory of Open Access Journals (Sweden)

E Walum

2009-12-01

Full Text Available Leptin is produced by adipose tissue, and its concentration in plasma is related to the amount of fat in the body. The leptin receptor (OBR is a member of the class I cytokine receptor family and several different isoforms, produced by alternative mRNA splicing are found in many tissues, including the hypothalamus. The two predominant isoforms includes a long form (OBRl with an intracellular domain of 303 amino acids and a shorter form (OBRs with an intracellular domain of 34 amino acids. Since OBRl is mainly expressed in the hypotalamus, it has been suggested to be the main signalling form. The peripheral production of leptin by adipocyte tissue and its effects as a signal of satiety in the central nervous system imply that leptin gains access to regions of the brain regulating in energy balance by crossing the blood-brain barrier. In an attempt to characterize the intracellular transport of leptin, we have followed binding internalization and degradation of leptin in HEK293 cells. We have also monitored the intracellular transport pathway of fluorescent conjugated leptin in HEK293 cells. Phenylarsine oxide, a general inhibitor of endocytosis, as well as incubation at mild hypertonic conditions, prevented the uptake of leptin, confirming a receptor-mediated internalization process. When internalized, 125I-leptin was rapidly accumulated inside the cells and reached a maximum after 10 min. After 70 minutes about 40-50% of total counts in each time point were found in the medium as TCA-soluble material. Leptin sorting, at the level of early endosomes, did not seem to involve recycling endosomes, since FITC-leptin was sorted from Cy3- transferrin containing compartments at 37°C. At 45 minutes of continuos internalization, FITC-leptin appeared mainly accumulated in late endocytic structures colocalizing with internalized rhodamine coupled epidermial growth factor (EGF and the lysosomal marker protein lamp-1. The transport of leptin was also shown

The cystic-fibrosis-associated ΔF508 mutation confers post-transcriptional destabilization on the C. elegans ABC transporter PGP-3

Directory of Open Access Journals (Sweden)

Liping He

2012-11-01

Membrane proteins make up ∼30% of the proteome. During the early stages of maturation, this class of proteins can experience localized misfolding in distinct cellular compartments, such as the cytoplasm, endoplasmic reticulum (ER lumen and ER membrane. ER quality control (ERQC mechanisms monitor folding and determine whether a membrane protein is appropriately folded or is misfolded and warrants degradation. ERQC plays crucial roles in human diseases, such as cystic fibrosis, in which deletion of a single amino acid (F508 results in the misfolding and degradation of the cystic fibrosis transmembrane conductance regulator (CFTR Cl– channel. We introduced the ΔF508 mutation into Caenorhabditis elegans PGP-3, a 12-transmembrane ABC transporter with 15% identity to CFTR. When expressed in intestinal epithelial cells, PGP-3wt was stable and efficiently trafficked to the apical plasma membrane through a COPII-dependent mechanism. However, PGP-3ΔF508 was post-transcriptionally destabilized, resulting in reduced total and apical membrane protein levels. Genetic or physiological activation of the osmotic stress response pathway, which causes accumulation of the chemical chaperone glycerol, stabilized PGP-3ΔF508. Efficient degradation of PGP-3ΔF508 required the function of several C. elegans ER-associated degradation (ERAD homologs, suggesting that destabilization occurs through an ERAD-type mechanism. Our studies show that the ΔF508 mutation causes post-transcriptional destabilization and degradation of PGP-3 in C. elegans epithelial cells. This model, combined with the power of C. elegans genetics, provides a new opportunity to genetically dissect metazoan ERQC.

Stochastic models of intracellular transport

KAUST Repository

Bressloff, Paul C.

2013-01-09

The interior of a living cell is a crowded, heterogenuous, fluctuating environment. Hence, a major challenge in modeling intracellular transport is to analyze stochastic processes within complex environments. Broadly speaking, there are two basic mechanisms for intracellular transport: passive diffusion and motor-driven active transport. Diffusive transport can be formulated in terms of the motion of an overdamped Brownian particle. On the other hand, active transport requires chemical energy, usually in the form of adenosine triphosphate hydrolysis, and can be direction specific, allowing biomolecules to be transported long distances; this is particularly important in neurons due to their complex geometry. In this review a wide range of analytical methods and models of intracellular transport is presented. In the case of diffusive transport, narrow escape problems, diffusion to a small target, confined and single-file diffusion, homogenization theory, and fractional diffusion are considered. In the case of active transport, Brownian ratchets, random walk models, exclusion processes, random intermittent search processes, quasi-steady-state reduction methods, and mean-field approximations are considered. Applications include receptor trafficking, axonal transport, membrane diffusion, nuclear transport, protein-DNA interactions, virus trafficking, and the self-organization of subcellular structures. © 2013 American Physical Society.

Exploring Anti-Bacterial Compounds against Intracellular Legionella

Science.gov (United States)

Harrison, Christopher F.; Kicka, Sébastien; Trofimov, Valentin; Berschl, Kathrin; Ouertatani-Sakouhi, Hajer; Ackermann, Nikolaus; Hedberg, Christian; Cosson, Pierre; Soldati, Thierry; Hilbi, Hubert

2013-01-01

Legionella pneumophila is a ubiquitous fresh-water bacterium which reproduces within its erstwhile predators, environmental amoeba, by subverting the normal pathway of phagocytosis and degradation. The molecular mechanisms which confer resistance to amoeba are apparently conserved and also allow replication within macrophages. Thus, L. pneumophila can act as an ‘accidental’ human pathogen and cause a severe pneumonia known as Legionnaires’ disease. The intracellular localisation of L. pneumophila protects it from some antibiotics, and this fact must be taken into account to develop new anti-bacterial compounds. In addition, the intracellular lifestyle of L. pneumophila may render the bacteria susceptible to compounds diminishing bacterial virulence and decreasing intracellular survival and replication of this pathogen. The development of a single infection cycle intracellular replication assay using GFP-producing L. pneumophila and Acanthamoeba castellanii amoeba is reported here. This fluorescence-based assay allows for continuous monitoring of intracellular replication rates, revealing the effect of bacterial gene deletions or drug treatment. To examine how perturbations of the host cell affect L. pneumophila replication, several known host-targeting compounds were tested, including modulators of cytoskeletal dynamics, vesicle scission and Ras GTPase localisation. Our results reveal a hitherto unrealized potential antibiotic property of the β-lactone-based Ras depalmitoylation inhibitor palmostatin M, but not the closely related inhibitor palmostatin B. Further characterisation indicated that this compound caused specific growth inhibition of Legionella and Mycobacterium species, suggesting that it may act on a common bacterial target. PMID:24058631

Exploring anti-bacterial compounds against intracellular Legionella.

Directory of Open Access Journals (Sweden)

Christopher F Harrison

Full Text Available Legionella pneumophila is a ubiquitous fresh-water bacterium which reproduces within its erstwhile predators, environmental amoeba, by subverting the normal pathway of phagocytosis and degradation. The molecular mechanisms which confer resistance to amoeba are apparently conserved and also allow replication within macrophages. Thus, L. pneumophila can act as an 'accidental' human pathogen and cause a severe pneumonia known as Legionnaires' disease. The intracellular localisation of L. pneumophila protects it from some antibiotics, and this fact must be taken into account to develop new anti-bacterial compounds. In addition, the intracellular lifestyle of L. pneumophila may render the bacteria susceptible to compounds diminishing bacterial virulence and decreasing intracellular survival and replication of this pathogen. The development of a single infection cycle intracellular replication assay using GFP-producing L. pneumophila and Acanthamoebacastellanii amoeba is reported here. This fluorescence-based assay allows for continuous monitoring of intracellular replication rates, revealing the effect of bacterial gene deletions or drug treatment. To examine how perturbations of the host cell affect L. pneumophila replication, several known host-targeting compounds were tested, including modulators of cytoskeletal dynamics, vesicle scission and Ras GTPase localisation. Our results reveal a hitherto unrealized potential antibiotic property of the β-lactone-based Ras depalmitoylation inhibitor palmostatin M, but not the closely related inhibitor palmostatin B. Further characterisation indicated that this compound caused specific growth inhibition of Legionella and Mycobacterium species, suggesting that it may act on a common bacterial target.

Destabilization of Akt Promotes the Death of Myeloma Cell Lines

Directory of Open Access Journals (Sweden)

Yanan Zhang

2014-01-01

Full Text Available Constitutive activation of Akt is believed to be an oncogenic signal in multiple myeloma and is associated with poor patient prognosis and resistance to available treatment. The stability of Akt proteins is regulated by phosphorylating the highly conserved turn motif (TM of these proteins and the chaperone protein HSP90. In this study we investigate the antitumor effects of inhibiting mTORC2 plus HSP90 in myeloma cell lines. We show that chronic exposure of cells to rapamycin can inhibit mTORC2 pathway, and AKT will be destabilized by administration of the HSP90 inhibitor 17-allylamino-geldanamycin (17-AAG. Finally, we show that the rapamycin synergizes with 17-AAG and inhibits myeloma cells growth and promotes cell death to a greater extent than either drug alone. Our studies provide a clinical rationale of use mTOR inhibitors and chaperone protein inhibitors in combination regimens for the treatment of human blood cancers.

Intracellular calcium levels can regulate Importin-dependent nuclear import

International Nuclear Information System (INIS)

Kaur, Gurpreet; Ly-Huynh, Jennifer D.; Jans, David A.

2014-01-01

Highlights: • High intracellular calcium inhibits Impα/β1- or Impβ1-dependent nuclear protein import. • The effect of Ca 2+ on nuclear import does not relate to changes in the nuclear pore. • High intracellular calcium can result in mislocalisation of Impβ1, Ran and RCC1. - Abstract: We previously showed that increased intracellular calcium can modulate Importin (Imp)β1-dependent nuclear import of SRY-related chromatin remodeling proteins. Here we extend this work to show for the first time that high intracellular calcium inhibits Impα/β1- or Impβ1-dependent nuclear protein import generally. The basis of this relates to the mislocalisation of the transport factors Impβ1 and Ran, which show significantly higher nuclear localization in contrast to various other factors, and RCC1, which shows altered subnuclear localisation. The results here establish for the first time that intracellular calcium modulates conventional nuclear import through direct effects on the nuclear transport machinery

Intracellular calcium levels can regulate Importin-dependent nuclear import

Energy Technology Data Exchange (ETDEWEB)

Kaur, Gurpreet; Ly-Huynh, Jennifer D.; Jans, David A., E-mail: David.Jans@monash.edu

2014-07-18

Highlights: • High intracellular calcium inhibits Impα/β1- or Impβ1-dependent nuclear protein import. • The effect of Ca{sup 2+} on nuclear import does not relate to changes in the nuclear pore. • High intracellular calcium can result in mislocalisation of Impβ1, Ran and RCC1. - Abstract: We previously showed that increased intracellular calcium can modulate Importin (Imp)β1-dependent nuclear import of SRY-related chromatin remodeling proteins. Here we extend this work to show for the first time that high intracellular calcium inhibits Impα/β1- or Impβ1-dependent nuclear protein import generally. The basis of this relates to the mislocalisation of the transport factors Impβ1 and Ran, which show significantly higher nuclear localization in contrast to various other factors, and RCC1, which shows altered subnuclear localisation. The results here establish for the first time that intracellular calcium modulates conventional nuclear import through direct effects on the nuclear transport machinery.

Mechanism of duplex DNA destabilization by RNA-guided Cas9 nuclease during target interrogation.

Science.gov (United States)

Mekler, Vladimir; Minakhin, Leonid; Severinov, Konstantin

2017-05-23

The prokaryotic clustered regularly interspaced short palindromic repeats (CRISPR)-associated 9 (Cas9) endonuclease cleaves double-stranded DNA sequences specified by guide RNA molecules and flanked by a protospacer adjacent motif (PAM) and is widely used for genome editing in various organisms. The RNA-programmed Cas9 locates the target site by scanning genomic DNA. We sought to elucidate the mechanism of initial DNA interrogation steps that precede the pairing of target DNA with guide RNA. Using fluorometric and biochemical assays, we studied Cas9/guide RNA complexes with model DNA substrates that mimicked early intermediates on the pathway to the final Cas9/guide RNA-DNA complex. The results show that Cas9/guide RNA binding to PAM favors separation of a few PAM-proximal protospacer base pairs allowing initial target interrogation by guide RNA. The duplex destabilization is mediated, in part, by Cas9/guide RNA affinity for unpaired segments of nontarget strand DNA close to PAM. Furthermore, our data indicate that the entry of double-stranded DNA beyond a short threshold distance from PAM into the Cas9/single-guide RNA (sgRNA) interior is hindered. We suggest that the interactions unfavorable for duplex DNA binding promote DNA bending in the PAM-proximal region during early steps of Cas9/guide RNA-DNA complex formation, thus additionally destabilizing the protospacer duplex. The mechanism that emerges from our analysis explains how the Cas9/sgRNA complex is able to locate the correct target sequence efficiently while interrogating numerous nontarget sequences associated with correct PAMs.

Modeling HIV-1 intracellular replication: two simulation approaches

NARCIS (Netherlands)

Zarrabi, N.; Mancini, E.; Tay, J.; Shahand, S.; Sloot, P.M.A.

2010-01-01

Many mathematical and computational models have been developed to investigate the complexity of HIV dynamics, immune response and drug therapy. However, there are not many models which consider the dynamics of virus intracellular replication at a single level. We propose a model of HIV intracellular

Experimental study of the stabilization process of a non-premixed flame via the destabilization analysis of the blue ring flame

Energy Technology Data Exchange (ETDEWEB)

Pinguet, Guillaume; Escudie, Dany [Centre de Thermique de Lyon (CETHIL) UMR 5008 CNRS-INSA-UCBL, INSA de Lyon, 20 av. A. Einstein, 69621 Villeurbanne cedex (France)

2007-04-15

The flame stabilization phenomenon remains a crucial issue. The experimental study of flame stabilization behind a tulip-shaped flame-holder is addressed in this paper. The process leading to the transition between specific modes - the blue ring flame and the instable ring - of a non-premixed flame stabilized on a tulip-shaped bluff-body is detailed. The aim of this study is to provide an accurate description of the destabilization of specific combustion modes, which enables a further understanding of the entire stabilization mechanism. The aerodynamic and mixing fields are described by laser Doppler anemometry and concentration measurements by sampling probe respectively. The behaviour of shear layers developing at the wake and jet boundaries are characterized by means of a spectral analysis of the fluctuating radial velocity. Results show that the destabilization process is related to the intensification of hot gas recirculation, inducing an upheaval of the dynamical condition of stabilization and a transition of mixing phenomena. (author)

Single-cell intracellular nano-pH probes†

Science.gov (United States)

Özel, Rıfat Emrah; Lohith, Akshar; Mak, Wai Han; Pourmand, Nader

2016-01-01

Within a large clonal population, such as cancerous tumor entities, cells are not identical, and the differences between intracellular pH levels of individual cells may be important indicators of heterogeneity that could be relevant in clinical practice, especially in personalized medicine. Therefore, the detection of the intracellular pH at the single-cell level is of great importance to identify and study outlier cells. However, quantitative and real-time measurements of the intracellular pH of individual cells within a cell population is challenging with existing technologies, and there is a need to engineer new methodologies. In this paper, we discuss the use of nanopipette technology to overcome the limitations of intracellular pH measurements at the single-cell level. We have developed a nano-pH probe through physisorption of chitosan onto hydroxylated quartz nanopipettes with extremely small pore sizes (~100 nm). The dynamic pH range of the nano-pH probe was from 2.6 to 10.7 with a sensitivity of 0.09 units. We have performed single-cell intracellular pH measurements using non-cancerous and cancerous cell lines, including human fibroblasts, HeLa, MDA-MB-231 and MCF-7, with the pH nanoprobe. We have further demonstrated the real-time continuous single-cell pH measurement capability of the sensor, showing the cellular pH response to pharmaceutical manipulations. These findings suggest that the chitosan-functionalized nanopore is a powerful nano-tool for pH sensing at the single-cell level with high temporal and spatial resolution. PMID:27708772

Single-cell intracellular nano-pH probes.

Science.gov (United States)

Özel, Rıfat Emrah; Lohith, Akshar; Mak, Wai Han; Pourmand, Nader

2015-01-01

Within a large clonal population, such as cancerous tumor entities, cells are not identical, and the differences between intracellular pH levels of individual cells may be important indicators of heterogeneity that could be relevant in clinical practice, especially in personalized medicine. Therefore, the detection of the intracellular pH at the single-cell level is of great importance to identify and study outlier cells. However, quantitative and real-time measurements of the intracellular pH of individual cells within a cell population is challenging with existing technologies, and there is a need to engineer new methodologies. In this paper, we discuss the use of nanopipette technology to overcome the limitations of intracellular pH measurements at the single-cell level. We have developed a nano-pH probe through physisorption of chitosan onto hydroxylated quartz nanopipettes with extremely small pore sizes (~100 nm). The dynamic pH range of the nano-pH probe was from 2.6 to 10.7 with a sensitivity of 0.09 units. We have performed single-cell intracellular pH measurements using non-cancerous and cancerous cell lines, including human fibroblasts, HeLa, MDA-MB-231 and MCF-7, with the pH nanoprobe. We have further demonstrated the real-time continuous single-cell pH measurement capability of the sensor, showing the cellular pH response to pharmaceutical manipulations. These findings suggest that the chitosan-functionalized nanopore is a powerful nano-tool for pH sensing at the single-cell level with high temporal and spatial resolution.

Mycobacterium intracellulare Infection Mimicking Progression of Scleroderma

DEFF Research Database (Denmark)

Krabbe, Simon; Engelhart, Merete; Thybo, Sören

2017-01-01

This case report describes a patient with scleroderma who developed Mycobacterium intracellulare infection, which for more than a year mimicked worsening of her connective tissue disorder. The patient was diagnosed with scleroderma based on puffy fingers that developed into sclerodactyly, abnormal......, unfortunately with significant scarring. Immunodeficiency testing was unremarkable. In summary, an infection with Mycobacterium intracellulare was mistaken for an unusually severe progression of scleroderma....

Improvement of the myocardial performance index in atrial fibrilation patients treated with amiodarone after cardioversion.

Science.gov (United States)

Besli, Feyzullah; Basar, Cengiz; Kecebas, Mesut; Turker, Yasin

2015-03-01

This study evaluated the response to electrical cardioversion (EC) and the effect on the myocardial performance index (MPI) in patients with persistent and long-standing persistent atrial fibrillation (AF). We enrolled 103 patients (mean age 69.6 ± 8.9 years, 40.7% males) with a diagnosis of persistent and long-standing persistent AF. EC was applied to all patients after one g of amiodarone administration. Echocardiographic findings before EC were compared in patients with successful versus unsuccessful cardioversions and in patients with maintained sinus rhythm (SR) versus those with AF recurrence at the end of the first month. We also compared echocardiographic data before EC versus at the end of the first month in the same patients with maintained SR. SR was achieved in 72.8% of patients and was continued at the end of the first month in 69.3% of the patients. The MPI value of all patients was found to be 0.73 ± 0.21. The size of the left atrium was determined to be an independent predictor of the maintenance of SR at 1 month. In subgroup analyses, when we compared echocardiographic findings before EC and at the end of the first month in patients with maintained SR, the MPI (0.66 ± 0.14 vs 0.56 ± 0.09, p < 0.001) values were significantly decreased. Our study is the first to show impairment of the MPI, which is an indicator of systolic and diastolic function, in patients with persistent and long-standing persistent AF and improvement of the MPI after successful EC.

Uptake and intracellular activity of AM-1155 in phagocytic cells.

Science.gov (United States)

Yamamoto, T; Kusajima, H; Hosaka, M; Fukuda, H; Oomori, Y; Shinoda, H

1996-01-01

The uptake and intracellular activity of AM-1155 in murine J774.1 macrophages and human polymorphonuclear leukocytes were investigated. AM-1155 penetrated phagocytic cells rapidly and reversibly, although the penetration process was not affected by metabolic inhibitors such as sodium fluoride, cyanide m-chlorophenylhydrazone, or ouabain or by nucleoside transport system inhibitors such as adenosine. The intracellular concentration-to-extracellular concentration ratio of AM-1155 in both cell types of phagocytes ranged from 5 to 7. These ratios were almost equal to those for sparfloxacin. The intracellular activity of AM-1155 in J774.1 macrophages, examined with Staphylococcus aureus 209P as a test bacterium, was dependent on the extracellular concentration. AM-1155 at a concentration of 1 microgram/ml reduced the number of viable cells of S. aureus ingested by more than 90%. The intracellular activity of AM-1155 was more potent than those of sparfloxacin, ofloxacin, ciprofloxacin, flomoxef, and erythromycin. These results suggest that the potent intracellular activity of AM-1155 might mainly be due to the high intracellular concentration and its potent in vitro activity. PMID:9124835

Relevance of intracellular polarity to accuracy of eukaryotic chemotaxis

International Nuclear Information System (INIS)

Hiraiwa, Tetsuya; Nishikawa, Masatoshi; Shibata, Tatsuo; Nagamatsu, Akihiro; Akuzawa, Naohiro

2014-01-01

Eukaryotic chemotaxis is usually mediated by intracellular signals that tend to localize at the front or back of the cell. Such intracellular polarities frequently require no extracellular guidance cues, indicating that spontaneous polarization occurs in the signal network. Spontaneous polarization activity is considered relevant to the persistent motions in random cell migrations and chemotaxis. In this study, we propose a theoretical model that connects spontaneous intracellular polarity and motile ability in a chemoattractant solution. We demonstrate that the intracellular polarity can enhance the accuracy of chemotaxis. Chemotactic accuracy should also depend on chemoattractant concentration through the concentration-dependent correlation time in the polarity direction. Both the polarity correlation time and the chemotactic accuracy depend on the degree of responsiveness to the chemical gradient. We show that optimally accurate chemotaxis occurs at an intermediate responsiveness of intracellular polarity. Experimentally, we find that the persistence time of randomly migrating Dictyostelium cells depends on the chemoattractant concentration, as predicted by our theory. At the optimum responsiveness, this ameboid cell can enhance its chemotactic accuracy tenfold. (paper)

Destabilization in the isomeric nitrobenzonitriles: an experimental thermochemical study

International Nuclear Information System (INIS)

Roux, Maria Victoria; Jimenez, Pilar; Davalos, Juan Z.; Temprado, Manuel; Liebman, Joel F.

2003-01-01

The enthalpies of combustion and of sublimation, respectively, of the three isomeric nitrobenzonitriles have been measured: o-, {(-3456.3±2.9), (88.1±1.4)} kJ·mol -1 ; m-, {(-3442.8±3.3), (92.8±0.3)} kJ·mol -1 ; p-, {(-3448.2±3.6), (91.1±1.3)} kJ·mol -1 . In turn, from these values, the standard molar enthalpies of formation for the condensed and gaseous state, respectively, have been derived: o-, {(130.1±3.1), (218.2±3.4)} kJ·mol -1 ; m-, {(116.5±3.5), (209.3±3.5)} kJ·mol -1 ; p-, {(122.0±3.8), (213.1±4.0)} kJ·mol -1 . Destabilization energies associated with the presence of the two electron-withdrawing groups have been determined, for o-, m-, and p-nitrobenzonitrile, {(17.6±4.1), (8.7±4.2), and (12.5±4.6)} kJ·mol -1 , respectively, and are consistent with those obtained for the corresponding sets of isomeric methyl benzenedicarboxylates, dicyanobenzenes, dinitrobenzenes, and (neutral and ionized) nitrobenzoic acids

Endogenous ribosomal frameshift signals operate as mRNA destabilizing elements through at least two molecular pathways in yeast.

Science.gov (United States)

Belew, Ashton T; Advani, Vivek M; Dinman, Jonathan D

2011-04-01

Although first discovered in viruses, previous studies have identified operational -1 ribosomal frameshifting (-1 RF) signals in eukaryotic genomic sequences, and suggested a role in mRNA stability. Here, four yeast -1 RF signals are shown to promote significant mRNA destabilization through the nonsense mediated mRNA decay pathway (NMD), and genetic evidence is presented suggesting that they may also operate through the no-go decay pathway (NGD) as well. Yeast EST2 mRNA is highly unstable and contains up to five -1 RF signals. Ablation of the -1 RF signals or of NMD stabilizes this mRNA, and changes in -1 RF efficiency have opposing effects on the steady-state abundance of the EST2 mRNA. These results demonstrate that endogenous -1 RF signals function as mRNA destabilizing elements through at least two molecular pathways in yeast. Consistent with current evolutionary theory, phylogenetic analyses suggest that -1 RF signals are rapidly evolving cis-acting regulatory elements. Identification of high confidence -1 RF signals in ∼10% of genes in all eukaryotic genomes surveyed suggests that -1 RF is a broadly used post-transcriptional regulator of gene expression.

Engineering FKBP-Based Destabilizing Domains to Build Sophisticated Protein Regulation Systems.

Directory of Open Access Journals (Sweden)

Wenlin An

Full Text Available Targeting protein stability with small molecules has emerged as an effective tool to control protein abundance in a fast, scalable and reversible manner. The technique involves tagging a protein of interest (POI with a destabilizing domain (DD specifically controlled by a small molecule. The successful construction of such fusion proteins may, however, be limited by functional interference of the DD epitope with electrostatic interactions required for full biological function of proteins. Another drawback of this approach is the remaining endogenous protein. Here, we combined the Cre-LoxP system with an advanced DD and generated a protein regulation system in which the loss of an endogenous protein, in our case the tumor suppressor PTEN, can be coupled directly with a conditionally fine-tunable DD-PTEN. This new system will consolidate and extend the use of DD-technology to control protein function precisely in living cells and animal models.

Analysis of Intracellular Metabolites from Microorganisms: Quenching and Extraction Protocols.

Science.gov (United States)

Pinu, Farhana R; Villas-Boas, Silas G; Aggio, Raphael

2017-10-23

Sample preparation is one of the most important steps in metabolome analysis. The challenges of determining microbial metabolome have been well discussed within the research community and many improvements have already been achieved in last decade. The analysis of intracellular metabolites is particularly challenging. Environmental perturbations may considerably affect microbial metabolism, which results in intracellular metabolites being rapidly degraded or metabolized by enzymatic reactions. Therefore, quenching or the complete stop of cell metabolism is a pre-requisite for accurate intracellular metabolite analysis. After quenching, metabolites need to be extracted from the intracellular compartment. The choice of the most suitable metabolite extraction method/s is another crucial step. The literature indicates that specific classes of metabolites are better extracted by different extraction protocols. In this review, we discuss the technical aspects and advancements of quenching and extraction of intracellular metabolite analysis from microbial cells.

Influence of intracellular acidosis on contractile function in the working rat heart

International Nuclear Information System (INIS)

Jeffrey, F.M.H.; Malloy, C.R.; Radda, G.K.

1987-01-01

The decrease in myocardial contractility during ischemia, hypoxia, and extracellular acidosis has been attributed to intracellular acidosis. Previous studies of the relationship between pH and contractile state have utilized respiratory or metabolic acidosis to alter intracellular pH. The authors developed a model in the working perfused rat heart to study the effects of intracellular acidosis with normal external pH and optimal O 2 delivery. Intracellular pH and high-energy phosphates were monitored by 31 P nuclear magnetic resonance spectroscopy. Hearts were perfused to a steady state with a medium containing 10 mM NH 4 Cl. Acidosis induced a substantial decrease in aortic flow and stroke volume which was associated with little change in peak systolic pressure. It was concluded that (1) for the same intracellular acidosis the influence on tension development was more pronounced with a combined extra- and intracellular acidosis than with an isolated intracellular acidosis, and (2) stroke volume at constant preload was impaired by intracellular acidosis even though changes in developed pressure were minimal. These observations suggest that isolated intracellular acidosis has adverse effects on diastolic compliance and/or relaxation

Intracellular trafficking of new anticancer therapeutics: antibody–drug conjugates

Science.gov (United States)

Kalim, Muhammad; Chen, Jie; Wang, Shenghao; Lin, Caiyao; Ullah, Saif; Liang, Keying; Ding, Qian; Chen, Shuqing; Zhan, Jinbiao

2017-01-01

Antibody–drug conjugate (ADC) is a milestone in targeted cancer therapy that comprises of monoclonal antibodies chemically linked to cytotoxic drugs. Internalization of ADC takes place via clathrin-mediated endocytosis, caveolae-mediated endocytosis, and pinocytosis. Conjugation strategies, endocytosis and intracellular trafficking optimization, linkers, and drugs chemistry present a great challenge for researchers to eradicate tumor cells successfully. This inventiveness of endocytosis and intracellular trafficking has given considerable momentum recently to develop specific antibodies and ADCs to treat cancer cells. It is significantly advantageous to emphasize the endocytosis and intracellular trafficking pathways efficiently and to design potent engineered conjugates and biological entities to boost efficient therapies enormously for cancer treatment. Current studies illustrate endocytosis and intracellular trafficking of ADC, protein, and linker strategies in unloading and also concisely evaluate practically applicable ADCs. PMID:28814834

Intracellular trafficking of new anticancer therapeutics: antibody-drug conjugates.

Science.gov (United States)

Kalim, Muhammad; Chen, Jie; Wang, Shenghao; Lin, Caiyao; Ullah, Saif; Liang, Keying; Ding, Qian; Chen, Shuqing; Zhan, Jinbiao

2017-01-01

Antibody-drug conjugate (ADC) is a milestone in targeted cancer therapy that comprises of monoclonal antibodies chemically linked to cytotoxic drugs. Internalization of ADC takes place via clathrin-mediated endocytosis, caveolae-mediated endocytosis, and pinocytosis. Conjugation strategies, endocytosis and intracellular trafficking optimization, linkers, and drugs chemistry present a great challenge for researchers to eradicate tumor cells successfully. This inventiveness of endocytosis and intracellular trafficking has given considerable momentum recently to develop specific antibodies and ADCs to treat cancer cells. It is significantly advantageous to emphasize the endocytosis and intracellular trafficking pathways efficiently and to design potent engineered conjugates and biological entities to boost efficient therapies enormously for cancer treatment. Current studies illustrate endocytosis and intracellular trafficking of ADC, protein, and linker strategies in unloading and also concisely evaluate practically applicable ADCs.

Spatial Cytoskeleton Organization Supports Targeted Intracellular Transport

Science.gov (United States)

Hafner, Anne E.; Rieger, Heiko

2018-03-01

The efficiency of intracellular cargo transport from specific source to target locations is strongly dependent upon molecular motor-assisted motion along the cytoskeleton. Radial transport along microtubules and lateral transport along the filaments of the actin cortex underneath the cell membrane are characteristic for cells with a centrosome. The interplay between the specific cytoskeleton organization and the motor performance realizes a spatially inhomogeneous intermittent search strategy. In order to analyze the efficiency of such intracellular search strategies we formulate a random velocity model with intermittent arrest states. We evaluate efficiency in terms of mean first passage times for three different, frequently encountered intracellular transport tasks: i) the narrow escape problem, which emerges during cargo transport to a synapse or other specific region of the cell membrane, ii) the reaction problem, which considers the binding time of two particles within the cell, and iii) the reaction-escape problem, which arises when cargo must be released at a synapse only after pairing with another particle. Our results indicate that cells are able to realize efficient search strategies for various intracellular transport tasks economically through a spatial cytoskeleton organization that involves only a narrow actin cortex rather than a cell body filled with randomly oriented actin filaments.

Analysis of Intracellular Metabolites from Microorganisms: Quenching and Extraction Protocols

Directory of Open Access Journals (Sweden)

Farhana R. Pinu

2017-10-01

Full Text Available Sample preparation is one of the most important steps in metabolome analysis. The challenges of determining microbial metabolome have been well discussed within the research community and many improvements have already been achieved in last decade. The analysis of intracellular metabolites is particularly challenging. Environmental perturbations may considerably affect microbial metabolism, which results in intracellular metabolites being rapidly degraded or metabolized by enzymatic reactions. Therefore, quenching or the complete stop of cell metabolism is a pre-requisite for accurate intracellular metabolite analysis. After quenching, metabolites need to be extracted from the intracellular compartment. The choice of the most suitable metabolite extraction method/s is another crucial step. The literature indicates that specific classes of metabolites are better extracted by different extraction protocols. In this review, we discuss the technical aspects and advancements of quenching and extraction of intracellular metabolite analysis from microbial cells.

Methanolic extract of Moringa oleifera leaf and low doses of gamma radiation alleviated amiodarone-induced lung toxicity in albino rats

Directory of Open Access Journals (Sweden)

Hasan Hesham F.

2016-01-01

Full Text Available This study aimed to evaluate the effects of methanolic extract of Moringa oleifera (MO and/or low doses of gamma radiation (LDR on amiodarone (AMD-induced lung toxicity in rats. AMD administered to female albino rats (100 mg/kg body weight for 10 consecutive days. Rats received methanolic extract of MO (250 mg/kg bwt for 15 successive days and/or were exposed to whole body LDR (0.25Gy on the 1st and 10th days, up to a total dose of 0.5Gy. MO administration induced a significant decrease in serum tumor necrosis factor-alpha (TNF-α and transforming growth factor-beta (TGF-β levels as well as lactate dehydrogenase (LDH activity. Also, the content of malondialdehyde (MDA and hydroxyproline (HYP was significantly decreased in lung tissue. Furthermore, MO significantly increased reduced glutathione (GSH content in lung tissue as compared with AMD. The histopathological investigation of lung tissue revealed the appearance of interstitial pneumonia in rats treated with AMD. The oral administration of MO and/or exposure to LDR reversed the biochemical and histopathological alterations induced by AMD. It can be posited that MO and LDR might have a considerable role in the prevention of lung toxicity induced by AMD.

Manipulation of Host Cholesterol by Obligate Intracellular Bacteria

Directory of Open Access Journals (Sweden)

Dhritiman Samanta

2017-05-01

Full Text Available Cholesterol is a multifunctional lipid that plays important metabolic and structural roles in the eukaryotic cell. Despite having diverse lifestyles, the obligate intracellular bacterial pathogens Chlamydia, Coxiella, Anaplasma, Ehrlichia, and Rickettsia all target cholesterol during host cell colonization as a potential source of membrane, as well as a means to manipulate host cell signaling and trafficking. To promote host cell entry, these pathogens utilize cholesterol-rich microdomains known as lipid rafts, which serve as organizational and functional platforms for host signaling pathways involved in phagocytosis. Once a pathogen gains entrance to the intracellular space, it can manipulate host cholesterol trafficking pathways to access nutrient-rich vesicles or acquire membrane components for the bacteria or bacteria-containing vacuole. To acquire cholesterol, these pathogens specifically target host cholesterol metabolism, uptake, efflux, and storage. In this review, we examine the strategies obligate intracellular bacterial pathogens employ to manipulate cholesterol during host cell colonization. Understanding how obligate intracellular pathogens target and use host cholesterol provides critical insight into the host-pathogen relationship.

A Thapsigargin-Resistant Intracellular Calcium Sequestering Compartment in Rat Brain

Science.gov (United States)

2000-03-31

have a major impact on neuronal intracellular signaling. Most of the ER in neurons and glia appears to accumulate calcium by energy driven ion pumps...secretion of exocrine, endocrine, and neurocrine products, regulation of glycogenolysis and gluconeogenesis , intracellular transport, secretion of fluids...the RyRs [140]. Furthermore, the intracellular expression of these receptor-channels in neuronal ER is also reciprocal with RyRs located primarily in

Nuclear magnetic resonance studies of intracellular ions in perfused from heart

International Nuclear Information System (INIS)

Burnstein, D.; Fossel, E.T.

1987-01-01

Intracellular sodium, potassium, and lithium were observed in a perfused frog heart by nuclear magnetic resonance (NMR) spectroscopy. A perfusate buffer containing the shift reagent, dysprosium tripolyphosphate, was used in combination with mathematical filtering or presaturation of the extracellular resonance to separate the intra- and extracellular sodium NMR signals. Addition of 10 μM ouabain to the perfusate, perfusion with a zero potassium, low-calcium buffer, and replacement of 66% of the perfusate sodium with lithium resulted in changes in the intracellular sodium levels. An increase of 45% in the intracellular sodium was observed when changing the pacing rate from 0 to 60 beats/min (with proportional changes for intermediate pacing rates). The ratio of intracellular potassium to sodium concentration was determined to be 2.3 by NMR, indicating that a substantial amount of the intracellular potassium is undetectable with these NMR method. In addition, intracellular lithium was observed during perfusion with a lithium-containing perfusate

Intracellular pH homeostasis in Leishmania donovani amastigotes and promastigotes

International Nuclear Information System (INIS)

Glaser, T.A.; Baatz, J.E.; Kreishman, G.P.; Mukkada, A.J.

1988-01-01

Intracellular pH and pH gradients of Leishmania donovani amastigotes and promastigotes were determined over a broad range of extracellular pH values. Intracellular pH was determined by 31 P NMR and by equilibrium distribution studies with 5,5-dimethyloxazolidine-2,4-dione or methylamine. Promastigotes maintain intracellular pH values close to neutral between extracellular pH values of 5.0 and 7.4. Amastigote intracellular pH is maintained close to neutral at external pH values as low as 4.0. Both life stages maintain a positive pH gradient to an extracellular pH of 7.4, which is important for active transport of substrates. Treatment with ionophores, such as nigericin and carbonyl cyanide m-chlorophenylhydrazone and the ATPase inhibitor dicyclohexylcarbodiimide, reduced pH gradients in both stages. Maintenance of intracellular pH in the physiologic range is especially relevant for the survival of the amastigote in its acidic in vivo environment

Crystallization and preliminary electron diffraction study to 3. 7 A of DNA helix-destabilizing protein gp32*I

Energy Technology Data Exchange (ETDEWEB)

Chiu, W; Hosoda, J

1978-01-01

A two-dimensionally large and thin crystal has been obtained from gp32*I, a proteolytically digested product of a DNA helix-destabilizing protein coded by gene 32 in bacteriophage T4. High-resolution electron diffraction patterns (approx. 3.7 A) are recorded from both unstained and stained protein crystals embedded in glucose. The crystal is of orthorhombic space group with a = 62.9 A and b = 47.3 A.

Is the posterior cruciate ligament destabilized after the tibial cut in a cruciate retaining total knee replacement? An anatomical study.

Science.gov (United States)

Liabaud, Barthelemy; Patrick, David A; Geller, Jeffrey A

2013-12-01

Cruciate retaining total knee replacement has been shown to effectively improve pain and quality of life. Successful outcomes depend on many factors, including the maintenance of a competent posterior cruciate ligament. This study sought to anatomically analyze the percentage of PCL injured during a full transverse, tibial cut, thus altering normal function. One hundred and thirty five consecutive knee MRIs taken from 2006 to 2011 were selected from a single surgeon's database for this study. Only subjects with non-arthritic knees were considered for this study; the lack of degenerative joint disease (DJD) was confirmed via a radiological report. The optimal view of the PCL's tibial attachment was observed using the sagittal view of the knee, with a T1 signal. One hundred and twenty two usable images were viewed electronically, and measurements were made using the standardized transverse cut implant guidelines. The percentage of PCL remaining following the cut was categorized into five different groups: 0% (no PCL undermined), 1-49%, 50-74%, 75-99% and 100% (PCL undermined entirely). Overall only 9.0% (n=11) would have not endured any damage to the PCL with a transverse tibial saw cut, while 79.6% (n=98) would have had 50% or more of the PCL undermined. Of the 98 patients with more than 50% resected, 52.1% (n=51 patients) presented complete destabilization of the PCL. The percentage of PCL destabilized was not significant across age groups (p=0.280), gender (p=0.586), or operative side (p=0.460). Independent of age, gender, and operative side, a majority of PCLs are more than 50% destabilized following the standard transverse tibial cut. II. Copyright © 2013 Elsevier B.V. All rights reserved.

Study of the man-caused destabilization consequences to the Semipalatinsk test site subsurface

International Nuclear Information System (INIS)

Gorbunova, Eh.M.

2004-01-01

Conduction of underground nuclear explosions (UNE) at the territory of the Semipalatinsk Test Site (STS) has led to irreversible deformation of the geological environment. The carried out experimental researches on study of the UNE effect on the rocks massif allowed to ascertain the basic mechanisms of man-caused subsurface destabilization, which is a change in the physic-mechanical characteristics and filtering structure of the background medium, hydrogeodynamic and radiation situation. The consequences of the post-explosion deformations detected in the massif conjugate not only with the epicenter zone of UNE, but are also related to the structural boundaries of various range (to faults, lithologic contacts, boundaries of inhomogeneous media). The consequences were partially detected on the ground surface as well. (author)

Directed antigen delivery as a vaccine strategy for an intracellular bacterial pathogen

Science.gov (United States)

Bouwer, H. G. Archie; Alberti-Segui, Christine; Montfort, Megan J.; Berkowitz, Nathan D.; Higgins, Darren E.

2006-03-01

We have developed a vaccine strategy for generating an attenuated strain of an intracellular bacterial pathogen that, after uptake by professional antigen-presenting cells, does not replicate intracellularly and is readily killed. However, after degradation of the vaccine strain within the phagolysosome, target antigens are released into the cytosol for endogenous processing and presentation for stimulation of CD8+ effector T cells. Applying this strategy to the model intracellular pathogen Listeria monocytogenes, we show that an intracellular replication-deficient vaccine strain is cleared rapidly in normal and immunocompromised animals, yet antigen-specific CD8+ effector T cells are stimulated after immunization. Furthermore, animals immunized with the intracellular replication-deficient vaccine strain are resistant to lethal challenge with a virulent WT strain of L. monocytogenes. These studies suggest a general strategy for developing safe and effective, attenuated intracellular replication-deficient vaccine strains for stimulation of protective immune responses against intracellular bacterial pathogens. CD8+ T cell | replication-deficient | Listeria monocytogenes

Intracellular Enzymes Contribution to the Biocatalytic Removal of Pharmaceuticals by Trametes hirsuta.

Science.gov (United States)

Haroune, Lounès; Saibi, Sabrina; Cabana, Hubert; Bellenger, Jean-Philippe

2017-01-17

The use of white rot fungi (WRF) for bioremediation of recalcitrant trace organic contaminants (TrOCs) is becoming greatly popular. Biosorption and lignin modifying enzymes (LMEs) are the most often reported mechanisms of action. Intracellular enzymes, such as cytochrome P450 (CYP450), have also been suggested to contribute. However, direct evidence of TrOCs uptake and intracellular transformation is lacking. The aim of this study was to evaluate the relative contribution of biosorption, extracellular LMEs activity, TrOCs uptake, and intracellular CYP450 on the removal of six nonsteroidal anti-inflammatories (NSAIs) by Trametes hirsuta. Results show that for most tested NSAIs, LMEs activity and biosorption failed to explain the observed removal. Most tested TrOCs are quickly taken up and intracellularly transformed. Fine characterization of intracellular transformation using ketoprofen showed that CYP450 is not the sole intracellular enzyme responsible for intracellular transformation. The contribution of CYP450 in further transformation of ketoprofen byproducts is also reported. These results illustrate that TrOCs transformation by WRF is a more complex process than previously reported. Rapid uptake of TrOCs and intracellular transformation through diverse enzymatic systems appears to be important components of WRF efficiency toward TrOCs.

Ortholog-based screening and identification of genes related to intracellular survival.

Science.gov (United States)

Yang, Xiaowen; Wang, Jiawei; Bing, Guoxia; Bie, Pengfei; De, Yanyan; Lyu, Yanli; Wu, Qingmin

2018-04-20

Bioinformatics and comparative genomics analysis methods were used to predict unknown pathogen genes based on homology with identified or functionally clustered genes. In this study, the genes of common pathogens were analyzed to screen and identify genes associated with intracellular survival through sequence similarity, phylogenetic tree analysis and the λ-Red recombination system test method. The total 38,952 protein-coding genes of common pathogens were divided into 19,775 clusters. As demonstrated through a COG analysis, information storage and processing genes might play an important role intracellular survival. Only 19 clusters were present in facultative intracellular pathogens, and not all were present in extracellular pathogens. Construction of a phylogenetic tree selected 18 of these 19 clusters. Comparisons with the DEG database and previous research revealed that seven other clusters are considered essential gene clusters and that seven other clusters are associated with intracellular survival. Moreover, this study confirmed that clusters screened by orthologs with similar function could be replaced with an approved uvrY gene and its orthologs, and the results revealed that the usg gene is associated with intracellular survival. The study improves the current understanding of intracellular pathogens characteristics and allows further exploration of the intracellular survival-related gene modules in these pathogens. Copyright © 2018. Published by Elsevier B.V.

DMPD: Intracellular DNA sensors in immunity. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 18573338 Intracellular DNA sensors in immunity. Takeshita F, Ishii KJ. Curr Opin Im...munol. 2008 Aug;20(4):383-8. Epub 2008 Jun 23. (.png) (.svg) (.html) (.csml) Show Intracellular DNA sensors ...in immunity. PubmedID 18573338 Title Intracellular DNA sensors in immunity. Authors Takeshita F, Ishii KJ. P

Stochastic models of intracellular transport

KAUST Repository

Bressloff, Paul C.; Newby, Jay M.

2013-01-01

mechanisms for intracellular transport: passive diffusion and motor-driven active transport. Diffusive transport can be formulated in terms of the motion of an overdamped Brownian particle. On the other hand, active transport requires chemical energy, usually

Response of knee fibrocartilage to joint destabilization.

Science.gov (United States)

Dyment, N A; Hagiwara, Y; Jiang, X; Huang, J; Adams, D J; Rowe, D W

2015-06-01

A major challenge to understanding osteoarthritis (OA) pathology is identifying the cellular events that precede the onset of cartilage damage. The objective of this study is to determine the effect of joint destabilization on early changes to fibrocartilage in the joint. The anterior cruciate ligament was transected in collagen reporter mice (Col1CFP and ColXRFP). Mineralization labels were given every 2 weeks to measure new mineralized cartilage apposition. Novel fluorescent histology of mineralized tissue was used to characterize the changes in fibrocartilage at 2 and 4 weeks post-injury. Changes in fibrocartilaginous structures of the joint occur as early as 2 weeks after injury and are well developed by 4 weeks. The alterations are seen in multiple entheses and in the medial surface of the femoral and tibial condyles. In the responding entheses, mineral apposition towards the ligament midsubstance results in thickening of the mineralize fibrocartilage. These changes are associated with increases in ColX-RFP, Col1-CFP reporter activity and alkaline phosphatase enzyme activity. Mineral apposition also occurs in the fibrocartilage of the non-articular regions of the medial condyles by 2 weeks and develops into osteophytes by 4 weeks post-injury. An unexpected observation is punctate expression of tartrate resistant acid phosphatase activity in unmineralized fibrochondrocytes adjacent to active appositional mineralization. These observations suggest that fibrocartilage activates prior to degradation of the articular cartilage. Thus clinical and histological imaging of fibrocartilage may be an earlier indicator of disease initiation and may indicate a more appropriate time to start preventative treatment. Copyright © 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Optimization of DIII-D discharges to avoid AE destabilization

Science.gov (United States)

Varela, Jacobo; Spong, Donald; Garcia, Luis; Huang, Juan; Murakami, Masanori

2017-10-01

The aim of the study is to analyze the stability of Alfven Eigenmodes (AE) perturbed by energetic particles (EP) during DIII-D operation. We identify the optimal NBI operational regimes that avoid or minimize the negative effects of AE on the device performance. We use the reduced MHD equations to describe the linear evolution of the poloidal flux and the toroidal component of the vorticity in a full 3D system, coupled with equations of density and parallel velocity moments for the energetic particles, including the effect of the acoustic modes. We add the Landau damping and resonant destabilization effects using a closure relation. We perform parametric studies of the MHD and AE stability, taking into account the experimental profiles of the thermal plasma and EP, also using a range of values of the energetic particles β, density and velocity as well the effect of the toroidal couplings. We reproduce the AE activity observed in high poloidal β discharge at the pedestal and reverse shear discharges. This material based on work is supported both by the U.S. Department of Energy, Office of Science, under Contract DE-AC05-00OR22725 with UT-Battelle, LLC. Research sponsored in part by the Ministerio de Economia y Competitividad of Spain under the project.

Rational design and synthesis of water-compatible molecularly imprinted polymers for selective solid phase extraction of amiodarone.

Science.gov (United States)

Muhammad, Turghun; Cui, Liu; Jide, Wang; Piletska, Elena V; Guerreiro, Antonio R; Piletsky, Sergey A

2012-01-04

Novel water-compatible molecularly imprinted polymers (MIPs) selective for amiodarone (AD) were designed via a new methodology which relies on screening library of non-imprinted polymers (NIPs). The NIP library consisted of eighteen cross-linked co-polymers synthesized from monomers commonly used in molecular imprinting. The binding capacity of each polymer in the library was analyzed in two different solvents. Binding in water was used to assess non-specific (hydrophobic) interactions and binding in an appropriate organic solvent was used to assess specific interactions. A good correlation was found between the screening tests and modeling of monomer-template interactions performed using computational approach. Additionally, analysis of template-monomer interactions was performed using UV-vis spectroscopy. As the result, 4-vinylpyridine (4-VP) was selected as the best monomer for developing MIP for AD. The 4-VP-based polymers demonstrated imprinting factor equal 3.9. The polymers performance in SPE was evaluated using AD and its structural analogues. The recovery of AD was as high as 96% when extracted from spiked phosphate buffer (pH 4.5) solution and 82.1% from spiked serum samples. The developed MIP shown as a material with specific binding to AD, comparing to its structural analogues, 1-(2-diethylaminoethoxy)-2,6-diiodo-4-nitrobenzene and lidocaine, which shown 9.9% and 25.4% of recovery from the buffer solution, correspondingly. We believe that the screening of NIP library could be proposed as an alternative to commonly used computational and combinatorial approaches. Copyright © 2011 Elsevier B.V. All rights reserved.

Intracellular Complement Activation Sustains T Cell Homeostasis and Mediates Effector Differentiation

Science.gov (United States)

Liszewski, M. Kathryn; Kolev, Martin; Le Friec, Gaelle; Leung, Marilyn; Bertram, Paula G.; Fara, Antonella F.; Subias, Marta; Pickering, Matthew C.; Drouet, Christian; Meri, Seppo; Arstila, T. Petteri; Pekkarinen, Pirkka T.; Ma, Margaret; Cope, Andrew; Reinheckel, Thomas; Rodriguez de Cordoba, Santiago; Afzali, Behdad; Atkinson, John P.; Kemper, Claudia

2013-01-01

Summary Complement is viewed as a critical serum-operative component of innate immunity, with processing of its key component, C3, into activation fragments C3a and C3b confined to the extracellular space. We report here that C3 activation also occurred intracellularly. We found that the T cell-expressed protease cathepsin L (CTSL) processed C3 into biologically active C3a and C3b. Resting T cells contained stores of endosomal and lysosomal C3 and CTSL and substantial amounts of CTSL-generated C3a. While “tonic” intracellular C3a generation was required for homeostatic T cell survival, shuttling of this intracellular C3-activation-system to the cell surface upon T cell stimulation induced autocrine proinflammatory cytokine production. Furthermore, T cells from patients with autoimmune arthritis demonstrated hyperactive intracellular complement activation and interferon-γ production and CTSL inhibition corrected this deregulated phenotype. Importantly, intracellular C3a was observed in all examined cell populations, suggesting that intracellular complement activation might be of broad physiological significance. PMID:24315997

Optimizing Nanoelectrode Arrays for Scalable Intracellular Electrophysiology.

Science.gov (United States)

Abbott, Jeffrey; Ye, Tianyang; Ham, Donhee; Park, Hongkun

2018-03-20

Electrode technology for electrophysiology has a long history of innovation, with some decisive steps including the development of the voltage-clamp measurement technique by Hodgkin and Huxley in the 1940s and the invention of the patch clamp electrode by Neher and Sakmann in the 1970s. The high-precision intracellular recording enabled by the patch clamp electrode has since been a gold standard in studying the fundamental cellular processes underlying the electrical activities of neurons and other excitable cells. One logical next step would then be to parallelize these intracellular electrodes, since simultaneous intracellular recording from a large number of cells will benefit the study of complex neuronal networks and will increase the throughput of electrophysiological screening from basic neurobiology laboratories to the pharmaceutical industry. Patch clamp electrodes, however, are not built for parallelization; as for now, only ∼10 patch measurements in parallel are possible. It has long been envisioned that nanoscale electrodes may help meet this challenge. First, nanoscale electrodes were shown to enable intracellular access. Second, because their size scale is within the normal reach of the standard top-down fabrication, the nanoelectrodes can be scaled into a large array for parallelization. Third, such a nanoelectrode array can be monolithically integrated with complementary metal-oxide semiconductor (CMOS) electronics to facilitate the large array operation and the recording of the signals from a massive number of cells. These are some of the central ideas that have motivated the research activity into nanoelectrode electrophysiology, and these past years have seen fruitful developments. This Account aims to synthesize these findings so as to provide a useful reference. Summing up from the recent studies, we will first elucidate the morphology and associated electrical properties of the interface between a nanoelectrode and a cellular membrane

Investigating Internalization and Intracellular Trafficking of GPCRs

DEFF Research Database (Denmark)

Foster, Simon R; Bräuner-Osborne, Hans

2017-01-01

for signal transduction. One of the major mechanisms for GPCR regulation involves their endocytic trafficking, which serves to internalize the receptors from the plasma membrane and thereby attenuate G protein-dependent signaling. However, there is accumulating evidence to suggest that GPCRs can signal...... independently of G proteins, as well as from intracellular compartments including endosomes. It is in this context that receptor internalization and intracellular trafficking have attracted renewed interest within the GPCR field. In this chapter, we will review the current understanding and methodologies...

The destabilizing influence of heat flow on the geological environment during underground nuclear explosions

International Nuclear Information System (INIS)

Politikov, M.I.; Kamberov, I.M.; Krivchenko, V.F.; Lukashenko, S.N.; Solodukhin, V.P.

2001-01-01

The study has determined the fact that the processes of gas-radioactive ectoplasm intrusion from nuclear cavities in the geological environment bring the significant contribution in bosom destabilizing besides the mechanical rock destruction as affected by underground nuclear explosions. Not only heat field forming that reduces the rock resistance and increases its porosity is related to it, but also the forming, on the way, of man-caused contamination aureoles of the geological environment, including the underground water bearing horizon. Unfortunately, this problem is hardly studied, mainly for the lack of reliable apparatus and methods. Judging by the results of information search, the best way to solve the problem is not yet known. (author)

The Influence of Oblique Angle Forced Exercise in Surgically Destabilized Stifle Joints Is Synergistic with Bone, but Antagonistic with Cartilage in an Ovine Model of Osteoarthritis

Directory of Open Access Journals (Sweden)

Rachel J. Hill

2017-01-01

Full Text Available Large animal models of osteoarthritis are a necessary testing ground for FDA approval of human medicine applications. Sheep models have advantages over other available large animals, but development and progression of osteoarthritis in sheep is exceedingly slow, which handicaps progress in development of potential treatments. We combined oblique angle forced exercise to increase stress on the stifle, with surgical destabilization to hasten the development of osteoarthritis in ewes. Methods for early detection of clinical signs included radiography, urine, and serum biomarker assays and gait analysis and ex vivo we used microcomputed tomography and macroscopic joint analysis. Our model was able to produce clinically detectable signs of osteoarthritis in a relatively short period (14 weeks. Changes in bone were highly correlated between microcomputed tomography and radiographic analysis and changes in cartilage correlated well between urinary glycosaminoglycan levels and serum aggrecanase analyses. Exercise improved the negative effects of destabilization in bone but exacerbated the negative effects of destabilization in cartilage. These observations suggest that we may need to consider treatments for bone and cartilage separately. These results represent an improved large animal model of osteoarthritis with rapid onset of disease and superior detection of bone and soft tissue changes.

The epithelial cell cytoskeleton and intracellular trafficking. I. Shiga toxin B-subunit system: retrograde transport, intracellular vectorization, and more.

Science.gov (United States)

Johannes, Ludger

2002-07-01

Many intracellular transport routes are still little explored. This is particularly true for retrograde transport between the plasma membrane and the endoplasmic reticulum. Shiga toxin B subunit has become a powerful tool to study this pathway, and recent advances on the molecular mechanisms of transport in the retrograde route and on its physiological function(s) are summarized. Furthermore, it is discussed how the study of retrograde transport of Shiga toxin B subunit allows one to design new methods for the intracellular delivery of therapeutic compounds.

New perspective in the assessment of total intracellular magnesium

Directory of Open Access Journals (Sweden)

Azzurra Sargenti

2014-01-01

Full Text Available Magnesium (Mg is essential for biological processes, but its cellular homeostasis has not been thoroughly elucidated, mainly because of the inadequacy of the available techniques to map intracellular Mg distribution. Recently, particular interest has been raised by a new family of fluorescent probes, diaza-18-crown-hydroxyquinoline (DCHQ, that shows remarkably high affinity and specificity for Mg, thus permitting the detection of the total intracellular Mg. The data obtained by fluori- metric and cytofluorimetric assays performed with DCHQ5 are in good agreement with atomic absorption spectroscopy, confirming that DCHQ5 probe allows both qualitative and quantitative determination of total intracellular Mg.

PROTEOLYTIC REMOVAL OF THE CARBOXYL TERMINUS OF THE T4 GENE 32 HELIX-DESTABILIZING PROTEIN ALTERS THE T4 IN VITRO REPLICATION COMPLEX

Energy Technology Data Exchange (ETDEWEB)

Burke, R.L.; Alberts, B.M.; Hosoda, J.

1980-07-01

The proteolytic removal of about 60 amino acids from the COOH terminus of the bacteriophage T4 helix-destabilizing protein (gene 32 protein) produces 32*I, a 27,000-dalton fragment which still binds tightly and cooperatively to single-stranded DNA. The substitution of 32*I protein for intact 32 protein in the seven-protein T4 replication complex results in dramatic changes in some of the reactions catalyzed by this in vitro DNA replication system, while leaving others largely unperturbed. (1) Like intact 32 protein, the 32*I protein promotes DNA synthesis by the DNA polymerase when the T4 polymerase accessory proteins (gene 44/62 and 45 proteins) are also present. The host helix-destabilizing protein (Escherichia coli ssb protein) cannot replace the 32*I protein for this synthesis. (2) Unlike intact 32 protein, 32*I protein strongly inhibits DNA synthesis catalyzed by the T4 DNA polymerase alone on a primed single-stranded DNA template. (3) Unlike intact 32 protein, the 32*I protein strongly inhibits RNA primer synthesis catalyzed by the T4 gene 41 and 61 proteins and also reduces the efficiency of RNA primer utilization. As a result, de novo DNA chain starts are blocked completely in the complete T4 replication system, and no lagging strand DNA synthesis occurs. (4) The 32*I protein does not bind to either the T4 DNA polymerase or to the T4 gene 61 protein in the absence of DNA; these associations (detected with intact 32 protein) would therefore appear to be essential for the normal control of 32 protein activity, and to account at least in part for observations 2 and 3, above. We propose that the COOH-terminal domain of intact 32 protein functions to guide its interactions with the T4 DNA polymerase and the T4 gene 61 RNA-priming protein. When this domain is removed, as in 32*I protein, the helix destabilization induced by the protein is controlled inadequately, so that polymerizing enzymes tend to be displaced from the growing 3{prime}-OH end of a

Destabilization of Human Insulin Fibrils by Peptides of Fruit Bromelain Derived From Ananas comosus (Pineapple).

Science.gov (United States)

Das, Sromona; Bhattacharyya, Debasish

2017-12-01

Deposition of insulin aggregates in human body leads to dysfunctioning of several organs. Effectiveness of fruit bromelain from pineapple in prevention of insulin aggregate was investigated. Proteolyses of bromelain was done as par human digestive system and the pool of small peptides was separated from larger peptides and proteins. Under conditions of growth of insulin aggregates from its monomers, this pool of peptides restricted the reaction upto formation of oligomers of limited size. These peptides also destabilized preformed insulin aggregates to oligomers. These processes were followed fluorimetrically using Thioflavin T and 1-ANS, size-exclusion HPLC, dynamic light scattering, atomic force microscopy, and transmission electron microscopy. Sequences of insulin (A and B chains) and bromelain were aligned using Clustal W software to predict most probable sites of interactions. Synthetic tripeptides corresponding to the hydrophobic interactive sites of bromelain showed disaggregation of insulin suggesting specificity of interactions. The peptides GG and AAA serving as negative controls showed no potency in destabilization of aggregates. Disaggregation potency of the peptides was also observed when insulin was deposited on HepG2 liver cells where no formation of toxic oligomers occurred. Amyloidogenic des-octapeptide (B23-B30 of insulin) incapable of cell signaling showed cytotoxicity similar to insulin. This toxicity could be neutralized by bromelain derived peptides. FT-IR and far-UV circular dichroism analysis indicated that disaggregated insulin had structure distinctly different from that of its hexameric (native) or monomeric states. Based on the stoichiometry of interaction and irreversibility of disaggregation, the mechanism/s of the peptides and insulin interactions has been proposed. J. Cell. Biochem. 118: 4881-4896, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Destabilizing protein polymorphisms in the genetic background direct phenotypic expression of mutant SOD1 toxicity.

Directory of Open Access Journals (Sweden)

Tali Gidalevitz

2009-03-01

Full Text Available Genetic background exerts a strong modulatory effect on the toxicity of aggregation-prone proteins in conformational diseases. In addition to influencing the misfolding and aggregation behavior of the mutant proteins, polymorphisms in putative modifier genes may affect the molecular processes leading to the disease phenotype. Mutations in SOD1 in a subset of familial amyotrophic lateral sclerosis (ALS cases confer dominant but clinically variable toxicity, thought to be mediated by misfolding and aggregation of mutant SOD1 protein. While the mechanism of toxicity remains unknown, both the nature of the SOD1 mutation and the genetic background in which it is expressed appear important. To address this, we established a Caenorhabditis elegans model to systematically examine the aggregation behavior and genetic interactions of mutant forms of SOD1. Expression of three structurally distinct SOD1 mutants in C. elegans muscle cells resulted in the appearance of heterogeneous populations of aggregates and was associated with only mild cellular dysfunction. However, introduction of destabilizing temperature-sensitive mutations into the genetic background strongly enhanced the toxicity of SOD1 mutants, resulting in exposure of several deleterious phenotypes at permissive conditions in a manner dependent on the specific SOD1 mutation. The nature of the observed phenotype was dependent on the temperature-sensitive mutation present, while its penetrance reflected the specific combination of temperature-sensitive and SOD1 mutations. Thus, the specific toxic phenotypes of conformational disease may not be simply due to misfolding/aggregation toxicity of the causative mutant proteins, but may be defined by their genetic interactions with cellular pathways harboring mildly destabilizing missense alleles.

Advances in genetic manipulation of obligate intracellular bacterial pathogens

Directory of Open Access Journals (Sweden)

Paul eBeare

2011-05-01

Full Text Available Infections by obligate intracellular bacterial pathogens result in significant morbidity and mortality worldwide. These bacteria include Chlamydia spp., which causes millions of cases of sexually transmitted disease and blinding trachoma annually, and members of the α-proteobacterial genera Anaplasma, Ehrlichia, Orientia and Rickettsia, agents of serious human illnesses including epidemic typhus. Coxiella burnetii, the agent of human Q fever, has also been considered a prototypical obligate intracellular bacterium, but recent host cell-free (axenic growth has rescued it from obligatism. The historic genetic intractability of obligate intracellular bacteria has severely limited molecular dissection of their unique lifestyles and virulence factors involved in pathogenesis. Host cell restricted growth is a significant barrier to genetic transformation that can make simple procedures for free-living bacteria, such as cloning, exceedingly difficult. Low transformation efficiency requiring long term culture in host cells to expand small transformant populations is another obstacle. Despite numerous technical limitations, the last decade has witnessed significant gains in genetic manipulation of obligate intracellular bacteria including allelic exchange. Continued development of genetic tools should soon enable routine mutation and complementation strategies for virulence factor discovery and stimulate renewed interest in these refractory pathogens. In this review, we discuss the technical challenges associated with genetic transformation of obligate intracellular bacteria and highlight advances made with individual genera.

Intracellular transport of fat-soluble vitamins A and E.

Science.gov (United States)

Kono, Nozomu; Arai, Hiroyuki

2015-01-01

Vitamins are compounds that are essential for the normal growth, reproduction and functioning of the human body. Of the 13 known vitamins, vitamins A, D, E and K are lipophilic compounds and are therefore called fat-soluble vitamins. Because of their lipophilicity, fat-soluble vitamins are solubilized and transported by intracellular carrier proteins to exert their actions and to be metabolized properly. Vitamin A and its derivatives, collectively called retinoids, are solubilized by intracellular retinoid-binding proteins such as cellular retinol-binding protein (CRBP), cellular retinoic acid-binding protein (CRABP) and cellular retinal-binding protein (CRALBP). These proteins act as chaperones that regulate the metabolism, signaling and transport of retinoids. CRALBP-mediated intracellular retinoid transport is essential for vision in human. α-Tocopherol, the main form of vitamin E found in the body, is transported by α-tocopherol transfer protein (α-TTP) in hepatic cells. Defects of α-TTP cause vitamin E deficiency and neurological disorders in humans. Recently, it has been shown that the interaction of α-TTP with phosphoinositides plays a critical role in the intracellular transport of α-tocopherol and is associated with familial vitamin E deficiency. In this review, we summarize the mechanisms and biological significance of the intracellular transport of vitamins A and E. © 2014 The Authors. Traffic published by John Wiley & Sons Ltd.

Ciprofloxacin nano-niosomes for targeting intracellular infections: an in vitro evaluation

International Nuclear Information System (INIS)

Akbari, Vajihe; Abedi, Daryoush; Pardakhty, Abbas; Sadeghi-Aliabadi, Hojjat

2013-01-01

In order to propose non-ionic surfactant vesicles (niosomes) for the treatment of intracellular infections, a remote loading method (active drug encapsulation) followed by sonication was used to prepare nano-niosome formulations containing ciprofloxacin (CPFX). Size analysis, size distribution and zeta potentials of niosomes were evaluated and then their antimicrobial activity, cellular uptake, cytotoxicity, intracellular distribution, and antibacterial activity against intracellular Staphylococcus aureus infection of murine macrophage-like, J774, cells were investigated in comparison to free drug. Our findings reveal that size and composition of the niosome formula can influence their in vitro biological properties. Vesicles in the 300–600 nm size range were phagocytosed to a greater degree by macrophages in comparison to other size vesicles. The minimum inhibitory concentrations (MICs) of CPFX-loaded niosomes were two to eightfold lower than MICs of free CPFX. In addition, niosome encapsulation of CPFX provided high intracellular antimicrobial activities while free CPFX is ineffective for eradicating intracellular forms of S. aureus. Encapsulation of CPFX in niosomes generally decreased its in vitro cytotoxicity. Our results show that niosomes are suitable drug delivery systems with good efficacy and safety properties to be proposed for drug targeting against intracellular infections.

Ciprofloxacin nano-niosomes for targeting intracellular infections: an in vitro evaluation

Energy Technology Data Exchange (ETDEWEB)

Akbari, Vajihe; Abedi, Daryoush [Isfahan University of Medical Sciences, Department of Pharmaceutical Biotechnology and Isfahan Pharmaceutical Research Center, Faculty of Pharmacy (Iran, Islamic Republic of); Pardakhty, Abbas [Kerman University of Medical Sciences, Pharmaceutics Research Center (Iran, Islamic Republic of); Sadeghi-Aliabadi, Hojjat, E-mail: sadeghi@pharm.mui.ac.ir [Isfahan University of Medical Sciences, Department of Pharmaceutical Biotechnology and Isfahan Pharmaceutical Research Center, Faculty of Pharmacy (Iran, Islamic Republic of)

2013-04-15

In order to propose non-ionic surfactant vesicles (niosomes) for the treatment of intracellular infections, a remote loading method (active drug encapsulation) followed by sonication was used to prepare nano-niosome formulations containing ciprofloxacin (CPFX). Size analysis, size distribution and zeta potentials of niosomes were evaluated and then their antimicrobial activity, cellular uptake, cytotoxicity, intracellular distribution, and antibacterial activity against intracellular Staphylococcus aureus infection of murine macrophage-like, J774, cells were investigated in comparison to free drug. Our findings reveal that size and composition of the niosome formula can influence their in vitro biological properties. Vesicles in the 300-600 nm size range were phagocytosed to a greater degree by macrophages in comparison to other size vesicles. The minimum inhibitory concentrations (MICs) of CPFX-loaded niosomes were two to eightfold lower than MICs of free CPFX. In addition, niosome encapsulation of CPFX provided high intracellular antimicrobial activities while free CPFX is ineffective for eradicating intracellular forms of S. aureus. Encapsulation of CPFX in niosomes generally decreased its in vitro cytotoxicity. Our results show that niosomes are suitable drug delivery systems with good efficacy and safety properties to be proposed for drug targeting against intracellular infections.

A bacteriophage endolysin that eliminates intracellular streptococci

Science.gov (United States)

Shen, Yang; Barros, Marilia; Vennemann, Tarek; Gallagher, D Travis; Yin, Yizhou; Linden, Sara B; Heselpoth, Ryan D; Spencer, Dennis J; Donovan, David M; Moult, John; Fischetti, Vincent A; Heinrich, Frank; Lösche, Mathias; Nelson, Daniel C

2016-01-01

PlyC, a bacteriophage-encoded endolysin, lyses Streptococcus pyogenes (Spy) on contact. Here, we demonstrate that PlyC is a potent agent for controlling intracellular Spy that often underlies refractory infections. We show that the PlyC holoenzyme, mediated by its PlyCB subunit, crosses epithelial cell membranes and clears intracellular Spy in a dose-dependent manner. Quantitative studies using model membranes establish that PlyCB interacts strongly with phosphatidylserine (PS), whereas its interaction with other lipids is weak, suggesting specificity for PS as its cellular receptor. Neutron reflection further substantiates that PlyC penetrates bilayers above a PS threshold concentration. Crystallography and docking studies identify key residues that mediate PlyCB–PS interactions, which are validated by site-directed mutagenesis. This is the first report that a native endolysin can traverse epithelial membranes, thus substantiating the potential of PlyC as an antimicrobial for Spy in the extracellular and intracellular milieu and as a scaffold for engineering other functionalities. DOI: http://dx.doi.org/10.7554/eLife.13152.001 PMID:26978792

Alpha particle destabilization of the TAE modes

International Nuclear Information System (INIS)

Cheng, C.Z.

1991-01-01

The high frequency, low mode number toroidicity-induced Alfven eigenmodes (TAE) are shown to be driven unstable by the circulating and/or trapped α-particles through the wave-particle resonances. For a poloidal harmonic to satisfy the resonance condition it requires that the α-particle birth speed v α ≥ v A /(2|m-nq|), where v A is the Alfven speed, m is the poloidal mode number, and n is the toroidal mode number. To destabilize the TAE modes, the inverse Landau damping associated with the α-particle pressure gradient free energy must overcome the velocity space Landau damping due to both the slowing-down α-particle and the core Maxwellian electron and ion distributions. Stability criteria in terms of the α-particle beta β α , α-particle pressure gradient parameter (ω * /ω A ) (ω * is the α-particle diamagnetic drift frequency), and (v α /v A ) parameters are presented for TFTR, CIT, and ITER tokamaks. The volume averaged α-particle beta threshold for TAE instability also depends sensitively on the core electron and ion temperature. Typically the volume averaged α-particle beta threshold is in the order of 10 -4 if the continuum damping effect is absent. Typical growth rates of the n = 1 TAE mode can be in the order of 10 -2 ω A , where ω A = v A /qR. Stability of higher n TAE modes is also studied. Other types of global Alfven waves are stable due to sideband mode continuum damping resulting from toroidal coupling effects. If the Alfven continuum gap does not exist across the whole minor radius, continuum damping exists for some poloidal harmonics. The continuum damping effect is studied by employing both a resistive MHD stability code (NOVA-R) and an analytical matching method, and the results are presented. 1 ref

Assessment of destabilizing factor for automatic control systems in propulsion systems of mechatronic and maritime transport objects

Science.gov (United States)

Zhilenkov, A. A.; Kapitonov, A. A.

2017-10-01

It is known that many of today’s ships and vessels have a shaft generator as a part of their power plants. Modern automatic control systems used in the world’s fleet do not enable their shaft generators to operate in parallel with the main diesel generators for long-term sustenance of the total load of the ship network. On the other hand, according to our calculations and experiments, a shaft generator operated in parallel with the main power plant helps save at least 10% of fuel while making the power system of the ship more efficient, reliable, and eco-friendly. The fouling and corrosion of the propeller as well as the weather conditions of navigation affect its modulus of resistance. It changes the free component of the transient process of shaft generator stress frequency changes in transient processes. While the shaft generator and the diesel generator of the ship power plant are paralleled, there emerges an angle between their EMF. This results in equalizing currents generated between them. The altering torque in the drive-shaft line—propeller system causes torsional fluctuations of the ship shaft line. To compensate for the effect of destabilizing factors and torsional fluctuations of the shaft line on the dynamic characteristics of the transient process that alters the RPM of the main engine, sliding mode controls can be used. To synthesize such a control, one has to evaluate the effect of destabilizing factors.

Genomic analysis suggests that mRNA destabilization by the microprocessor is specialized for the auto-regulation of Dgcr8.

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Archana Shenoy

2009-09-01

Full Text Available The Microprocessor, containing the RNA binding protein Dgcr8 and RNase III enzyme Drosha, is responsible for processing primary microRNAs to precursor microRNAs. The Microprocessor regulates its own levels by cleaving hairpins in the 5'UTR and coding region of the Dgcr8 mRNA, thereby destabilizing the mature transcript.To determine whether the Microprocessor has a broader role in directly regulating other coding mRNA levels, we integrated results from expression profiling and ultra high-throughput deep sequencing of small RNAs. Expression analysis of mRNAs in wild-type, Dgcr8 knockout, and Dicer knockout mouse embryonic stem (ES cells uncovered mRNAs that were specifically upregulated in the Dgcr8 null background. A number of these transcripts had evolutionarily conserved predicted hairpin targets for the Microprocessor. However, analysis of deep sequencing data of 18 to 200nt small RNAs in mouse ES, HeLa, and HepG2 indicates that exonic sequence reads that map in a pattern consistent with Microprocessor activity are unique to Dgcr8.We conclude that the Microprocessor's role in directly destabilizing coding mRNAs is likely specifically targeted to Dgcr8 itself, suggesting a specialized cellular mechanism for gene auto-regulation.

Destabilization of strigolactone receptor DWARF14 by binding of ligand and E3-ligase signaling effector DWARF3

Science.gov (United States)

Zhao, Li-Hua; Zhou, X Edward; Yi, Wei; Wu, Zhongshan; Liu, Yue; Kang, Yanyong; Hou, Li; de Waal, Parker W; Li, Suling; Jiang, Yi; Scaffidi, Adrian; Flematti, Gavin R; Smith, Steven M; Lam, Vinh Q; Griffin, Patrick R; Wang, Yonghong; Li, Jiayang; Melcher, Karsten; Xu, H Eric

2015-01-01

Strigolactones (SLs) are endogenous hormones and exuded signaling molecules in plant responses to low levels of mineral nutrients. Key mediators of the SL signaling pathway in rice include the α/β-fold hydrolase DWARF 14 (D14) and the F-box component DWARF 3 (D3) of the ubiquitin ligase SCFD3 that mediate ligand-dependent degradation of downstream signaling repressors. One perplexing feature is that D14 not only functions as the SL receptor but is also an active enzyme that slowly hydrolyzes diverse natural and synthetic SLs including GR24, preventing the crystallization of a binary complex of D14 with an intact SL as well as the ternary D14/SL/D3 complex. Here we overcome these barriers to derive a structural model of D14 bound to intact GR24 and identify the interface that is required for GR24-mediated D14-D3 interaction. The mode of GR24-mediated signaling, including ligand recognition, hydrolysis by D14, and ligand-mediated D14-D3 interaction, is conserved in structurally diverse SLs. More importantly, D14 is destabilized upon the binding of ligands and D3, thus revealing an unusual mechanism of SL recognition and signaling, in which the hormone, the receptor, and the downstream effectors are systematically destabilized during the signal transduction process. PMID:26470846

A cellular model of memory reconsolidation involves reactivation-induced destabilization and restabilization at the sensorimotor synapse in Aplysia.

Science.gov (United States)

Lee, Sue-Hyun; Kwak, Chuljung; Shim, Jaehoon; Kim, Jung-Eun; Choi, Sun-Lim; Kim, Hyoung F; Jang, Deok-Jin; Lee, Jin-A; Lee, Kyungmin; Lee, Chi-Hoon; Lee, Young-Don; Miniaci, Maria Concetta; Bailey, Craig H; Kandel, Eric R; Kaang, Bong-Kiun

2012-08-28

The memory reconsolidation hypothesis suggests that a memory trace becomes labile after retrieval and needs to be reconsolidated before it can be stabilized. However, it is unclear from earlier studies whether the same synapses involved in encoding the memory trace are those that are destabilized and restabilized after the synaptic reactivation that accompanies memory retrieval, or whether new and different synapses are recruited. To address this issue, we studied a simple nonassociative form of memory, long-term sensitization of the gill- and siphon-withdrawal reflex in Aplysia, and its cellular analog, long-term facilitation at the sensory-to-motor neuron synapse. We found that after memory retrieval, behavioral long-term sensitization in Aplysia becomes labile via ubiquitin/proteasome-dependent protein degradation and is reconsolidated by means of de novo protein synthesis. In parallel, we found that on the cellular level, long-term facilitation at the sensory-to-motor neuron synapse that mediates long-term sensitization is also destabilized by protein degradation and is restabilized by protein synthesis after synaptic reactivation, a procedure that parallels memory retrieval or retraining evident on the behavioral level. These results provide direct evidence that the same synapses that store the long-term memory trace encoded by changes in the strength of synaptic connections critical for sensitization are disrupted and reconstructed after signal retrieval.

Determination of Intracellular Vitrification Temperatures for Unicellular Micro Organisms under Conditions Relevant for Cryopreservation.

Science.gov (United States)

Fonseca, Fernanda; Meneghel, Julie; Cenard, Stéphanie; Passot, Stéphanie; Morris, G John

2016-01-01

During cryopreservation ice nucleation and crystal growth may occur within cells or the intracellular compartment may vitrify. Whilst previous literature describes intracellular vitrification in a qualitative manner, here we measure the intracellular vitrification temperature of bacteria and yeasts under conditions relevant to cryopreservation, including the addition of high levels of permeating and nonpermeating additives and the application of rapid rates of cooling. The effects of growth conditions that are known to modify cellular freezing resistance on the intracellular vitrification temperature are also examined. For bacteria a plot of the activity on thawing against intracellular glass transition of the maximally freeze-concentrated matrix (Tg') shows that cells with the lowest value of intracellular Tg' survive the freezing process better than cells with a higher intracellular Tg'. This paper demonstrates the role of the physical state of the intracellular environment in determining the response of microbial cells to preservation and could be a powerful tool to be manipulated to allow the optimization of methods for the preservation of microorganisms.

Antiproliferative efficacy of curcumin mimics through microtubule destabilization.

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Khwaja, Sadiya; Fatima, Kaneez; Hasanain, Mohammad; Behera, Chittaranjan; Kour, Avneet; Singh, Arjun; Luqman, Suaib; Sarkar, Jayanta; Chanda, Debabrata; Shanker, Karuna; Gupta, A K; Mondhe, D M; Negi, Arvind S

2018-05-10

Curcumin possesses an attractive chemical structure with highly conjugated diferuloylmethane core. Curcumin mimics have been designed and prepared with an additional bridged phenyl ring in conjugation. Fourteen diverse analogues were evaluated against a panel of human cancer cell lines. The best analogue of the series i.e. compound 6a exhibited potent cytotoxicity against A431, epidermoid carcinoma cell line (IC 50  = 1.5 μM) and DLD1, colorectal adenocarcinoma cell line (IC 50  = 6.9 μM). In tubulin kinetics experiment, compound 6a destabilized polymerisation process (IC 50  = 4.68 μM). In cell cycle analysis, compound 6a exerted G2/M phase arrest in A431 cells and induced apoptosis. In Ehrlich Ascites Carcinoma in Swiss-albino mice, compound 6a showed 78.6% tumour reduction at 80 mg/kg dose and 57% solid tumour reduction at 150 mg/kg dose. Further, in acute-oral toxicity experiment in rodent model, compound 6a was given in three different oral doses to Swiss albino mice. There were non-significant changes in various biochemical parameters and major body organs studied, including their absolute and relative weights. It was tolerable up to 300 mg/kg dose in Swiss-albino mice. The present study shows that the novel curcumin mimic 6a is a safe and efficacious anticancer compound. However, it needs to be optimized for better efficacy. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Positioning the Intracellular Salt Potassium Glutamate in the Hofmeister Series by Chemical Unfolding Studies of NTL9.

Science.gov (United States)

Sengupta, Rituparna; Pantel, Adrian; Cheng, Xian; Shkel, Irina; Peran, Ivan; Stenzoski, Natalie; Raleigh, Daniel P; Record, M Thomas

2016-04-19

In vitro, replacing KCl with potassium glutamate (KGlu), the Escherichia coli cytoplasmic salt and osmolyte, stabilizes folded proteins and protein-nucleic acid complexes. To understand the chemical basis for these effects and rank Glu- in the Hofmeister anion series for protein unfolding, we quantify and interpret the strong stabilizing effect of KGlu on the ribosomal protein domain NTL9, relative to the effects of other stabilizers (KCl, KF, and K2SO4) and destabilizers (GuHCl and GuHSCN). GuHSCN titrations at 20 ° C, performed as a function of the concentration of KGlu or another salt and monitored by NTL9 fluorescence, are analyzed to obtain R-values quantifying the Hofmeister salt concentration (m3) dependence of the unfolding equilibrium constant K(obs) [r-value = −d ln K(obs)/dm3 = (1/RT) dΔG(obs) ° /dm3 = m-value/RT]. r-Values for both stabilizing K+ salts and destabilizing GuH+ salts are compared with predictions from model compound data. For two-salt mixtures, we find that contributions of stabilizing and destabilizing salts to observed r-values are additive and independent. At 20 ° C, we determine a KGlu r-value of 3.22 m(−1) and K2SO4, KF, KCl, GuHCl, and GuHSCN r-values of 5.38, 1.05, 0.64, −1.38, and −3.00 m(−1), respectively. The KGlu r-value represents a 25-fold (1.9 kcal) stabilization per molal KGlu added. KGlu is much more stabilizing than KF, and the stabilizing effect of KGlu is larger in magnitude than the destabilizing effect of GuHSCN. Interpretation of the data reveals good agreement between predicted and observed relative r-values and indicates the presence of significant residual structure in GuHSCN-unfolded NTL9 at 20 ° C.

Phosphorylation of ARD1 by IKKβ contributes to its destabilization and degradation

International Nuclear Information System (INIS)

Kuo, Hsu-Ping; Lee, Dung-Fang; Xia, Weiya; Lai, Chien-Chen; Li, Long-Yuan; Hung, Mien-Chie

2009-01-01

IκB kinase β (IKKβ), a major kinase downstream of various proinflammatory signals, mediates multiple cellular functions through phosphorylation and regulation of its substrates. On the basis of protein sequence analysis, we identified arrest-defective protein 1 (ARD1), a protein involved in apoptosis and cell proliferation processes in many human cancer cells, as a new IKKβ substrate. We provided evidence showing that ARD1 is indeed a bona fide substrate of IKKβ. IKKβ physically associated with ARD1 and phosphorylated it at Ser209. Phosphorylation by IKKβ destabilized ARD1 and induced its proteasome-mediated degradation. Impaired growth suppression was observed in ARD1 phosphorylation-mimic mutant (S209E)-transfected cells as compared with ARD1 non-phosphorylatable mutant (S209A)-transfected cells. Our findings of molecular interactions between ARD1 and IKKβ may enable further understanding of the upstream regulation mechanisms of ARD1 and of the diverse functions of IKKβ.

SIRT3 opposes reprogramming of cancer cell metabolism through HIF1α destabilization

Science.gov (United States)

Finley, Lydia W.S.; Carracedo, Arkaitz; Lee, Jaewon; Souza, Amanda; Egia, Ainara; Zhang, Jiangwen; Teruya-Feldstein, Julie; Moreira, Paula I.; Cardoso, Sandra M.; Clish, Clary B.; Pandolfi, Pier Paolo; Haigis, Marcia C.

2011-01-01

Summary Tumor cells exhibit aberrant metabolism characterized by high glycolysis even in the presence of oxygen. This metabolic reprogramming, known as the Warburg effect, provides tumor cells with the substrates required for biomass generation. Here, we show that the mitochondrial NAD-dependent deacetylase SIRT3 is a crucial regulator of the Warburg effect. Mechanistically, SIRT3 mediates metabolic reprogramming by destabilizing hypoxia-inducible factor-1α (HIF1α), a transcription factor that controls glycolytic gene expression. SIRT3 loss increases reactive oxygen species production, leading to HIF1α stabilization. SIRT3 expression is reduced in human breast cancers, and its loss correlates with the upregulation of HIF1α target genes. Finally, we find that SIRT3 overexpression represses glycolysis and proliferation in breast cancer cells, providing a metabolic mechanism for tumor suppression. PMID:21397863

The mass media destabilizes the cultural homogenous regime in Axelrod's model

Energy Technology Data Exchange (ETDEWEB)

Peres, Lucas R; Fontanari, Jose F [Instituto de Fisica de Sao Carlos, Universidade de Sao Paulo, Caixa Postal 369, 13560-970 Sao Carlos SP (Brazil)

2010-02-05

An important feature of Axelrod's model for culture dissemination or social influence is the emergence of many multicultural absorbing states, despite the fact that the local rules that specify the agents interactions are explicitly designed to decrease the cultural differences between agents. Here we re-examine the problem of introducing an external, global interaction-the mass media-in the rules of Axelrod's model: in addition to their nearest neighbors, each agent has a certain probability p to interact with a virtual neighbor whose cultural features are fixed from the outset. Most surprisingly, this apparently homogenizing effect actually increases the cultural diversity of the population. We show that, contrary to previous claims in the literature, even a vanishingly small value of p is sufficient to destabilize the homogeneous regime for very large lattice sizes.

The interferon response to intracellular DNA: why so many receptors?

Science.gov (United States)

Unterholzner, Leonie

2013-11-01

The detection of intracellular DNA has emerged to be a key event in the innate immune response to viruses and intracellular bacteria, and during conditions of sterile inflammation and autoimmunity. One of the consequences of the detection of DNA as a 'stranger' and a 'danger' signal is the production of type I interferons and pro-inflammatory cytokines. Much work has been dedicated to the elucidation of the signalling cascades that activate this DNA-induced gene expression programme. However, while many proteins have been proposed to act as sensors for intracellular DNA in recent years, none has been met with universal acceptance, and a theory linking all the recent observations is, as yet, lacking. This review presents the evidence for the various interferon-inducing DNA receptors proposed to date, and examines the hypotheses that might explain why so many different receptors appear to be involved in the innate immune recognition of intracellular DNA. Copyright © 2013 Elsevier GmbH. All rights reserved.

The protective effect of grape seed and Ginkgo biloba against hepatotoxicity induced by the antidysrhythmic drug “amiodarone” in male albino rats

Directory of Open Access Journals (Sweden)

Manal Abdul-Hamid

2018-06-01

Full Text Available Amiodarone was an orally effective antiarrhythmic drug widely used throughout the world, had long-term administration side effects such as hepatotoxicity. The actions of two antioxidants; grape seed and Ginkgo biloba on the extent of tissue damage in amiodarone-induced hepatotoxicity were elucidated in this study. We equally divided thirty-six albino rats into six groups given doses by gastric tube daily for 8 weeks as follow; the 1st group (G1 served as an untreated control group under the same laboratory conditions and was given distilled water, the 2nd group (G2 grape seed-treated group that received (100 mg/kg/day, the 3rd group (G3 Ginkgo biloba-treated group that received (100 mg/kg/day, the 4th group (G4 amiodarone-treated group that received (40 mg/kg/day, the 5th group (G5 received amiodarone parallel with grape seed at the same time and the 6th group (G6 received amiodarone parallel with Ginkgo biloba at the same time. The current histological study revealed that amiodarone caused marked change in the liver including degeneration, proliferation of bile duct, inflammatory cells infiltration and fatty changes of hepatocytes in addition to deposition of collagen fibers in the hepatic tissue moreover, ultra-structural observations in the liver including vacuolation, fibrosis and pyknotic nuclei. In addition, histochemical study revealed depletion of glycogen and comet assay revealed marked of DNA damage.Treatment with the two used antioxidants reduced the extent of liver damage induced by amiodarone as indicated by decreased Aspartate aminotransferase (AST and Alanine aminotransferase (ALT activities. These antioxidants ameliorated the histopathological, histochemical and ultrastructure alternations of the liver tissue. In conclusion, grape seed was markedly effective than Ginkgo biloba in protecting rats against amiodarone. Keywords: Amiodarone, Grape seed, Ginkgo biloba, Comet assay, Hepatotoxicity, Histopathology, Ultrastructure

The Effect of Size and Species on Lens Intracellular Hydrostatic Pressure

Science.gov (United States)

Gao, Junyuan; Sun, Xiurong; Moore, Leon C.; Brink, Peter R.; White, Thomas W.; Mathias, Richard T.

2013-01-01

Purpose. Previous experiments showed that mouse lenses have an intracellular hydrostatic pressure that varied from 335 mm Hg in central fibers to 0 mm Hg in surface cells. Model calculations predicted that in larger lenses, all else equal, pressure should increase as the lens radius squared. To test this prediction, lenses of different radii from different species were studied. Methods. All studies were done in intact lenses. Intracellular hydrostatic pressures were measured with a microelectrode-manometer–based system. Membrane conductances were measured by frequency domain impedance analysis. Intracellular Na+ concentrations were measured by injecting the Na+-sensitive dye sodium-binding benzofuran isophthalate. Results. Intracellular hydrostatic pressures were measured in lenses from mice, rats, rabbits, and dogs with radii (cm) 0.11, 0.22, 0.49, and 0.57, respectively. In each species, pressure varied from 335 ± 6 mm Hg in central fiber cells to 0 mm Hg in surface cells. Further characterization of transport in lenses from mice and rats showed that the density of fiber cell gap junction channels was approximately the same, intracellular Na+ concentrations varied from 17 mM in central fiber cells to 7 mM in surface cells, and intracellular voltages varied from −45 mV in central fiber cells to −60 mV in surface cells. Fiber cell membrane conductance was a factor of 2.7 times larger in mouse than in rat lenses. Conclusions. Intracellular hydrostatic pressure is an important physiological parameter that is regulated in lenses from these different species. The most likely mechanism of regulation is to reduce the density of open Na+-leak channels in fiber cells of larger lenses. PMID:23211824

Surveillance for Intracellular Antibody by Cytosolic Fc Receptor TRIM21

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William A. McEwan

2016-11-01

Full Text Available TRIM21 has emerged as an atypical Fc receptor that is broadly conserved and widely expressed in the cytoplasm of mammalian cells. Viruses that traffic surface-bound antibodies into the cell during infection recruit TRIM21 via a high affinity interaction between Fc and TRIM21 PRYSPRY domain. Following binding of intracellular antibody, TRIM21 acts as both antiviral effector and sensor for innate immune signalling. These activities serve to reduce viral replication by orders of magnitude in vitro and contribute to host survival during in vivo infection. Neutralization occurs rapidly after detection and requires the activity of the ubiquitin-proteasome system. The microbial targets of this arm of intracellular immunity are still being identified: TRIM21 activity has been reported following infection by several non-enveloped viruses and intracellular bacteria. These findings extend the sphere of influence of antibodies to the intracellular domain and have broad implications for immunity. TRIM21 has been implicated in the chronic auto-immune condition systemic lupus erythematosus and is itself an auto-antigen in Sjögren’s syndrome. This review summarises our current understanding of TRIM21’s role as a cytosolic Fc receptor and briefly discusses pathological circumstances where intracellular antibodies have been described, or are hypothesized to occur, and may benefit from further investigations of the role of TRIM21.

Legionella pneumophila transcriptome during intracellular multiplication in human macrophages

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Sebastien P Faucher

2011-04-01

Full Text Available Legionella pneumophila is the causative agent of Legionnaires’ disease, an acute pulmonary infection. L. pneumophila is able to infect and multiply in both phagocytic protozoa, such as Acanthamoeba castellanii, and mammalian professional phagocytes. The best-known L. pneumophila virulence determinant is the Icm/Dot Type IVB secretion system (TFBSS, which is used to translocate more than 150 effector proteins to host cells. While the transcriptional response of Legionella to the intracellular environment of A. castellanii has been investigated, much less is known about the Legionella transcriptional response inside human macrophages. In this study, the transcriptome of L. pneumophila was monitored during exponential and post-exponential phase in rich AYE broth as well as during infection of human cultured macrophages. This was accomplished with microarrays and an RNA amplification procedure called SCOTS to detect small amounts of mRNA from low numbers of intracellular bacteria. Among the genes induced intracellularly are those involved in amino acid biosynthetic pathways leading to L-arginine, L-histidine and L-proline as well as many transport systems involved in amino acid and iron uptake. Gene involved in catabolism of glycerol is also induced during intracellular growth and could be used as a carbon source. The genes encoding the Icm/Dot system are not differentially expressed inside cells compared to control bacteria grown in rich broth, but the genes encoding several translocated effectors are strongly induced. Moreover, we used the transcriptome data to predict previously unrecognized Icm/Dot effector genes based on their expression pattern and confirmed translocation for three candidates. This study provides a comprehensive view of how L. pneumophila responds to the human macrophage intracellular environment.

Long-term pretreatment with desethylamiodarone (DEA) or amiodarone (AMIO) protects against coronary artery occlusion induced ventricular arrhythmias in conscious rats.

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Morvay, Nikolett; Baczkó, István; Sztojkov-Ivanov, Anita; Falkay, György; Papp, Julius Gy; Varró, András; Leprán, István

2015-09-01

The aim of this investigation was to compare the effectiveness of long-term pretreatment with amiodarone (AMIO) and its active metabolite desethylamiodarone (DEA) on arrhythmias induced by acute myocardial infarction in rats. Acute myocardial infarction was induced in conscious, male, Sprague-Dawley rats by pulling a previously inserted loose silk loop around the left main coronary artery. Long-term oral pretreatment with AMIO (30 or 100 mg·(kg body mass)(-1)·day(-1), loading dose 100 or 300 mg·kg(-1) for 3 days) or DEA (15 or 50 mg·kg(-1)·day(-1), loading dose 100 or 300 mg·kg(-1) for 3 days), was applied for 1 month before the coronary artery occlusion. Chronic oral treatment with DEA (50 mg·kg(-1)·day(-1)) resulted in a similar myocardial DEA concentration as chronic AMIO treatment (100 mg·kg(-1)·day(-1)) in rats (7.4 ± 0.7 μg·g(-1) and 8.9 ± 2.2 μg·g(-1)). Both pretreatments in the larger doses significantly improved the survival rate during the acute phase of experimental myocardial infarction (82% and 64% by AMIO and DEA, respectively, vs. 31% in controls). Our results demonstrate that chronic oral treatment with DEA resulted in similar cardiac tissue levels to that of chronic AMIO treatment, and offered an equivalent degree of antiarrhythmic effect against acute coronary artery ligation induced ventricular arrhythmias in conscious rats.

Quantifying intracellular hydrogen peroxide perturbations in terms of concentration

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Beijing K. Huang

2014-01-01

Full Text Available Molecular level, mechanistic understanding of the roles of reactive oxygen species (ROS in a variety of pathological conditions is hindered by the difficulties associated with determining the concentration of various ROS species. Here, we present an approach that converts fold-change in the signal from an intracellular sensor of hydrogen peroxide into changes in absolute concentration. The method uses extracellular additions of peroxide and an improved biochemical measurement of the gradient between extracellular and intracellular peroxide concentrations to calibrate the intracellular sensor. By measuring peroxiredoxin activity, we found that this gradient is 650-fold rather than the 7–10-fold that is widely cited. The resulting calibration is important for understanding the mass-action kinetics of complex networks of redox reactions, and it enables meaningful characterization and comparison of outputs from endogenous peroxide generating tools and therapeutics across studies.

Antibody- and TRIM21-dependent intracellular restriction of Salmonella enterica.

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Rakebrandt, Nikolas; Lentes, Sabine; Neumann, Heinz; James, Leo C; Neumann-Staubitz, Petra

2014-11-01

TRIM21 ('tripartite motif-containing protein 21', Ro52) is a ubiquitously expressed cytosolic Fc receptor, which has a potent role in protective immunity against nonenveloped viruses. TRIM21 mediates intracellular neutralisation of antibody-coated viruses, a process called ADIN (antibody-dependent intracellular neutralisation). Our results reveal a similar mechanism to fight bacterial infections. TRIM21 is recruited to the intracellular pathogen Salmonella enterica in epithelial cells early in infection. TRIM21 does not bind directly to S. enterica, but to antibodies opsonising it. Most importantly, bacterial restriction is dependent on TRIM21 as well as on the opsonisation state of the bacteria. Finally, Salmonella and TRIM21 colocalise with the autophagosomal marker LC3, and intracellular defence is enhanced in starved cells suggesting an involvement of the autophagocytic pathway. Our data extend the protective role of TRIM21 from viruses to bacteria and thereby strengthening the general role of ADIN in cellular immunity. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Data for automated, high-throughput microscopy analysis of intracellular bacterial colonies using spot detection

DEFF Research Database (Denmark)

Ernstsen, Christina Lundgaard; Login, Frédéric H.; Jensen, Helene Halkjær

2017-01-01

Quantification of intracellular bacterial colonies is useful in strategies directed against bacterial attachment, subsequent cellular invasion and intracellular proliferation. An automated, high-throughput microscopy-method was established to quantify the number and size of intracellular bacteria...

Cationic Antimicrobial Peptide LL-37 Is Effective against both Extra- and Intracellular Staphylococcus aureus

Science.gov (United States)

Noore, Jabeen; Noore, Adly

2013-01-01

The increasing resistance of bacteria to conventional antibiotics and the challenges posed by intracellular bacteria, which may be responsible for chronic and recurrent infections, have driven the need for advanced antimicrobial drugs for effective elimination of both extra- and intracellular pathogens. The purpose of this study was to determine the killing efficacy of cationic antimicrobial peptide LL-37 compared to conventional antibiotics against extra- and intracellular Staphylococcus aureus. Bacterial killing assays and an infection model of osteoblasts and S. aureus were studied to determine the bacterial killing efficacy of LL-37 and conventional antibiotics against extra- and intracellular S. aureus. We found that LL-37 was effective in killing extracellular S. aureus at nanomolar concentrations, while lactoferricin B was effective at micromolar concentrations and doxycycline and cefazolin at millimolar concentrations. LL-37 was surprisingly more effective in killing the clinical strain than in killing an ATCC strain of S. aureus. Moreover, LL-37 was superior to conventional antibiotics in eliminating intracellular S. aureus. The kinetic studies further revealed that LL-37 was fast in eliminating both extra- and intracellular S. aureus. Therefore, LL-37 was shown to be very potent and prompt in eliminating both extra- and intracellular S. aureus and was more effective in killing extra- and intracellular S. aureus than commonly used conventional antibiotics. LL-37 could potentially be used to treat chronic and recurrent infections due to its effectiveness in eliminating not only extracellular but also intracellular pathogens. PMID:23274662

Intracellular mechanisms of solar water disinfection

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Castro-Alférez, María; Polo-López, María Inmaculada; Fernández-Ibáñez, Pilar

2016-12-01

Solar water disinfection (SODIS) is a zero-cost intervention measure to disinfect drinking water in areas of poor access to improved water sources, used by more than 6 million people in the world. The bactericidal action of solar radiation in water has been widely proven, nevertheless the causes for this remain still unclear. Scientific literature points out that generation of reactive oxygen species (ROS) inside microorganisms promoted by solar light absorption is the main reason. For the first time, this work reports on the experimental measurement of accumulated intracellular ROS in E. coli during solar irradiation. For this experimental achievement, a modified protocol based on the fluorescent probe dichlorodihydrofluorescein diacetate (DCFH-DA), widely used for oxidative stress in eukaryotic cells, has been tested and validated for E. coli. Our results demonstrate that ROS and their accumulated oxidative damages at intracellular level are key in solar water disinfection.

Subversion of the cytoskeleton by intracellular bacteria: lessons from Listeria, Salmonella, and Vibrio

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de Souza Santos, Marcela; Orth, Kim

2018-01-01

Summary Entry into host cells and intracellular persistence by invasive bacteria are tightly coupled to the ability of the bacterium to disrupt the eukaryotic cytoskeletal machinery. Herein we review the main strategies used by three intracellular pathogens to harness key modulators of the cytoskeleton. Two of these bacteria, namely Listeria monocytogenes and Salmonella enterica serovar Typhimurium, exhibit quite distinct intracellular lifestyles, and therefore, provide a comprehensive panel for the understanding of the intricate bacteria-cytoskeleton interplay during infections. The emerging intracellular pathogen Vibrio parahaemolyticus is depicted as a developing model for the uncovering of novel mechanisms used to hijack the cytoskeleton. PMID:25440316

MgATP hydrolysis destabilizes the interaction between subunit H and yeast V1-ATPase, highlighting H's role in V-ATPase regulation by reversible disassembly.

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Sharma, Stuti; Oot, Rebecca A; Wilkens, Stephan

2018-05-12

Vacuolar H+-ATPases (V-ATPases; V1Vo-ATPases) are rotary motor proton pumps that acidify intracellular compartments and in some tissues, the extracellular space. V-ATPase is regulated by reversible disassembly into autoinhibited V1-ATPase and Vo proton channel sectors. An important player in V-ATPase regulation is subunit H, which binds at the interface of V1 and Vo. H is required for MgATPase activity in holo V-ATPase, but also for stabilizing the MgADP inhibited state in membrane detached V1. However, how H fulfills these two functions is poorly understood. To characterize the H-V1 interaction and its role in reversible disassembly, we determined binding affinities of full length H and its N-terminal domain (HNT) for an isolated heterodimer of subunits E and G (EG), the N-terminal domain of subunit a (aNT), and V1 lacking subunit H (V1ΔH). Using isothermal titration calorimetry (ITC) and biolayer interferometry (BLI), we show that HNT binds EG with moderate affinity, that full length H binds aNT weakly, and that both H and HNT bind V1ΔH with high affinity. We also found that only one molecule of HNT binds V1ΔH with high affinity, suggesting conformational asymmetry of the three EG heterodimers in V1ΔH. Moreover, MgATP hydrolysis-driven conformational changes in V1 destabilized the interaction of H, or HNT, with V1ΔH, suggesting an interplay between MgADP inhibition and subunit H. Our observation that H binding is affected by MgATP hydrolysis in V1 points to H's role in the mechanism of reversible disassembly. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

Tethering factors as organizers of intracellular vesicular traffic.

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Yu, I-Mei; Hughson, Frederick M

2010-01-01

Intracellular trafficking entails the budding, transport, tethering, and fusion of transport vesicles and other membrane carriers. Here we review recent progress toward a mechanistic understanding of vesicle tethering. The known tethering factors are large complexes important for one or more intracellular trafficking pathways and are capable of interacting directly with many of the other principal components of the cellular trafficking machinery. Our review emphasizes recent developments in the in vitro reconstitution of vesicle tethering and the structural characterization of multisubunit tethering factors. The combination of these and other approaches has led to exciting progress toward understanding how these essential nanomachines work.

Concentration-dependent fluorescence live-cell imaging and tracking of intracellular nanoparticles

Energy Technology Data Exchange (ETDEWEB)

Seo, Ji Hye; Joo, Sang-Woo [Department of Chemistry, Soongsil University, Seoul 156-743 (Korea, Republic of); Cho, Keunchang [Logos Biosystems, Incorporated, Anyang 431-070 (Korea, Republic of); Lee, So Yeong, E-mail: leeso@snu.ac.kr, E-mail: sjoo@ssu.ac.kr [Laboratory of Pharmacology, College of Veterinary Medicine and Research Institute for Veterinary Science, Seoul National University, Seoul 151-742 (Korea, Republic of)

2011-06-10

Using live-cell imaging techniques we investigated concentration-dependent intracellular movements of fluorescence nanoparticles (NPs) in real-time after their entry into HeLa cells via incubation. Intracellular particle traces appeared to be a mixture of both random and fairly unidirectional movements of the particles. At rather low concentrations of NPs, a majority of the non-random intracellular particle trajectories are assumed to mostly go along microtubule networks after endocytosis, as evidenced from the inhibition test with nocodazole. On the other hand, as the concentrations of NPs increased, random motions were more frequently observed inside the cells.

Concentration-dependent fluorescence live-cell imaging and tracking of intracellular nanoparticles

International Nuclear Information System (INIS)

Seo, Ji Hye; Joo, Sang-Woo; Cho, Keunchang; Lee, So Yeong

2011-01-01

Using live-cell imaging techniques we investigated concentration-dependent intracellular movements of fluorescence nanoparticles (NPs) in real-time after their entry into HeLa cells via incubation. Intracellular particle traces appeared to be a mixture of both random and fairly unidirectional movements of the particles. At rather low concentrations of NPs, a majority of the non-random intracellular particle trajectories are assumed to mostly go along microtubule networks after endocytosis, as evidenced from the inhibition test with nocodazole. On the other hand, as the concentrations of NPs increased, random motions were more frequently observed inside the cells.

Fluorescent probes and nanoparticles for intracellular sensing of pH values

Science.gov (United States)

Shi, Wen; Li, Xiaohua; Ma, Huimin

2014-12-01

Intracellular pH regulates a number of cell metabolism processes and its sensing is thus of great importance for cell studies. Among various methods, fluorescent probes have been widely used for sensing intracellular pH values because of their high sensitivity and spatiotemporal resolution capability. In this article, the development of fluorescent probes with good practicability in sensing intracellular pH values and pH variation during 2009 - 2014 is reviewed. These fluorescence probes are divided into two kinds: small molecules and nanoparticles. Photophysical properties, advantages/disadvantages and applications of the two kinds of probes are discussed in detail.

Destabilization of the Outer and Inner Mitochondrial Membranes by Core and Linker Histones

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Cascone, Annunziata; Bruelle, Celine; Lindholm, Dan; Bernardi, Paolo; Eriksson, Ove

2012-01-01

Background Extensive DNA damage leads to apoptosis. Histones play a central role in DNA damage sensing and may mediate signals of genotoxic damage to cytosolic effectors including mitochondria. Methodology/Principal Findings We have investigated the effects of histones on mitochondrial function and membrane integrity. We demonstrate that both linker histone H1 and core histones H2A, H2B, H3, and H4 bind strongly to isolated mitochondria. All histones caused a rapid and massive release of the pro-apoptotic intermembrane space proteins cytochrome c and Smac/Diablo, indicating that they permeabilize the outer mitochondrial membrane. In addition, linker histone H1, but not core histones, permeabilized the inner membrane with a collapse of the membrane potential, release of pyridine nucleotides, and mitochondrial fragmentation. Conclusions We conclude that histones destabilize the mitochondrial membranes, a mechanism that may convey genotoxic signals to mitochondria and promote apoptosis following DNA damage. PMID:22523586

Combinatorial release of dexamethasone and amiodarone from a nano-structured parylene-C film to reduce perioperative inflammation and atrial fibrillation

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Robinson, Erik; Kaushal, Sunjay; Alaboson, Justice; Sharma, Sudhish; Belagodu, Amogh; Watkins, Claire; Walker, Brandon; Webster, Gregory; McCarthy, Patrick; Ho, Dean

2016-02-01

Suppressing perioperative inflammation and post-operative atrial fibrillation requires effective drug delivery platforms (DDP). Localized anti-inflammatory and anti-arrhythmic agent release may be more effective than intravenous treatment to improve patient outcomes. This study utilized a dexamethasone (DEX) and amiodarone (AMIO)-loaded Parylene-C (PPX) nano-structured film to inhibit inflammation and atrial fibrillation. The PPX film was tested in an established pericardial adhesion rabbit model. Following sternotomy, the anterior pericardium was resected and the epicardium was abraded. Rabbits were randomly assigned to five treatment groups: control, oxidized PPX (PPX-Oxd), PPX-Oxd infused with DEX (PPX-Oxd[DEX]), native PPX (PPX), and PPX infused with DEX and AMIO (PPX[AMIO, DEX]). 4 weeks post-sternotomy, pericardial adhesions were evaluated for gross adhesions using a 4-point grading system and histological evaluation for epicardial neotissue fibrosis (NTF). Atrial fibrillation duration and time per induction were measured. The PPX[AMIO, DEX] group had a significant reduction in mean adhesion score compared with the control group (control 2.75 +/- 0.42 vs. PPX[AMIO, DEX] 0.25 +/- 0.42, P atrial fibrillation was decreased in rabbits with PPX[AMIO, DEX] films compared to control (9.5 +/- 6.8 s vs. 187.6 +/- 174.7 s, p = 0.003). Time of atrial fibrillation per successful induction decreased among PPX[AMIO, DEX] films compared to control (2.8 +/- 1.2 s vs. 103.2 +/- 178 s, p = 0.004). DEX/AMIO-loaded PPX films are associated with reduced perioperative inflammation and a diminished atrial fibrillation duration. Epicardial application of AMIO, DEX films is a promising strategy to prevent post-operative cardiac complications.Suppressing perioperative inflammation and post-operative atrial fibrillation requires effective drug delivery platforms (DDP). Localized anti-inflammatory and anti-arrhythmic agent release may be more effective than intravenous treatment to

Biomineralization Patterns of Intracellular Carbonatogenesis in Cyanobacteria: Molecular Hypotheses

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Jinhua Li

2016-02-01

Full Text Available The recent discovery of intracellular carbonatogenesis in several cyanobacteria species has challenged the traditional view that this process was extracellular and not controlled. However, a detailed analysis of the size distribution, chemical composition and 3-D-arrangement of carbonates in these cyanobacteria is lacking. Here, we characterized these features in Candidatus Gloeomargarita lithophora C7 and Candidatus Synechococcus calcipolaris G9 by conventional transmission electron microscopy, tomography, ultramicrotomy, and scanning transmission X-ray microscopy (STXM. Both Ca. G. lithophora C7 and Ca. S. calcipolaris G9 formed numerous polyphosphate granules adjacent or engulfing Ca-carbonate inclusions when grown in phosphate-rich solutions. Ca-carbonates were scattered within Ca. G. lithophora C7 cells under these conditions, but sometimes arranged in one or several chains. In contrast, Ca-carbonates formed at cell septa in Ca. S. calcipolaris G9 and were segregated equally between daughter cells after cell division, arranging as distorted disks at cell poles. The size distribution of carbonates evolved from a positively to a negatively skewed distribution as particles grew. Conventional ultramicrotomy did not preserve Ca-carbonates explaining partly why intracellular calcification has been overlooked in the past. All these new observations allow discussing with unprecedented insight some nucleation and growth processes occurring in intracellularly calcifying cyanobacteria with a particular emphasis on the possible involvement of intracellular compartments and cytoskeleton.

Detection and quantification of intracellular bacterial colonies by automated, high-throughput microscopy.

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Ernstsen, Christina L; Login, Frédéric H; Jensen, Helene H; Nørregaard, Rikke; Møller-Jensen, Jakob; Nejsum, Lene N

2017-08-01

To target bacterial pathogens that invade and proliferate inside host cells, it is necessary to design intervention strategies directed against bacterial attachment, cellular invasion and intracellular proliferation. We present an automated microscopy-based, fast, high-throughput method for analyzing size and number of intracellular bacterial colonies in infected tissue culture cells. Cells are seeded in 48-well plates and infected with a GFP-expressing bacterial pathogen. Following gentamicin treatment to remove extracellular pathogens, cells are fixed and cell nuclei stained. This is followed by automated microscopy and subsequent semi-automated spot detection to determine the number of intracellular bacterial colonies, their size distribution, and the average number per host cell. Multiple 48-well plates can be processed sequentially and the procedure can be completed in one working day. As a model we quantified intracellular bacterial colonies formed by uropathogenic Escherichia coli (UPEC) during infection of human kidney cells (HKC-8). Urinary tract infections caused by UPEC are among the most common bacterial infectious diseases in humans. UPEC can colonize tissues of the urinary tract and is responsible for acute, chronic, and recurrent infections. In the bladder, UPEC can form intracellular quiescent reservoirs, thought to be responsible for recurrent infections. In the kidney, UPEC can colonize renal epithelial cells and pass to the blood stream, either via epithelial cell disruption or transcellular passage, to cause sepsis. Intracellular colonies are known to be clonal, originating from single invading UPEC. In our experimental setup, we found UPEC CFT073 intracellular bacterial colonies to be heterogeneous in size and present in nearly one third of the HKC-8 cells. This high-throughput experimental format substantially reduces experimental time and enables fast screening of the intracellular bacterial load and cellular distribution of multiple

Intracellular sodium hydrogen exchange inhibition and clinical myocardial protection.

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Mentzer, Robert M; Lasley, Robert D; Jessel, Andreas; Karmazyn, Morris

2003-02-01

Although the mechanisms underlying ischemia/reperfusion injury remain elusive, evidence supports the etiologic role of intracellular calcium overload and oxidative stress induced by reactive oxygen species. Activation of the sodium hydrogen exchanger (NHE) is associated with intracellular calcium accumulation. Inhibition of the NHE-1 isoform may attenuate the consequences of this injury. Although there is strong preclinical and early clinical evidence that NHE inhibitors may be cardioprotective, definitive proof of this concept in humans awaits the results of ongoing clinical trials.

The effect of sodium bicarbonate on intracellular pH using 31P-MR spectroscopy

International Nuclear Information System (INIS)

Nakashima, Kazuya; Kashiwagi, Shiro; Ito, Haruhide; Yamashita, Tetsuo; Kitahara, Tetsuhiro; Nakayama, Naoto; Saito, Kennichi

1997-01-01

This report deals with the effects of sodium bicarbonate on the intracellular pH of the brain and cerebral blood flow (CBF); five normal volunteers were studied. Intracellular pH and CBF were measured by phosphorus 31 magnetic resonance spectroscopy ( 31 P-MRS) and stable xenon computed tomography (Xe-CT), respectively. Each individual received 7% sodium bicarbonate (3.5 ml/kg body weight), infused intravenously over a 15-min period. Intracellular pH, CBF, and physiological parameters were determined before and after the injection. Intracellular pH was significantly decreased and CBF was increased. Among the physiological parameters, the hematocrit was significantly decreased and arterial pressure of carbon dioxide (PaCO 2 ), increased. These results suggest that increasing CO 2 contributes to the decrease in intracellular pH. In conclusion, three factors increase CBF during the administration of sodium bicarbonate to humans: arterial dilatation in response to carbon dioxide; decrease of the hematocrit, and intracellular cerebral acidosis. (author)

Fluorescent nanoparticles for intracellular sensing: A review

International Nuclear Information System (INIS)

Ruedas-Rama, Maria J.; Walters, Jamie D.; Orte, Angel; Hall, Elizabeth A.H.

2012-01-01

Highlights: ► Analytical applications of fluorescent nanoparticles (NPs) in intracellular sensing. ► Critical review on performance of QDots, metal NPs, silica NPs, and polymer NPs. ► Highlighted potential of fluorescence lifetime imaging microscopy (FLIM). - Abstract: Fluorescent nanoparticles (NPs), including semiconductor NPs (Quantum Dots), metal NPs, silica NPs, polymer NPs, etc., have been a major focus of research and development during the past decade. The fluorescent nanoparticles show unique chemical and optical properties, such as brighter fluorescence, higher photostability and higher biocompatibility, compared to classical fluorescent organic dyes. Moreover, the nanoparticles can also act as multivalent scaffolds for the realization of supramolecular assemblies, since their high surface to volume ratio allow distinct spatial domains to be functionalized, which can provide a versatile synthetic platform for the implementation of different sensing schemes. Their excellent properties make them one of the most useful tools that chemistry has supplied to biomedical research, enabling the intracellular monitoring of many different species for medical and biological purposes. In this review, we focus on the developments and analytical applications of fluorescent nanoparticles in chemical and biological sensing within the intracellular environment. The review also points out the great potential of fluorescent NPs for fluorescence lifetime imaging microscopy (FLIM). Finally, we also give an overview of the current methods for delivering of fluorescent NPs into cells, where critically examine the benefits and liabilities of each strategy.

Intracellular renin disrupts chemical communication between heart cells. Pathophysiological implications

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Walmor eDe Mello

2015-01-01

Full Text Available The influence of intracellular renin on the process of chemical communication between cardiac cells was investigated in cell pairs isolated from the left ventricle of adult Wistar Kyoto rats. The enzyme together with Lucifer yellow CH was dialyzed into one cell of the pair using the whole cell clamp technique. The diffusion of the dye in the dialyzed and in non-dialyzed cell was followed by measuring the intensity of fluorescence in both cells as a function of time. The results indicated that; 1 under normal conditions, Lucifer Yellow flows from cell-to-cell through gap junctions; 2 the intracellular dialysis of renin (100nM disrupts chemical communication-an effect enhanced by simultaneous administration of angiotensinogen (100nM; 3 enalaprilat (10-9M administered to the cytosol together with renin reduced drastically the uncoupling action of the enzyme; 4 aliskiren (10-8M inhibited the effect of renin on chemical communication;5 the possible role of intracellular renin independently of angiotensin II (Ang II was evaluated including the increase of the inward calcium current elicited by the enzyme and the possible role of oxidative stress on the disruption of cell communication; 6 the possible harmful versus the beneficial effect of intracellular renin during myocardial infarction was discussed;7 the present results indicate that intracellular renin due to internalization or in situ synthesis, causes a severe impairment of chemical communication in the heart resulting in derangement of metabolic cooperation with serious consequences for heart function.

Stabilizing and destabilizing forces in the nursing work environment: a qualitative study on turnover intention.

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Choi, Sandy Pin-Pin; Pang, Samantha Mei-Che; Cheung, Kin; Wong, Thomas Kwok-Shing

2011-10-01

The nursing work environment, which provides the context of care delivery, has been gaining increasing attention in recent years. A growing body of evidence points to an inseparable link between attributes of the nursing work environment and nurse and patient outcomes. While most studies have adopted a survey design to examine the workforce and work environment issues, this study employed a phenomenological approach to provide empirical evidence regarding nurses' perceptions of their work and work environment. The aim of this study was to advance our understanding of the phenomenon of increasing nurse turnover through exploring frontline registered nurses' lived experiences of working in Hong Kong public hospitals. A modified version of Van Kaam's controlled explication method was adopted. Individual semi-structured interviews were conducted with 26 frontline nurses recruited from ten acute regional, district and non-acute public hospitals in Hong Kong. Their perspectives in regard to their work and work environment, such as workload, manpower demand and professional values, were extensively examined, and a hypothetical description relating the nursing work environment with nurses' turnover intention was posited. Contemplation of nurses' experiences revealed the vulnerable aspects of nursing work and six essential constituents of the nursing work environment, namely staffing level, work responsibility, management, co-worker relationships, job, and professional incentives. These essential constituents have contributed to two sets of forces, stabilizing and destabilizing forces, which originate from the attributes of the nursing work environment. Nurses viewed harmonious co-worker relationships, recognition and professional development as the crucial retaining factors. However, nurses working in an unfavorable environment were overwhelmed by destabilizing forces; they expressed frustration and demonstrated an intention to leave their work environment. The nursing

Fatty Acid Signaling: The New Function of Intracellular Lipases

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Zuzana Papackova

2015-02-01

Full Text Available Until recently, intracellular triacylglycerols (TAG stored in the form of cytoplasmic lipid droplets have been considered to be only passive “energy conserves”. Nevertheless, degradation of TAG gives rise to a pleiotropic spectrum of bioactive intermediates, which may function as potent co-factors of transcription factors or enzymes and contribute to the regulation of numerous cellular processes. From this point of view, the process of lipolysis not only provides energy-rich equivalents but also acquires a new regulatory function. In this review, we will concentrate on the role that fatty acids liberated from intracellular TAG stores play as signaling molecules. The first part provides an overview of the transcription factors, which are regulated by fatty acids derived from intracellular stores. The second part is devoted to the role of fatty acid signaling in different organs/tissues. The specific contribution of free fatty acids released by particular lipases, hormone-sensitive lipase, adipose triacylglycerol lipase and lysosomal lipase will also be discussed.

Data for automated, high-throughput microscopy analysis of intracellular bacterial colonies using spot detection.

Science.gov (United States)

Ernstsen, Christina L; Login, Frédéric H; Jensen, Helene H; Nørregaard, Rikke; Møller-Jensen, Jakob; Nejsum, Lene N

2017-10-01

Quantification of intracellular bacterial colonies is useful in strategies directed against bacterial attachment, subsequent cellular invasion and intracellular proliferation. An automated, high-throughput microscopy-method was established to quantify the number and size of intracellular bacterial colonies in infected host cells (Detection and quantification of intracellular bacterial colonies by automated, high-throughput microscopy, Ernstsen et al., 2017 [1]). The infected cells were imaged with a 10× objective and number of intracellular bacterial colonies, their size distribution and the number of cell nuclei were automatically quantified using a spot detection-tool. The spot detection-output was exported to Excel, where data analysis was performed. In this article, micrographs and spot detection data are made available to facilitate implementation of the method.

Fluorescent probes and nanoparticles for intracellular sensing of pH values

International Nuclear Information System (INIS)

Shi, Wen; Li, Xiaohua; Ma, Huimin

2014-01-01

Intracellular pH regulates a number of cell metabolism processes and its sensing is thus of great importance for cell studies. Among various methods, fluorescent probes have been widely used for sensing intracellular pH values because of their high sensitivity and spatiotemporal resolution capability. In this article, the development of fluorescent probes with good practicability in sensing intracellular pH values and pH variation during 2009 − 2014 is reviewed. These fluorescence probes are divided into two kinds: small molecules and nanoparticles. Photophysical properties, advantages/disadvantages and applications of the two kinds of probes are discussed in detail. (topical review)

A method for functional trans-complementation of intracellular Francisella tularensis.

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Shaun Steele

Full Text Available Francisella tularensis is a highly infectious bacterial pathogen that invades and replicates within numerous host cell types. After uptake, F. tularensis bacteria escape the phagosome, replicate within the cytosol, and suppress cytokine responses. However, the mechanisms employed by F. tularensis to thrive within host cells are mostly unknown. Potential F. tularensis mutants involved in host-pathogen interactions are typically discovered by negative selection screens for intracellular replication or virulence. Mutants that fulfill these criteria fall into two categories: mutants with intrinsic intracellular growth defects and mutants that fail to modify detrimental host cell processes. It is often difficult and time consuming to discriminate between these two possibilities. We devised a method to functionally trans-complement and thus identify mutants that fail to modify the host response. In this assay, host cells are consistently and reproducibly infected with two different F. tularensis strains by physically tethering the bacteria to antibody-coated beads. To examine the efficacy of this protocol, we tested phagosomal escape, cytokine suppression, and intracellular replication for F. tularensis ΔripA and ΔpdpC. ΔripA has an intracellular growth defect that is likely due to an intrinsic defect and fails to suppress IL-1β secretion. In the co-infection model, ΔripA was unable to replicate in the host cell when wild-type bacteria infected the same cell, but cytokine suppression was rescued. Therefore, ΔripA intracellular growth is due to an intrinsic bacterial defect while cytokine secretion results from a failed host-pathogen interaction. Likewise, ΔpdpC is deficient for phagosomal escape, intracellular survival and suppression of IL-1β secretion. Wild-type bacteria that entered through the same phagosome as ΔpdpC rescued all of these phenotypes, indicating that ΔpdpC failed to properly manipulate the host. In summary, functional

Optimized Reaction Conditions for Removal of Cellular Organic Matter of Microcystis aeruginosa During the Destabilization and Aggregation Process Using Ferric Sulfate in Water Purification

Czech Academy of Sciences Publication Activity Database

Pivokonský, Martin; Polášek, Pavel; Pivokonská, Lenka; Tomášková, Hana

2009-01-01

Roč. 81, č. 5 (2009), s. 514-522 ISSN 1061-4303 R&D Projects: GA ČR GA103/07/0295 Institutional research plan: CEZ:AV0Z20600510 Keywords : Microcystis aeruginosa * cellular organic matter * destabilization * aggregation * optimized reaction conditions * water purification Subject RIV: BK - Fluid Dynamics Impact factor: 0.965, year: 2009

Leishmania hijacking of the macrophage intracellular compartments.

Science.gov (United States)

Liévin-Le Moal, Vanessa; Loiseau, Philippe M

2016-02-01

Leishmania spp., transmitted to humans by the bite of the sandfly vector, are responsible for the three major forms of leishmaniasis, cutaneous, diffuse mucocutaneous and visceral. Leishmania spp. interact with membrane receptors of neutrophils and macrophages. In macrophages, the parasite is internalized within a parasitophorous vacuole and engages in a particular intracellular lifestyle in which the flagellated, motile Leishmania promastigote metacyclic form differentiates into non-motile, metacyclic amastigote form. This phenomenon is induced by Leishmania-triggered events leading to the fusion of the parasitophorous vacuole with vesicular members of the host cell endocytic pathway including recycling endosomes, late endosomes and the endoplasmic reticulum. Maturation of the parasitophorous vacuole leads to the intracellular proliferation of the Leishmania amastigote forms by acquisition of host cell nutrients while escaping host defense responses. © 2015 FEBS.

pH-sensitive intracellular photoluminescence of carbon nanotube-fluorescein conjugates in human ovarian cancer cells

International Nuclear Information System (INIS)

Chen, M T; Ishikawa, F N; Gundersen, M A; Gomez, L M; Vernier, P T; Zhou, C

2009-01-01

To add to the understanding of the properties of functionalized carbon nanotubes in biological applications, we report a monotonic pH sensitivity of the intracellular fluorescence emission of single-walled carbon nanotube-fluorescein carbazide (SWCNT-FC) conjugates in human ovarian cancer cells. Light-stimulated intracellular hydrolysis of the amide linkage and localized intracellular pH changes are proposed as mechanisms. SWCNT-FC conjugates may serve as intracellular pH sensors.

Isoflurane reversibly destabilizes hippocampal dendritic spines by an actin-dependent mechanism.

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Jimcy Platholi

Full Text Available General anesthetics produce a reversible coma-like state through modulation of excitatory and inhibitory synaptic transmission. Recent evidence suggests that anesthetic exposure can also lead to sustained cognitive dysfunction. However, the subcellular effects of anesthetics on the structure of established synapses are not known. We investigated effects of the widely used volatile anesthetic isoflurane on the structural stability of hippocampal dendritic spines, a postsynaptic structure critical to excitatory synaptic transmission in learning and memory. Exposure to clinical concentrations of isoflurane induced rapid and non-uniform shrinkage and loss of dendritic spines in mature cultured rat hippocampal neurons. Spine shrinkage was associated with a reduction in spine F-actin concentration. Spine loss was prevented by either jasplakinolide or cytochalasin D, drugs that prevent F-actin disassembly. Isoflurane-induced spine shrinkage and loss were reversible upon isoflurane elimination. Thus, isoflurane destabilizes spine F-actin, resulting in changes to dendritic spine morphology and number. These findings support an actin-based mechanism for isoflurane-induced alterations of synaptic structure in the hippocampus. These reversible alterations in dendritic spine structure have important implications for acute anesthetic effects on excitatory synaptic transmission and synaptic stability in the hippocampus, a locus for anesthetic-induced amnesia, and have important implications for anesthetic effects on synaptic plasticity.

Alanine Counteracts the Destabilizing Effect that Urea has on RNase-A.

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Chowhan, Rimpy K; Ali, Fasil; Bhat, Mohd Y; Rahman, Safikur; Singh, Laishram R; Ahmad, Faizan; Dar, Tanveer A

2016-01-01

It is generally believed that organisms use and accumulate methylamine osmolytes to prevent urea's damaging effect on protein stability and activity. However, urea-rich cells not only accumulate methylamines but also many other methylated and non-methylated compounds as well. But, so far it is not known whether osmolytes that are not accumulated in urea-rich cells could also confer urea-counteracting properties. We investigated the behavior of a non-methylamine osmolyte, alanine for its counteracting effect against urea denaturation of a model protein, ribonuclease A (RNase-A). We have measured structure and thermodynamic parameters (Tm, ΔHm, and ΔGD°) of RNase-A in the presence of alanine, urea and their combination. The results were also compared with the ability of glycine (osmolyte lacking one methyl group when compared with alanine) to counter urea's effect on protein stability. We observed that alanine but not glycine counteracts urea's harmful effect on RNase-A stability. The results indicated that alanine (in addition to methylamine osmolytes) may serve as an alternate urea-counteractant. Since glycine fails to protect RNase-A from urea's destabilizing effect, it seems that methylation to glycine might have some evolutionary significance to protect proteins against harmful effects of urea.

Cortisol and testosterone increase financial risk taking and may destabilize markets

Science.gov (United States)

Cueva, Carlos; Roberts, R. Edward; Spencer, Tom; Rani, Nisha; Tempest, Michelle; Tobler, Philippe N.; Herbert, Joe; Rustichini, Aldo

2015-01-01

It is widely known that financial markets can become dangerously unstable, yet it is unclear why. Recent research has highlighted the possibility that endogenous hormones, in particular testosterone and cortisol, may critically influence traders’ financial decision making. Here we show that cortisol, a hormone that modulates the response to physical or psychological stress, predicts instability in financial markets. Specifically, we recorded salivary levels of cortisol and testosterone in people participating in an experimental asset market (N = 142) and found that individual and aggregate levels of endogenous cortisol predict subsequent risk-taking and price instability. We then administered either cortisol (single oral dose of 100 mg hydrocortisone, N = 34) or testosterone (three doses of 10 g transdermal 1% testosterone gel over 48 hours, N = 41) to young males before they played an asset trading game. We found that both cortisol and testosterone shifted investment towards riskier assets. Cortisol appears to affect risk preferences directly, whereas testosterone operates by inducing increased optimism about future price changes. Our results suggest that changes in both cortisol and testosterone could play a destabilizing role in financial markets through increased risk taking behaviour, acting via different behavioural pathways. PMID:26135946

PKC-η-MARCKS Signaling Promotes Intracellular Survival of Unopsonized Burkholderia thailandensis.

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Micheva-Viteva, Sofiya N; Shou, Yulin; Ganguly, Kumkum; Wu, Terry H; Hong-Geller, Elizabeth

2017-01-01

Pathogenic Burkholderia rely on host factors for efficient intracellular replication and are highly refractory to antibiotic treatment. To identify host genes that are required by Burkholderia spp. during infection, we performed a RNA interference (RNAi) screen of the human kinome and identified 35 host kinases that facilitated Burkholderia thailandensis intracellular survival in human monocytic THP-1 cells. We validated a selection of host kinases using imaging flow cytometry to assess efficiency of B. thailandensis survival in the host upon siRNA-mediated knockdown. We focused on the role of the novel protein kinase C isoform, PKC-η, in Burkholderia infection and characterized PKC-η/MARCKS signaling as a key event that promotes the survival of unopsonized B. thailandensis CDC2721121 within host cells. While infection of lung epithelial cells with unopsonized Gram-negative bacteria stimulated phosphorylation of Ser175/160 in the MARCKS effector domain, siRNA-mediated knockdown of PKC-η expression reduced the levels of phosphorylated MARCKS by >3-fold in response to infection with Bt CDC2721121. We compared the effect of the conventional PKC-α and novel PKC-η isoforms on the growth of B. thailandensis CDC2721121 within monocytic THP-1 cells and found that ≥75% knock-down of PRKCH transcript levels reduced intracellular bacterial load 100% more efficiently when compared to growth in cells siRNA-depleted of the classical PKC-α, suggesting that the PKC-η isoform can specifically mediate Burkholderia intracellular survival. Based on imaging studies of intracellular B. thailandensis , we found that PKC-η function stimulates phagocytic pathways that promote B. thailandensis escape into the cytoplasm leading to activation of autophagosome flux. Identification of host kinases that are targeted by Burkholderia during infection provides valuable molecular insights in understanding Burkholderia pathogenesis, and ultimately, in designing effective host

Microsporidia are natural intracellular parasites of the nematode Caenorhabditis elegans.

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Troemel, Emily R; Félix, Marie-Anne; Whiteman, Noah K; Barrière, Antoine; Ausubel, Frederick M

2008-12-09

For decades the soil nematode Caenorhabditis elegans has been an important model system for biology, but little is known about its natural ecology. Recently, C. elegans has become the focus of studies of innate immunity and several pathogens have been shown to cause lethal intestinal infections in C. elegans. However none of these pathogens has been shown to invade nematode intestinal cells, and no pathogen has been isolated from wild-caught C. elegans. Here we describe an intracellular pathogen isolated from wild-caught C. elegans that we show is a new species of microsporidia. Microsporidia comprise a large class of eukaryotic intracellular parasites that are medically and agriculturally important, but poorly understood. We show that microsporidian infection of the C. elegans intestine proceeds through distinct stages and is transmitted horizontally. Disruption of a conserved cytoskeletal structure in the intestine called the terminal web correlates with the release of microsporidian spores from infected cells, and appears to be part of a novel mechanism by which intracellular pathogens exit from infected cells. Unlike in bacterial intestinal infections, the p38 MAPK and insulin/insulin-like growth factor (IGF) signaling pathways do not appear to play substantial roles in resistance to microsporidian infection in C. elegans. We found microsporidia in multiple wild-caught isolates of Caenorhabditis nematodes from diverse geographic locations. These results indicate that microsporidia are common parasites of C. elegans in the wild. In addition, the interaction between C. elegans and its natural microsporidian parasites provides a system in which to dissect intracellular intestinal infection in vivo and insight into the diversity of pathogenic mechanisms used by intracellular microbes.

DMPD: NOD-like receptors (NLRs): bona fide intracellular microbial sensors. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 18585455 NOD-like receptors (NLRs): bona fide intracellular microbial sensors. Shaw...tml) (.csml) Show NOD-like receptors (NLRs): bona fide intracellular microbial sensors. PubmedID 18585455 Ti...tle NOD-like receptors (NLRs): bona fide intracellular microbial sensors. Authors

Intracellular Cholesterol Trafficking and Impact in Neurodegeneration

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Fabian Arenas

2017-11-01

Full Text Available Cholesterol is a critical component of membrane bilayers where it plays key structural and functional roles by regulating the activity of diverse signaling platforms and pathways. Particularly enriched in brain, cholesterol homeostasis in this organ is singular with respect to other tissues and exhibits a heterogeneous regulation in distinct brain cell populations. Due to the key role of cholesterol in brain physiology and function, alterations in cholesterol homeostasis and levels have been linked to brain diseases and neurodegeneration. In the case of Alzheimer disease (AD, however, this association remains unclear with evidence indicating that either increased or decreased total brain cholesterol levels contribute to this major neurodegenerative disease. Here, rather than analyzing the role of total cholesterol levels in neurodegeneration, we focus on the contribution of intracellular cholesterol pools, particularly in endolysosomes and mitochondria through its trafficking via specialized membrane domains delineated by the contacts between endoplasmic reticulum and mitochondria, in the onset of prevalent neurodegenerative diseases such as AD, Parkinson disease, and Huntington disease as well as in lysosomal disorders like Niemann-Pick type C disease. We dissect molecular events associated with intracellular cholesterol accumulation, especially in mitochondria, an event that results in impaired mitochondrial antioxidant defense and function. A better understanding of the mechanisms involved in the distribution of cholesterol in intracellular compartments may shed light on the role of cholesterol homeostasis disruption in neurodegeneration and may pave the way for specific intervention opportunities.

Fluorescent nanoparticles for intracellular sensing: A review

Energy Technology Data Exchange (ETDEWEB)

Ruedas-Rama, Maria J., E-mail: mjruedas@ugr.esmailto [Department of Physical Chemistry, Faculty of Pharmacy, University of Granada, Campus Cartuja, 18071, Granada (Spain); Walters, Jamie D. [Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, UK CB2 1QT (United Kingdom); Orte, Angel [Department of Physical Chemistry, Faculty of Pharmacy, University of Granada, Campus Cartuja, 18071, Granada (Spain); Hall, Elizabeth A.H., E-mail: lisa.hall@biotech.cam.ac.uk [Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QT (United Kingdom)

2012-11-02

Highlights: Black-Right-Pointing-Pointer Analytical applications of fluorescent nanoparticles (NPs) in intracellular sensing. Black-Right-Pointing-Pointer Critical review on performance of QDots, metal NPs, silica NPs, and polymer NPs. Black-Right-Pointing-Pointer Highlighted potential of fluorescence lifetime imaging microscopy (FLIM). - Abstract: Fluorescent nanoparticles (NPs), including semiconductor NPs (Quantum Dots), metal NPs, silica NPs, polymer NPs, etc., have been a major focus of research and development during the past decade. The fluorescent nanoparticles show unique chemical and optical properties, such as brighter fluorescence, higher photostability and higher biocompatibility, compared to classical fluorescent organic dyes. Moreover, the nanoparticles can also act as multivalent scaffolds for the realization of supramolecular assemblies, since their high surface to volume ratio allow distinct spatial domains to be functionalized, which can provide a versatile synthetic platform for the implementation of different sensing schemes. Their excellent properties make them one of the most useful tools that chemistry has supplied to biomedical research, enabling the intracellular monitoring of many different species for medical and biological purposes. In this review, we focus on the developments and analytical applications of fluorescent nanoparticles in chemical and biological sensing within the intracellular environment. The review also points out the great potential of fluorescent NPs for fluorescence lifetime imaging microscopy (FLIM). Finally, we also give an overview of the current methods for delivering of fluorescent NPs into cells, where critically examine the benefits and liabilities of each strategy.

Reduction of intracellular glutathione content and radiosensitivity

International Nuclear Information System (INIS)

Vos, O.; Schans, G.P. van der; Roos-Verheij, W.S.D.

1986-05-01

The intracellular glutathione (GSH) content in HeLa, CHO and V79 cells was reduced by incubating the cells in growth medium containing buthionine sulfoximine (BSO) or diethyl maleate (DEM). Clonogenicity, single strand DNA breaks (ssb) and double strand DNA breaks (dsb) were used as criteria for radiation induced damage after X- or γ irradiation. In survival experiments DEM gave a slightly larger sensitization although it gave a smaller reduction of the intracellular GSH. In general, sensitization was larger for dsb than for ssb, also the reduction of the OER was generally larger for dsb than for ssb. This may be due to the higher dose rate in case of dsb experiments resulting in a higher rate of radiochemical oxygen consumption. In general, no effect was found on post-irradiation repair of ssb and dsb. (Auth.)

Reduction of intracellular glutathione content and radiosensitivity

International Nuclear Information System (INIS)

Vos, O.; Schans, G.P. van der; Roos-Verheij, W.S.D.

1986-01-01

The intracellular glutathione (GSH) content of HeLa, CHO and V79 cells was reduced by incubating the cells in growth medium containing buthionine sulphoximine or diethyl maleate (DEM). Clonogenicity, single-strand DNA breaks (ssb) and double-strand DNA breaks (dsb) were used as criteria for radiation-induced damage after X- or γ-irradiation. In survival experiments, DEM gave a slightly larger sensitization although it gave a smaller reduction of the intracellular GSH. In general, sensitization was larger for dsb than for ssb, also the reduction of the o.e.r. was generally larger for dsb than for ssb. This may be due to the higher dose rate in case of dsb experiments resulting in a higher rate of radiochemical oxygen consumption. In general, no effect was found on post-irradiation repair of ssb and dsb. (author)

Destabilization of Heterologous Proteins Mediated by the GSK3β Phosphorylation Domain of the β-Catenin Protein

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Yuhan Kong

2013-11-01

Full Text Available Background and Aims: Wnt/β-catenin signaling plays important roles in development and cellular processes. The hallmark of canonical Wnt signaling activation is the stabilization of β-catenin protein in cytoplasm and/or nucleus. The stability of β-catenin is the key to its biological functions and is controlled by the phosphorylation of its amino-terminal degradation domain. Aberrant activation of β-catenin signaling has been implicated in the development of human cancers. It has been recently suggested that GSK3βmay play an essential role in regulating global protein turnover. Here, we investigate if the GSK3β phosphorylation site-containing degradation domain of β-catenin is sufficient to destabilize heterologous proteins. Methods and Results: We engineer chimeric proteins by fusing β-catenin degradation domain at the N- and/or C-termini of the enhanced green fluorescent protein (eGFP. In both transient and stable expression experiments, the chimeric GFP proteins exhibit a significantly decreased stability, which can be effectively antagonized by lithium and Wnt1. An activating mutation in the destruction domain significantly stabilizes the fusion protein. Furthermore, GSK3 inhibitor SB-216763 effectively increases the GFP signal of the fusion protein. Conversely, the inhibition of Wnt signaling with tankyrase inhibitor XAV939 results in a decrease in GFP signal of the fusion proteins, while these small molecules have no significant effects on the mutant destruction domain-GFP fusion protein. Conclusion: Our findings strongly suggest that the β-catenin degradation domain may be sufficient to destabilize heterologous proteins in Wnt signaling-dependent manner. It is conceivable that the chimeric GFP proteins may be used as a functional reporter to measure the dynamic status of β-catenin signaling, and to identify potential anticancer drugs that target β-catenin signaling.

HFE mRNA expression is responsive to intracellular and extracellular iron loading: short communication.

Science.gov (United States)

Mehta, Kosha J; Farnaud, Sebastien; Patel, Vinood B

2017-10-01

In liver hepatocytes, the HFE gene regulates cellular and systemic iron homeostasis by modulating cellular iron-uptake and producing the iron-hormone hepcidin in response to systemic iron elevation. However, the mechanism of iron-sensing in hepatocytes remain enigmatic. Therefore, to study the effect of iron on HFE and hepcidin (HAMP) expressions under distinct extracellular and intracellular iron-loading, we examined the effect of holotransferrin treatment (1, 2, 5 and 8 g/L for 6 h) on intracellular iron levels, and mRNA expressions of HFE and HAMP in wild-type HepG2 and previously characterized iron-loaded recombinant-TfR1 HepG2 cells. Gene expression was analyzed by real-time PCR and intracellular iron was measured by ferrozine assay. Data showed that in the wild-type cells, where intracellular iron content remained unchanged, HFE expression remained unaltered at low holotransferrin treatments but was upregulated upon 5 g/L (p HFE and HAMP expressions were elevated only at low 1 g/L treatment (p HFE (p HFE mRNA was independently elevated by extracellular and intracellular iron-excess. Thus, it may be involved in sensing both, extracellular and intracellular iron. Repression of HAMP expression under simultaneous intracellular and extracellular iron-loading resembles non-hereditary iron-excess pathologies.

[Thyroid hormones and cardiovascular system].

Science.gov (United States)

Límanová, Zdeňka; Jiskra, Jan

Cardiovascular system is essentially affected by thyroid hormones by way of their genomic and non-genomic effects. Untreated overt thyroid dysfunction is associated with higher cardiovascular risk. Although it has been studied more than 3 decades, in subclinical thyroid dysfunction the negative effect on cardiovascular system is much more controversial. Large meta-analyses within last 10 years have shown that subclinical hyperthyroidism is associated with higher cardiovascular risk than subclinical hypothyroidism. Conversely, in patients of age > 85 years subclinical hypothyroidism was linked with lower mortality. Therefore, subclinical hyperthyroidism should be rather treated in the elderly while subclinical hypothyroidism in the younger patients and the older may be just followed. An important problem on the border of endocrinology and cardiology is amiodarone thyroid dysfunction. Effective and safe treatment is preconditioned by distinguishing of type 1 and type 2 amiodarone induced hyperthyroidism. The type 1 should be treated with methimazol, therapeutic response is prolonged, according to recent knowledge immediate discontinuation of amiodarone is not routinely recommended and patient should be usually prepared to total thyroidectomy, or rather rarely 131I radioiodine ablation may be used if there is appropriate accumulation. In the type 2 there is a promt therapeutic response on glucocorticoids (within 1-2 weeks) with permanent remission or development of hypothyroidism. If it is not used for life-threatening arrhytmias, amiodarone may be discontinuated earlier (after several weeks). Amiodarone induced hypothyroidism is treated with levothyroxine without amiodarone interruption.Key words: amiodarone induced thyroid dysfunction - atrial fibrillation - cardiovascular risk - heart failure - hyperthyroidism - hypothyroidism - thyroid stimulating hormone.

Pathogenic mechanisms of intracellular bacteria.

Science.gov (United States)

Niller, Hans Helmut; Masa, Roland; Venkei, Annamária; Mészáros, Sándor; Minarovits, Janos

2017-06-01

We wished to overview recent data on a subset of epigenetic changes elicited by intracellular bacteria in human cells. Reprogramming the gene expression pattern of various host cells may facilitate bacterial growth, survival, and spread. DNA-(cytosine C5)-methyltransferases of Mycoplasma hyorhinis targeting cytosine-phosphate-guanine (CpG) dinucleotides and a Mycobacterium tuberculosis methyltransferase targeting non-CpG sites methylated the host cell DNA and altered the pattern of gene expression. Gene silencing by CpG methylation and histone deacetylation, mediated by cellular enzymes, also occurred in M. tuberculosis-infected macrophages. M. tuberculosis elicited cell type-specific epigenetic changes: it caused increased DNA methylation in macrophages, but induced demethylation, deposition of euchromatic histone marks and activation of immune-related genes in dendritic cells. A secreted transposase of Acinetobacter baumannii silenced a cellular gene, whereas Mycobacterium leprae altered the epigenotype, phenotype, and fate of infected Schwann cells. The 'keystone pathogen' oral bacterium Porphyromonas gingivalis induced local DNA methylation and increased the level of histone acetylation in host cells. These epigenetic changes at the biofilm-gingiva interface may contribute to the development of periodontitis. Epigenetic regulators produced by intracellular bacteria alter the epigenotype and gene expression pattern of host cells and play an important role in pathogenesis.

Destabilization of Oil-in-Water Emulsions Formed Using Highly Hydrolyzed Whey Proteins.

Science.gov (United States)

Agboola; Singh; Munro; Dalgleish; Singh

1998-01-19

Oil-in-water emulsions (4 wt % soy oil) were prepared with 0.5-5 wt % whey protein hydrolysate (WPH) (27% degree of hydrolysis), in a two-stage homogenizer using various first-stage pressures of 10.3, 20.6, and 34.3 MPa and a constant second-stage pressure of 3.4 MPa. Destabilization studies on the emulsions were carried out for up to 24 h, using both laser light scattering and confocal laser microscopy. It was found that emulsions formed with oiling off and coalescence at all homogenization pressures. Emulsions formed with 2, 3, and 4% WPH showed coalescence and creaming only, while slight flocculation but no creaming occurred in emulsions formed with 5% WPH. Furthermore, the apparent rate of coalescence increased with homogenization pressure but decreased with WPH concentration. In contrast, the surface concentration of WPH increased with the WPH concentration in the emulsions but decreased with homogenization pressure. Analysis of WPH by high-performance liquid chromatography showed an increase in the concentration of high molecular weight peptides at the droplet surface compared to the WPH solution. This was considered very important for the stability of these oil-in-water emulsions.

Glutathione provides a source of cysteine essential for intracellular multiplication of Francisella tularensis.

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Khaled Alkhuder

2009-01-01

Full Text Available Francisella tularensis is a highly infectious bacterium causing the zoonotic disease tularemia. Its ability to multiply and survive in macrophages is critical for its virulence. By screening a bank of HimarFT transposon mutants of the F. tularensis live vaccine strain (LVS to isolate intracellular growth-deficient mutants, we selected one mutant in a gene encoding a putative gamma-glutamyl transpeptidase (GGT. This gene (FTL_0766 was hence designated ggt. The mutant strain showed impaired intracellular multiplication and was strongly attenuated for virulence in mice. Here we present evidence that the GGT activity of F. tularensis allows utilization of glutathione (GSH, gamma-glutamyl-cysteinyl-glycine and gamma-glutamyl-cysteine dipeptide as cysteine sources to ensure intracellular growth. This is the first demonstration of the essential role of a nutrient acquisition system in the intracellular multiplication of F. tularensis. GSH is the most abundant source of cysteine in the host cytosol. Thus, the capacity this intracellular bacterial pathogen has evolved to utilize the available GSH, as a source of cysteine in the host cytosol, constitutes a paradigm of bacteria-host adaptation.

Delivery of rifampicin-chitin nanoparticles into the intracellular compartment of polymorphonuclear leukocytes.

Science.gov (United States)

Smitha, K T; Nisha, N; Maya, S; Biswas, Raja; Jayakumar, R

2015-03-01

Polymorphonuclear leukocytes (PMNs) provide the primary host defence against invading pathogens by producing reactive oxygen species (ROS) and microbicidal products. However, few pathogens can survive for a prolonged period of time within the PMNs. Additionally their intracellular lifestyle within the PMNs protect themselves from the additional lethal action of host immune systems such as antibodies and complements. Antibiotic delivery into the intracellular compartments of PMNs is a major challenge in the field of infectious diseases. In order to deliver antibiotics within the PMNs and for the better treatment of intracellular bacterial infections we synthesized rifampicin (RIF) loaded amorphous chitin nanoparticles (RIF-ACNPs) of 350±50 nm in diameter. RIF-ACNPs nanoparticles are found to be non-hemolytic and non-toxic against a variety of host cells. The release of rifampicin from the prepared nanoparticles was ∼60% in 24 h, followed by a sustained pattern till 72 h. The RIF-ACNPs nanoparticles showed 5-6 fold enhanced delivery of RIF into the intracellular compartments of PMNs. The RIF-ACNPs showed anti-microbial activity against Escherichia coli, Staphylococcus aureus and a variety of other bacteria. In summary, our results suggest that RIF-ACNPs could be used to treat a variety of intracellular bacterial infections. Copyright © 2014 Elsevier B.V. All rights reserved.

Intracellular phase for an extracellular bacterial pathogen: MgtC shows the way

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Audrey Bernut

2015-08-01

Full Text Available Pseudomonas aeruginosa is an extracellular pathogen known to impair host phagocytic functions. However, our recent results identify MgtC as a novel actor in P. aeruginosa virulence, which plays a role in an intramacrophage phase of this pathogen. In agreement with its intracellular function, P. aeruginosa mgtC gene expression is strongly induced when the bacteria reside within macrophages. MgtC was previously known as a horizontally-acquired virulence factor important for multiplication inside macrophages in several intracellular bacterial pathogens. MgtC thus provides a singular example of a virulence determinant that subverts macrophages both in intracellular and extracellular pathogens. Moreover, we demonstrate that P. aeru-ginosa MgtC is required for optimal growth in Mg2+ deprived medium, a property shared by MgtC factors from intracellular pathogens and, under Mg2+ limitation, P. aeruginosaMgtC prevents biofilm formation. We propose that MgtC has a similar function in intracellular and extracellular pathogens, which contributes to macrophage resistance and fine-tune adaptation to the host in relation to the different bacterial lifestyles. MgtC thus appears as an attractive target for antivirulence strategies and our work provides a natural peptide as MgtC antagonist, which paves the way for the development of MgtC inhibitors.

Destabilization of the 6H-SrIrO3 polymorph through partial substitution of zinc and lithium

DEFF Research Database (Denmark)

Bremholm, Martin; K. Kim, Cindi; Hirai, Daigo

2012-01-01

We report on the destabilization of the 6H-SrIrO3 polymorph through partial substitutions of zinc and lithium for iridium to form perovskites. The perovskites crystallize in the orthorhombic space group Pbnm: SrIr1−xZnxO3 is found for 0.25 ≤ x ≤ 0.33, while SrIr1−xLixO3 is found only for x = 0...... show Curie–Weiss behavior, with relatively large temperature independent contributions, and that the iridium atoms have low effective moments, 0.52 to 1.08 μB per Ir. The resistivity of SrIr0.67Zn0.33O3, characterized by Mott variable range hopping type semiconducting behavior, indicates...

Model Linking Plasma and Intracellular Tenofovir/Emtricitabine with Deoxynucleoside Triphosphates.

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Xinhui Chen

Full Text Available The coformulation of the nucleos(tide analogs (NA tenofovir (TFV disoproxil fumarate (TDF and emtricitabine (FTC is approved for HIV-infection treatment and prevention. Plasma TFV and FTC undergo complicated hybrid processes to form, accumulate, and retain as their active intracellular anabolites: TFV-diphosphate (TFV-DP and FTC-triphosphate (FTC-TP. Such complexities manifest in nonlinear intracellular pharmacokinetics (PK. In target cells, TFV-DP/FTC-TP compete with endogenous deoxynucleoside triphosphates (dNTP at the active site of HIV reverse transcriptase, underscoring the importance of analog:dNTP ratios for antiviral efficacy. However, NA such as TFV and FTC have the potential to disturb the dNTP pool, which could augment or reduce their efficacies. We conducted a pharmacokinetics-pharmacodynamics (PKPD study among forty subjects receiving daily TDF/FTC (300 mg/200 mg from the first-dose to pharmacological intracellular steady-state (30 days. TFV/FTC in plasma, TFV-DP/FTC-TP and dNTPs in peripheral blood mononuclear cells (PBMC were quantified using validated LC/MS/MS methodologies. Concentration-time data were analyzed using nonlinear mixed effects modeling (NONMEM. Formations and the accumulation of intracellular TFV-DP/FTC-TP was driven by plasma TFV/FTC, which was described by a hybrid of first-order formation and saturation. An indirect response link model described the interplay between TFV-DP/FTC-TP and the dNTP pool change. The EC50 (interindividual variability, (%CV of TFV-DP and FTC-TP on the inhibition of deoxyadenosine triphosphate (dATP and deoxycytidine triphosphate (dCTP production were 1020 fmol/106 cells (130% and 44.4 pmol/106 cells (82.5%, resulting in (90% prediction interval 11% (0.45%, 53% and 14% (2.6%, 35% reductions. Model simulations of analog:dNTP molar ratios using IPERGAY dosing suggested that FTC significantly contributes to the protective effect of preexposure prophylaxis (PrEP. Simulation

Molecular detection and characterization of sustainable intracellular ...

African Journals Online (AJOL)

3Centre for Biopolymer and Bio-Molecular Research, Athlone College of Technology, Republic of Ireland. ... cells was associated with the elongation of micro-villar extension that ... Keywords: Intracellular contaminants, cell cultures, bacteria culture, pre-clinical studies. ... production work involving culture technology.

Cationic polymers for intracellular delivery of proteins

NARCIS (Netherlands)

Coué, G.M.J.P.C.; Engbersen, Johannes F.J.; Samal, Sangram; Dubruel, Peter

2015-01-01

Many therapeutic proteins exert their pharmaceutical action inside the cytoplasm or onto individual organelles inside the cell. Intracellular protein delivery is considered to be the most direct, fastest and safest approach for curing gene-deficiency diseases, enhancing vaccination and triggering

Biological synthesis and characterization of intracellular gold ...

Indian Academy of Sciences (India)

thods of reduction of metal ions using plants or microorganisms are often ... have several advantages over bacteria, they are often pre- ferred. ... in static condition for a period of 7 days. ... work was focused on the production of intracellular gold.

The effect of sodium bicarbonate on intracellular pH using {sup 31}P-MR spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Nakashima, Kazuya; Kashiwagi, Shiro; Ito, Haruhide [Yamaguchi Univ., Ube (Japan). School of Medicine; Yamashita, Tetsuo; Kitahara, Tetsuhiro; Nakayama, Naoto; Saito, Kennichi

1997-03-01

This report deals with the effects of sodium bicarbonate on the intracellular pH of the brain and cerebral blood flow (CBF); five normal volunteers were studied. Intracellular pH and CBF were measured by phosphorus 31 magnetic resonance spectroscopy ({sup 31}P-MRS) and stable xenon computed tomography (Xe-CT), respectively. Each individual received 7% sodium bicarbonate (3.5 ml/kg body weight), infused intravenously over a 15-min period. Intracellular pH, CBF, and physiological parameters were determined before and after the injection. Intracellular pH was significantly decreased and CBF was increased. Among the physiological parameters, the hematocrit was significantly decreased and arterial pressure of carbon dioxide (PaCO{sub 2}), increased. These results suggest that increasing CO{sub 2} contributes to the decrease in intracellular pH. In conclusion, three factors increase CBF during the administration of sodium bicarbonate to humans: arterial dilatation in response to carbon dioxide; decrease of the hematocrit, and intracellular cerebral acidosis. (author)

Intracellular insulin processing is altered in monocytes from patients with type II diabetes mellitus

International Nuclear Information System (INIS)

Trischitta, V.; Benzi, L.; Brunetti, A.; Cecchetti, P.; Marchetti, P.; Vigneri, R.; Navalesi, R.

1987-01-01

We studied total cell-associated A14-[ 125 I]insulin radioactivity (including surface-bound and internalized radioactivity), insulin internalization, and its intracellular degradation at 37 C in monocytes from nonobese type II untreated diabetic patients (n = 9) and normal subjects (n = 7). Total cell-associated radioactivity was decreased in diabetic patients [2.65 +/- 1.21% (+/- SD) vs. 4.47 +/- 1.04% of total radioactivity. Insulin internalization was also reduced in diabetic patients (34.0 +/- 6.8% vs. 59.0 +/- 11.3% of cell-associated radioactivity. Using high performance liquid chromatography six intracellular forms of radioactivity derived from A14-[ 125 I] insulin were identified; 10-20% of intracellular radioactivity had approximately 300,000 mol wt and was identified as radioactivity bound to the insulin receptor, and the remaining intracellular radioactivity included intact A14-[ 125 I]insulin, [ 125 I]iodide, or [ 125 I]tyrosine, and three intermediate compounds. A progressive reduction of intact insulin and a corresponding increase in iodine were found when the incubation time was prolonged. Intracellular insulin degradation was reduced in monocytes from diabetic patients; intracellular intact insulin was 65.6 +/- 18.1% vs. 37.4 +/- 18.0% of intracellular radioactivity after 2 min and 23.6 +/- 22.3% vs. 3.9 +/- 2.3% after 60 min in diabetic patients vs. normal subjects, respectively. In conclusion, 1) human monocytes internalize and degrade insulin in the intracellular compartment in a stepwise time-dependent manner; and 2) in monocytes from type II diabetic patients total cell-associated radioactivity, insulin internalization, and insulin degradation are significantly reduced. These defects may be related to the cellular insulin resistance present in these patients

Simple Recovery of Intracellular Gold Nanoparticles from Peanut Seedling Roots.

Science.gov (United States)

Raju, D; Mehta, Urmil J; Ahmad, Absar

2015-02-01

Fabrication of inorganic nanomaterials via a biological route witnesses the formation either extracellularly, intracellulary or both. Whereas extracellular formation of these nanomaterials is cherished owing to their easy and economical extraction and purification processes; the intracellular formation of nanomaterials, due to the lack of a proper recovery protocol has always been dreaded, as the extraction processes used so far were tedious, costly, time consuming and often resulting in very low recovery. The aim of the present study was to overcome the problems related with the extraction and recovery of intracellularly synthesized inorganic nanoparticles, and to devise a method to increasing the output, the shape, size, composition and dispersal of nanoparticles is not altered. Water proved to be much better system as it provided well dispersed, stable gold nanoparticles and higher recovery. This is the first report, where intracellular nanoparticles have been recovered using a very cost-effective and eco-friendly approach.

Intracellular Ca2+ Regulation in Calcium Sensitive Phenotype of Saccharomyces cerevisiae

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HERMANSYAH

2010-03-01

Full Text Available Intracellular cytosolic Ca2+ concentration accumulation plays an essential information in Saccharomyces cerevisiae i.e. to explain cellular mechanism of Ca2+ sensitive phenotype. Disruption both S. cerevisiae PPase PTP2 and MSG5 genes showed an inhibited growth in the presence of Ca2+. On the other hand, by using Luminocounter with apoaequorin system, a method based upon luminescent photoprotein aequorin, intracellular Ca2+ concentration was accumulated as a consequence of calcium sensitive phenotype of S. cerevisiae. This fact indicated that PPase ptp2Δ and msg5Δ were involved in intracellular Ca2+ transport in addition their already known pathways i.e Mitogen Activated Protein Kinase cell wall integrity pathway, high osmolarity glycerol (HOG pathway, and pheromone response FUS3 pathway.

Mycobacterium avium-intracellulare cellulitis occurring with septic arthritis after joint injection: a case report

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Murdoch David M

2007-02-01

Full Text Available Abstract Background Cellulitis caused by Mycobacterium avium-intracellulare has rarely been described. Mycobacterium avium-intracellulare is a rare cause of septic arthritis after intra-articular injection, though the causative role of injection is difficult to ascertain in such cases. Case presentation A 57-year-old with rheumatoid arthritis treated with prednisone and azathioprine developed bilateral painful degenerative shoulder arthritis. After corticosteroid injections into both acromioclavicular joints, he developed bilateral cellulitis centered over the injection sites. Skin biopsy showed non-caseating granulomas, and culture grew Mycobacterium avium-intracellulare. Joint aspiration also revealed Mycobacterium avium-intracellulare infection. Conclusion Although rare, skin and joint infections caused by Mycobacterium avium-intracellulare should be considered in any immunocompromised host, particularly after intra-articular injection. Stains for acid-fast bacilli may be negative in pathologic samples even in the presence of infection; cultures of tissue specimens should always be obtained.

Intracellular nitrate of marine diatoms as a driver of anaerobic nitrogen cycling in sinking aggregates

DEFF Research Database (Denmark)

Kamp, Anja; Stief, Peter; Bristow, Laura A.

2016-01-01

% was recovered as nitrite. Hence, aggregate-associated diatoms accumulate nitrate from the surrounding water and sustain complex nitrogen transformations, including loss of fixed nitrogen, in anoxic, pelagic microniches. Additionally, it may be expected that intracellular nitrate not converted before...... store nitrate intracellularly, we explored the fate of intracellular nitrate and its availability for microbial metabolism within anoxic diatom-bacteria aggregates. The ubiquitous nitrate-storing diatom Skeletonema marinoi was studied as both axenic cultures and laboratory-produced diatom......-bacteria aggregates. Stable 15N isotope incubations under dark and anoxic conditions revealed that axenic S. marinoi is able to reduce intracellular nitrate to ammonium that is immediately excreted by the cells. When exposed to a light:dark cycle and oxic conditions, S. marinoi stored nitrate intracellularly...

Intracellular Nitrate of Marine Diatoms as a Driver of Anaerobic Nitrogen Cycling in Sinking Aggregates

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Anja Kamp

2016-11-01

Full Text Available Diatom-bacteria aggregates are key for the vertical transport of organic carbon in the ocean. Sinking aggregates also represent pelagic microniches with intensified microbial activity, oxygen depletion in the center, and anaerobic nitrogen cycling. Since some of the aggregate-forming diatom species store nitrate intracellularly, we explored the fate of intracellular nitrate and its availability for microbial metabolism within anoxic diatom-bacteria aggregates. The ubiquitous nitrate-storing diatom Skeletonema marinoi was studied as both axenic cultures and laboratory-produced diatom-bacteria aggregates. Stable 15N isotope incubations under dark and anoxic conditions revealed that axenic S. marinoi is able to reduce intracellular nitrate to ammonium that is immediately excreted by the cells. When exposed to a light:dark cycle and oxic conditions, S. marinoi stored nitrate intracellularly in concentrations > 60 mmol L-1 both as free-living cells and associated to aggregates. Intracellular nitrate concentrations exceeded extracellular concentrations by three orders of magnitude. Intracellular nitrate was used up within 2-3 days after shifting diatom-bacteria aggregates to dark and anoxic conditions. Thirty-one percent of the diatom-derived nitrate was converted to nitrogen gas, indicating that a substantial fraction of the intracellular nitrate pool of S. marinoi becomes available to the aggregate-associated bacterial community. Only 5% of the intracellular nitrate was reduced to ammonium, while 59% was recovered as nitrite. Hence, aggregate-associated diatoms accumulate nitrate from the surrounding water and sustain complex nitrogen transformations, including loss of fixed nitrogen, in anoxic, pelagic microniches. Additionally, it may be expected that intracellular nitrate not converted before the aggregates have settled onto the seafloor could fuel benthic nitrogen transformations.

Flight Results of the NF-15B Intelligent Flight Control System (IFCS) Aircraft with Adaptation to a Longitudinally Destabilized Plant

Science.gov (United States)

Bosworth, John T.

2008-01-01

Adaptive flight control systems have the potential to be resilient to extreme changes in airplane behavior. Extreme changes could be a result of a system failure or of damage to the airplane. The goal for the adaptive system is to provide an increase in survivability in the event that these extreme changes occur. A direct adaptive neural-network-based flight control system was developed for the National Aeronautics and Space Administration NF-15B Intelligent Flight Control System airplane. The adaptive element was incorporated into a dynamic inversion controller with explicit reference model-following. As a test the system was subjected to an abrupt change in plant stability simulating a destabilizing failure. Flight evaluations were performed with and without neural network adaptation. The results of these flight tests are presented. Comparison with simulation predictions and analysis of the performance of the adaptation system are discussed. The performance of the adaptation system is assessed in terms of its ability to stabilize the vehicle and reestablish good onboard reference model-following. Flight evaluation with the simulated destabilizing failure and adaptation engaged showed improvement in the vehicle stability margins. The convergent properties of this initial system warrant additional improvement since continued maneuvering caused continued adaptation change. Compared to the non-adaptive system the adaptive system provided better closed-loop behavior with improved matching of the onboard reference model. A detailed discussion of the flight results is presented.

CONTRIBUTIONS OF INTRACELLULAR IONS TO Kv CHANNEL VOLTAGE SENSOR DYNAMICS.

Directory of Open Access Journals (Sweden)

Samuel eGoodchild

2012-06-01

Full Text Available Voltage sensing domains of Kv channels control ionic conductance through coupling of the movement of charged residues in the S4 segment to conformational changes at the cytoplasmic region of the pore domain, that allow K+ ions to flow. Conformational transitions within the voltage sensing domain caused by changes in the applied voltage across the membrane field are coupled to the conducting pore region and the gating of ionic conductance. However, several other factors not directly linked to the voltage dependent movement of charged residues within the voltage sensor impact the dynamics of the voltage sensor, such as inactivation, ionic conductance, intracellular ion identity and block of the channel by intracellular ligands. The effect of intracellular ions on voltage sensor dynamics is of importance in the interpretation of gating current measurements and the physiology of pore/voltage sensor coupling. There is a significant amount of variability in the reported kinetics of voltage sensor deactivation kinetics of Kv channels attributed to different mechanisms such as open state stabilization, immobilization and relaxation processes of the voltage sensor. Here we separate these factors and focus on the causal role that intracellular ions can play in allosterically modulating the dynamics of Kv voltage sensor deactivation kinetics. These considerations are of critical importance in understanding the molecular determinants of the complete channel gating cycle from activation to deactivation.

Killing of intracellular Mycobacterium tuberculosis by receptor-mediated drug delivery

International Nuclear Information System (INIS)

Majumdar, S.; Basu, S.K.

1991-01-01

p-Aminosalicylic acid (PAS) conjugated to maleylated bovine serum albumin (MBSA) was taken up efficiently through high-affinity MBSA-binding sites on macrophages. Binding of the radiolabeled conjugate to cultured mouse peritoneal macrophages at 4 degrees C was competed for by MBSA but not by PAS. At 37 degrees C, the radiolabeled conjugate was rapidly degraded by the macrophages, leading to release of acid-soluble degradation products in the medium. The drug conjugate was nearly 100 times as effective as free PAS in killing the intracellular mycobacteria in mouse peritoneal macrophages infected in culture with Mycobacterium tuberculosis. The killing of intracellular mycobacteria mediated by the drug conjugate was effectively prevented by simultaneous addition of excess MBSA (100 micrograms/ml) or chloroquine (3 microM) to the medium, whereas these agents did not affect the microbicidal action of free PAS. These results suggest that (i) uptake of the PAS-MBSA conjugate was mediated by cell surface receptors on macrophages which recognize MBSA and (ii) lysosomal hydrolysis of the internalized conjugate resulted in intracellular release of a pharmacologically active form of the drug, which led to selective killing of the M. tuberculosis harbored by mouse macrophages infected in culture. This receptor-mediated modality of delivering drugs to macrophages could contribute to greater therapeutic efficacy and minimization of toxic side effects in the management of tuberculosis and other intracellular mycobacterial infections

Complexation of intracellular cyanide by hydroxocobalamin using a human cellular model.

Science.gov (United States)

Astier, A; Baud, F J

1996-01-01

1. The rational for administering hydroxocobalamin (OHCbl) as an antidote to cyanide poisoning is based on the high affinity of CN ion for cobalt compounds. However, only few data are available on the influence of OHCbl on the intracellular cyanide pool. 2. In human fibroblasts incubated for 10 min with 500 microM of [14C] cyanide, the accumulation ratio was 25 at 37 degrees C (10.45 +/- 1.51 mM) and 11.9 at 4 degrees C. 3. Using the monoblastic U-937 cell line, a rapid uptake of radioactive cyanide was observed with a maximum accumulation ratio of 1.97 at 5 min. 4. A linear relationship between cyanide uptake by U-937 cells and cyanide concentration in incubation medium (10-500 microM; 5 min) was found suggesting a first order process (k = 0.25 min-1). 5. After incubation of fibroblasts with 500 microM of OHCbl, a 75% decrease of intracellular cyanide was observed, with concomittant formation of intracellular cyanocobalamin CNCbl (intracellular/extracellular ratio: 158). 6. These findings suggest that OHCbl is able to penetrate into heavily cyanide loaded cells and to complex cyanide to the non-toxic CNCbl form.

Dynamics of gradient formation by intracellular shuttling

Energy Technology Data Exchange (ETDEWEB)

Berezhkovskii, Alexander M. [Mathematical and Statistical Computing Laboratory, Division of Computational Bioscience, Center for Information Technology, National Institutes of Health, Bethesda, Maryland 20892 (United States); Shvartsman, Stanislav Y. [Department of Chemical and Biological Engineering and Lewis-Sigler Institute for Integrative Genomics, Princeton University, Princeton, New Jersey 08544 (United States)

2015-08-21

A number of important cellular functions rely on the formation of intracellular protein concentration gradients. Experimental studies discovered a number of mechanisms for the formation of such gradients. One of the mechanisms relies on the intracellular shuttling of a protein that interconverts between the two states with different diffusivities, under the action of two enzymes, one of which is localized to the plasma membrane, whereas the second is uniformly distributed in the cytoplasm. Recent work reported an analytical solution for the steady state gradient in this mechanism, obtained in the framework of a one-dimensional reaction-diffusion model. Here, we study the dynamics in this model and derive analytical expressions for the Laplace transforms of the time-dependent concentration profiles in terms of elementary transcendental functions. Inverting these transforms numerically, one can obtain time-dependent concentration profiles of the two forms of the protein.

Lithosphere destabilization by melt percolation during pre-oceanic rifting: Evidence from Alpine-Apennine ophiolitic peridotites

Science.gov (United States)

Piccardo, Giovanni; Ranalli, Giorgio

2017-04-01

conditions. This indicates that thermal advection by percolation of hot asthenospheric melts significantly heated the lithospheric mantle column above the melting asthenosphere. Numerical and analogue models show that infiltration of melts results in considerable softening of mantle rocks. Total ithospheric strength can be decreased from 10 to 1 TN m-1 as orders of magnitude and the sin-rift thermo-mechanical erosion of the lithospheric mantle induces significant rheological softening along the axial zone of extension (Corti et al., 2007; Ranalli et al., 2007). Softening of the lithospheric mantle may lead to whole lithospheric failure and consequently to transition from continental extension to oceanic spreading. Therefore, rheological softening caused destabilization of the lithospheric mantle between the future continental margins (Piccardo et al., 2014; Piccardo, 2016) of the Ligurian Tethys. The wedge of destabilized lithosphere favored faster divergence of the continental blocks and enhanced doming and thermal buoyancy of deeper/hotter asthenosphere that rose between the future continental margins and originated aggregated MORB melts (i.e., the oceanic magmatism that formed olivine-gabbro intrusions and pillowed basalt extrusions). Lithosphere destabilization by melt percolation can play a fundamental role in the geodynamic evolution of lithosphere extension causing transition from continental extension to continental break-up to oceanic spreading. Corti, G., Bonini, M., Innocenti, F., Manetti, P., Piccardo, G.B., Ranalli, G., 2007. Journal of Geodynamics, 43, 465-483. Piccardo, G.B., Padovano, M., Guarnieri, L. 2014. Earth-Science Reviews, 138, 409-434. Piccardo, G.B., 2016. Gondwana Research, 39, 230-249. Piccardo, G.B., Vissers, R.L.M., 2007. Journal of Geodynamics, 43, 417-449. Piccardo, G.B., Guarnieri, L., 2011. Lithos, 124, 210-214. Ranalli, G., Piccardo, G.B., Corona-Chavez, P., 2007. Journal of Geodynamics, 43, 450-464.

Electron Microscopy of Intracellular Protozoa

Science.gov (United States)

1988-12-20

Classification) " ELECTRON MICROSCOPY OF INTRACELLULAR PROTOZOA 12. PERSONAL AUTHOR(S) Aikawa, Masamichi 13a. TYPE OF REPORT I13b. TIME COVERED 114...authors suggest that anti-CS protein antibody is important in reducing the prevalence of malaria with increasing age among persons in such areas and... Hygine 33, 220-226. 0Giudice, G.D., Engers, H.D., Tougne, C., Biro, S.S., Weiss, N., Verdini, A.S., Pessi, A., Degremont, A.A., Freyvogel, T.A., Lambert

Emerging Paradigm of Intracellular Targeting of G Protein-Coupled Receptors.

Science.gov (United States)

Chaturvedi, Madhu; Schilling, Justin; Beautrait, Alexandre; Bouvier, Michel; Benovic, Jeffrey L; Shukla, Arun K

2018-05-04

G protein-coupled receptors (GPCRs) recognize a diverse array of extracellular stimuli, and they mediate a broad repertoire of signaling events involved in human physiology. Although the major effort on targeting GPCRs has typically been focused on their extracellular surface, a series of recent developments now unfold the possibility of targeting them from the intracellular side as well. Allosteric modulators binding to the cytoplasmic surface of GPCRs have now been described, and their structural mechanisms are elucidated by high-resolution crystal structures. Furthermore, pepducins, aptamers, and intrabodies targeting the intracellular face of GPCRs have also been successfully utilized to modulate receptor signaling. Moreover, small molecule compounds, aptamers, and synthetic intrabodies targeting β-arrestins have also been discovered to modulate GPCR endocytosis and signaling. Here, we discuss the emerging paradigm of intracellular targeting of GPCRs, and outline the current challenges, potential opportunities, and future outlook in this particular area of GPCR biology. Copyright © 2018 Elsevier Ltd. All rights reserved.

The effect of high pressure on the intracellular trehalose synthase activity of Thermus aquaticus.

Science.gov (United States)

Dong, Yongsheng; Ma, Lei; Duan, Yuanliang

2016-01-01

To understand the effect of high pressure on the intracellular trehalose synthase activity, Thermus aquaticus (T. aquaticus) in the logarithmic growth phase was treated with high-pressure air, and its intracellular trehalose synthase (TSase) activity was determined. Our results indicated that pressure is a factor strongly affecting the cell growth. High pressure significantly attenuated the growth rate of T. aquaticus and shortened the duration of stationary phase. However, after 2 h of culture under 1.0 MPa pressure, the activity of intracellular TSase in T. aquaticus reached its maximum value, indicating that pressure can significantly increase the activity of intracellular TSase in T. aquaticus. Thus the present study provides an important guide for the enzymatic production of trehalose.

Intrinsic Toxin-Derived Peptides Destabilize and Inactivate Clostridium difficile TcdB

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Jason L. Larabee

2017-05-01

Full Text Available Clostridium difficile infection (CDI is a major cause of hospital-associated, antibiotic-induced diarrhea, which is largely mediated by the production of two large multidomain clostridial toxins, TcdA and TcdB. Both toxins coordinate the action of specific domains to bind receptors, enter cells, and deliver a catalytic fragment into the cytosol. This results in GTPase inactivation, actin disassembly, and cytotoxicity. TcdB in particular has been shown to encode a region covering amino acids 1753 to 1851 that affects epitope exposure and cytotoxicity. Surprisingly, studies here show that several peptides derived from this region, which share the consensus sequence 1769NVFKGNTISDK1779, protect cells from the action of TcdB. One peptide, PepB2, forms multiple interactions with the carboxy-terminal region of TcdB, destabilizes TcdB structure, and disrupts cell binding. We further show that these effects require PepB2 to form a higher-order polymeric complex, a process that requires the central GN amino acid pair. These data suggest that TcdB1769–1779 interacts with repeat sequences in the proximal carboxy-terminal domain of TcdB (i.e., the CROP domain to alter the conformation of TcdB. Furthermore, these studies provide insights into TcdB structure and functions that can be exploited to inactivate this critical virulence factor and ameliorate the course of CDI.

Intracellular diffusion restrictions in isolated cardiomyocytes from rainbow trout

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Birkedal Rikke

2009-12-01

Full Text Available Abstract Background Restriction of intracellular diffusion of adenine nucleotides has been studied intensively on adult rat cardiomyocytes. However, their cause and role in vivo is still uncertain. Intracellular membrane structures have been suggested to play a role. We therefore chose to study cardiomyocytes from rainbow trout (Oncorhynchus mykiss, which are thinner and have fewer intracellular membrane structures than adult rat cardiomyocytes. Previous studies suggest that trout permeabilized cardiac fibers also have diffusion restrictions. However, results from fibers may be affected by incomplete separation of the cells. This is avoided when studying permeabilized, isolated cardiomyocytes. The aim of this study was to verify the existence of diffusion restrictions in trout cardiomyocytes by comparing ADP-kinetics of mitochondrial respiration in permeabilized fibers, permeabilized cardiomyocytes and isolated mitochondria from rainbow trout heart. Experiments were performed at 10, 15 and 20°C in the absence and presence of creatine. Results Trout cardiomyocytes hypercontracted in the solutions used for mammalian cardiomyocytes. We developed a new solution in which they retained their shape and showed stable steady state respiration rates throughout an experiment. The apparent ADP-affinity of permeabilized cardiomyocytes was different from that of fibers. It was higher, independent of temperature and not increased by creatine. However, it was still about ten times lower than in isolated mitochondria. Conclusions The differences between fibers and cardiomyocytes suggest that results from trout heart fibers were affected by incomplete separation of the cells. However, the lower ADP-affinity of cardiomyocytes compared to isolated mitochondria indicate that intracellular diffusion restrictions are still present in trout cardiomyocytes despite their lower density of intracellular membrane structures. The lack of a creatine effect indicates that

Uncoupling PIP2-calmodulin regulation of Kv7.2 channels by an assembly destabilizing epileptogenic mutation.

Science.gov (United States)

Alberdi, Araitz; Gomis-Perez, Carolina; Bernardo-Seisdedos, Ganeko; Alaimo, Alessandro; Malo, Covadonga; Aldaregia, Juncal; Lopez-Robles, Carlos; Areso, Pilar; Butz, Elisabeth; Wahl-Schott, Christian; Villarroel, Alvaro

2015-11-01

We show that the combination of an intracellular bi-partite calmodulin (CaM)-binding site and a distant assembly region affect how an ion channel is regulated by a membrane lipid. Our data reveal that regulation by phosphatidylinositol(4,5)bisphosphate (PIP2) and stabilization of assembled Kv7.2 subunits by intracellular coiled-coil regions far from the membrane are coupled molecular processes. Live-cell fluorescence energy transfer measurements and direct binding studies indicate that remote coiled-coil formation creates conditions for different CaM interaction modes, each conferring different PIP2 dependency to Kv7.2 channels. Disruption of coiled-coil formation by epilepsy-causing mutation decreases apparent CaM-binding affinity and interrupts CaM influence on PIP2 sensitivity. © 2015. Published by The Company of Biologists Ltd.

The surface destabilization effect of nitrate on the calcite (104). Water interface and yttrium(III) sorption thereon

Energy Technology Data Exchange (ETDEWEB)

Hellebrandt, S.E.; Hofmann, Sascha; Schmidt, Moritz [Helmholtz-Zentrum Dresden-Rossendorf e.V., Dresden (Germany). Div. Surface Processes; Stubbs, J.E.; Eng, P.J. [Chicago Univ., IL (United States). Center for Advanced Radiation Sources; Stumpf, Thorsten [Helmholtz-Zentrum Dresden-Rossendorf e.V., Dresden (Germany). Inst. of Resource Ecology

2016-07-01

Calcite, as a most abundant mineral on earth, was studied with X-ray reflectivity under the influence of NaNO{sub 3} [1]. The calcite (104) surface undergoes significant destabilization effects in the presence of NaNO{sub 3}, which occurs as partial dissolution and the formation of an amorphous layer at the interface. The disordering of the surface reaches more than 15 Aa into the crystal bulk. Furthermore, this surface modification has also an effect on the sorption behavior of the rare earth element Y. Without NaNO{sub 3} Y{sup 3+} adsorbs as both inner and outer sphere complexes, this was verified with resonant anomalous X-ray reflectivity (RAXR). If NaNO{sub 3} is present, both species desorbs from the surface completely.

DMPD: Ubiquitin: tool and target for intracellular NF-kappaB inhibitors. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 16982211 Ubiquitin: tool and target for intracellular NF-kappaB inhibitors. Wullaer...vg) (.html) (.csml) Show Ubiquitin: tool and target for intracellular NF-kappaB inhibitors. PubmedID 1698221...1 Title Ubiquitin: tool and target for intracellular NF-kappaB inhibitors. Author

Intracellular pH and inorganic phosphate content of heart in vivo: A 31P-NMR study

International Nuclear Information System (INIS)

Katz, L.A.; Swain, J.A.; Portman, M.A.; Balaban, R.S.

1988-01-01

Studies were performed to determine the contribution of red blood cells to the 31 P-nuclear magnetic resonance (NMR) spectrum of the canine heart in vivo and the feasibility of measuring myocardial intracellular phosphate and pH. This was accomplished by replacing whole blood with a perfluorochemical perfusion emulsion blood substitute, Oxypherol, and noting the difference in the 31 P-NMR spectrum of the heart. NMR data were collected with a NMR transmitter-receiver coil on the surface of the distal portion of the left ventricle. These studies demonstrated that a small contribution from 2,3-diphosphoglycerate (2,3-DPG) and phosphodiesters in the blood could be detected. The magnitude and shift of these blood-borne signals permitted the relative quantification of intracellular inorganic phosphate (P i ) content as well as intracellular pH. Under resting conditions, the intracellular ATP/P i was 7.0 ± 0.08. This corresponds to a free intracellular P 1 content of ∼ 0.8 μmol./g wet wt. The intracellular pH was 7.10 ± 0.01. Acute respiratory alkalosis and acidosis, with the arterial pH ranging from ∼7.0 to 7.7, resulted in only small changes in the intracellular pH. These latter results demonstrate an effective myocardial intracellular proton-buffering mechanism in vivo

Chelation of intracellular calcium blocks insulin action in the adipocyte

International Nuclear Information System (INIS)

Pershadsingh, H.A.; Shade, D.L.; Delfert, D.M.; McDonald, J.M.

1987-01-01

The hypothesis that intracellular Ca 2+ is an essential component of the intracellular mechanism of insulin action in the adipocyte was evaluated. Cells were loaded with the Ca 2+ chelator quin-2, by preincubating them with quin-2 AM, the tetrakis(acetoxymethyl) ester of quin-2. Quin-2 loading inhibited insulin-stimulated glucose transport without affecting basal activity. The ability of insulin to stimulate glucose uptake in quin-2-loaded cells could be partially restored by preincubating cells with buffer supplemented with 1.2 mM CaCl 2 and the Ca 2+ ionophore A23187. These conditions had no effect on basal activity and omission of CaCl 2 from the buffer prevented the restoration of insulin-stimulated glucose uptake by A23187. Quin-2 loading also inhibited insulin-stimulated glucose oxidation and the ability of insulin to inhibit cAMP-stimulated lipolysis without affecting their basal activities. Incubation of cells with 100 μM quin-2 or quin-2 AM had no effect on intracellular ATP concentration or the specific binding of 125 I=labeled insulin to adipocytes. These findings suggest that intracellular Ca 2+ is an essential component in the coupling of the insulin-activated receptor complex to cellular physiological/metabolic machinery. Furthermore, differing quin-2 AM dose-response profiles suggest the presence of dual Ca 2+ -dependent pathways in the adipocyte. One involves insulin stimulation of glucose transport and oxidation, whereas the other involves the antilipolytic action of insulin

Effect of insulin resistance on intracellular signal transduction of vessels in diabetic

International Nuclear Information System (INIS)

Cen Rongguang; Wei Shaoying; Mo Xingju

2003-01-01

To investigate the relationship between the insulin resistance (IR) and the intracellular signal transduction of vessels, changes in fasting blood glucose (FBG), fasting insulin (FINS), triglyceride (TG), total cholesterol (TC), inositol triphosphate (IP 3 ), protein kinase C(PKC) and intracellular total calcium concentration in 31 diabetic patients were compared with those of 39 normal controls. The levels of FBG, FINS, TG and TC in diabetic patients were significantly higher than those of normal controls (P 3 and PKC in diabetic patients were significantly lower than those of normal controls (P<0.01). The results suggest that there is a causal relation between insulin resistance and abnormalities of cellular calcium metabolism and intracellular signal transduction of vessels

Intracellular delivery of poly(I:C) induces apoptosis of fibroblast-like synoviocytes via an unknown dsRNA sensor

Energy Technology Data Exchange (ETDEWEB)

Karpus, Olga N.; Hsiao, Cheng-Chih; Kort, Hanneke de; Tak, Paul P.; Hamann, Jörg, E-mail: j.hamann@amc.uva.nl

2016-08-26

Fibroblast-like synoviocytes (FLS) express functional membranous and cytoplasmic sensors for double-stranded (ds)RNA. Notably, FLS undergo apoptosis upon transfection with the synthetic dsRNA analog poly(I:C). We here studied the mechanism of intracellular poly(I:C) recognition and subsequent cell death in FLS. FLS responded similarly to poly(I:C) or 3pRNA transfection; however, only intracellular delivery of poly(I:C) induced significant cell death, accompanied by upregulation of pro-apoptotic proteins Puma and Noxa, caspase 3 cleavage, and nuclear segregation. Knockdown of the DExD/H-box helicase MDA5 did not affect the response to intracellular poly(I:C); in contrast, knockdown of RIG-I abrogated the response to 3pRNA. Knockdown of the downstream adaptor proteins IPS, STING, and TRIF or inhibition of TBK1 did not affect the response to intracellular poly(I:C), while knockdown of IFNAR blocked intracellular poly(I:C)-mediated signaling and cell death. We conclude that a so far unknown intracellular sensor recognizes linear dsRNA and induces apoptosis in FLS. - Highlights: • Intracellular poly(I:C) and 3pRNA evoke immune responses in FLS. • Only intracellular delivery of poly(I:C) induces FLS apoptosis. • FLS do not require MDA5 for their response to intracellular poly(I:C). • FLS respond to intracellular poly(I:C) independent of IPS and STING. • An unknown intracellular sensor recognizes linear dsRNA in FLS.

Intracellular pH gradients in migrating cells

DEFF Research Database (Denmark)

Martin, Christine; Pedersen, Stine Helene Falsig; Schwab, Albrecht

2011-01-01

might function as such unevenly distributed regulators as they modulate the interaction of focal adhesion proteins and components of the cytoskeleton in vitro. However, an intracellular pH (pH(i)) gradient reflecting a spatial asymmetry of protons has not been shown so far. One major regulator of p...

Biological synthesis and characterization of intracellular gold ...

Indian Academy of Sciences (India)

In the present study, Aspergillus fumigatus was used for the intracellular synthesis of gold nanoparticles. Stable nanoparticles were produced when an aqueous solution of chloroauric acid (HAuCl4) was reduced by A. fumigatus biomass as the reducing agent. Production of nanoparticles was confirmed by the colour ...

Intracellular calcium signal at the leading edge regulates mesodermal sheet migration during Xenopus gastrulation.

Science.gov (United States)

Hayashi, Kentaro; Yamamoto, Takamasa S; Ueno, Naoto

2018-02-05

During the gastrulation stage in animal embryogenesis, the cells leading the axial mesoderm migrate toward the anterior side of the embryo, vigorously extending cell protrusions such as lamellipodia. It is thought that the leading cells sense gradients of chemoattractants emanating from the ectodermal cells and translate them to initiate and maintain the cell movements necessary for gastrulation. However, it is unclear how the extracellular information is converted to the intracellular chemical reactions that lead to motion. Here we demonstrated that intracellular Ca 2+ levels in the protrusion-forming leading cells are markedly higher than those of the following cells and the axial mesoderm cells. We also showed that inhibiting the intracellular Ca 2+ significantly retarded the gastrulation cell movements, while increasing the intracellular Ca 2+ with an ionophore enhanced the migration. We further found that the ionophore treatment increased the active form of the small GTPase Rac1 in these cells. Our results suggest that transient intracellular Ca 2+ signals play an essential role in the active cell migration during gastrulation.

Pulsed magneto-motive ultrasound imaging to detect intracellular accumulation of magnetic nanoparticles

International Nuclear Information System (INIS)

Mehrmohammadi, Mohammad; Qu Min; Sokolov, Konstantin V; Emelianov, Stanislav Y; Ma, Li L; Johnston, Keith P; Romanovicz, Dwight K

2011-01-01

As applications of nanoparticles in medical imaging and biomedicine rapidly expand, the interactions of nanoparticles with living cells have become an area of active interest. For example, intracellular accumulation of nanoparticles-an important part of cell-nanoparticle interaction-has been well studied using plasmonic nanoparticles and optical or optics-based techniques due to the change in optical properties of the nanoparticle aggregates. However, magnetic nanoparticles, despite their wide range of clinical applications, do not exhibit plasmonic-resonant properties and therefore their intracellular aggregation cannot be detected by optics-based imaging techniques. In this study, we investigated the feasibility of a novel imaging technique-pulsed magneto-motive ultrasound (pMMUS)-to identify intracellular accumulation of endocytosed magnetic nanoparticles. In pMMUS imaging a focused, high intensity, pulsed magnetic field is used to excite the cells labeled with magnetic nanoparticles, and ultrasound imaging is then used to monitor the mechanical response of the tissue. We demonstrated previously that clusters of magnetic nanoparticles amplify the pMMUS signal in comparison to the signal from individual nanoparticles. Here we further demonstrate that pMMUS imaging can identify interaction between magnetic nanoparticles and living cells, i.e. intracellular accumulation of nanoparticles within the cells. The results of our study suggest that pMMUS imaging can not only detect the presence of magnetic nanoparticles but also provides information about their intracellular accumulation non-invasively and in real-time.

Destabilization of hydromagnetic drift-Alfven waves in a finite pressure collisional plasma

International Nuclear Information System (INIS)

Tang, J.T.

1974-01-01

In a finite beta (β = 8πn 0 kT 0 /B 0 2 ) plasma, where the plasma pressure n 0 kT 0 is an appreciable fraction of the confining magnetic field energy-density B 0 2 /8π, density-gradient driven drift waves couple with Alfven waves when the phase velocities of the two waves become comparable. The resulting hydromagnetic drift-Alfven waves separate into two branches--a drift mode and an Alfven mode, with both modes exhibiting magnetic field and localized density fluctuations near the coupling point. The dispersion relation of the collisional drift-Alfven wave is derived by using a slab-geometry, two-fluid model which includes finite beta, electron-ion collisions, ion-ion collisions, finite ion larmar radius, temperature fluctuations, and an axial electron current. A hydromagnetic drift mode is found to be unstable in a moderately dense plasma. A localized ''Alfven'' mode is destabilized only with the passage of an axial current along the plasma column. In order to check the theoretical predictions an experiment is performed in a finite-beta plasma of density n 0 = 10 13 -10 15 cm -3 and temperature T/sub e/ approximately T/sub i/ = 1-7 eV. (U.S.)

Brucella abortus nicotinamidase (PncA) contributes to its intracellular replication and infectivity in mice.

Science.gov (United States)

Kim, Suk; Kurokawa, Daisuke; Watanabe, Kenta; Makino, Sou-Ichi; Shirahata, Toshikazu; Watarai, Masahisa

2004-05-15

Brucella spp. are facultative intracellular pathogens that have the ability to survive and multiply in professional and non-professional phagocytes, and cause abortion in domestic animals and undulant fever in humans. The mechanism and factors of virulence are not fully understood. Nicotinamidase/pyrazinamidase mutant (pncA mutant) of Brucella abortus failed to replicate in HeLa cells, and showed a lower rate of intracellular replication than that of wild-type strain in macrophages. Addition of nicotinic acid, but not nicotinamide, into medium supported intracellular replication of pncA mutant in HeLa cells and macrophages. The pncA mutant was not co-localizing with either late endosomes or lysosomes. The B. abortus virB4 mutant was completely cleared from the spleens of mice after 4 weeks, while the pncA mutant showed a 1.5-log reduction of the number of bacteria isolated from spleens after 10 weeks. Although pncA mutant showed reduced virulence in mice and defective intracellular replication, its ability to confer protection against the virulent B. abortus strain 544 was fully retained. These results suggest that PncA does not contribute to intracellular trafficking of B. abortus, but contributes to utilization of nutrients required for intracellular growth. Our results indicate that detailed characterizations of the pncA mutant may help the improvement of currently available live vaccines. Copyright 2004 Federation of European Microbiological Societies

Getting “Inside” Type I IFNs: Type I IFNs in Intracellular Bacterial Infections

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Deann T. Snyder

2017-01-01

Full Text Available Type I interferons represent a unique and complex group of cytokines, serving many purposes during innate and adaptive immunity. Discovered in the context of viral infections, type I IFNs are now known to have myriad effects in infectious and autoimmune disease settings. Type I IFN signaling during bacterial infections is dependent on many factors including whether the infecting bacterium is intracellular or extracellular, as different signaling pathways are activated. As such, the repercussions of type I IFN induction can positively or negatively impact the disease outcome. This review focuses on type I IFN induction and downstream consequences during infection with the following intracellular bacteria: Chlamydia trachomatis, Listeria monocytogenes, Mycobacterium tuberculosis, Salmonella enterica serovar Typhimurium, Francisella tularensis, Brucella abortus, Legionella pneumophila, and Coxiella burnetii. Intracellular bacterial infections are unique because the bacteria must avoid, circumvent, and even co-opt microbial “sensing” mechanisms in order to reside and replicate within a host cell. Furthermore, life inside a host cell makes intracellular bacteria more difficult to target with antibiotics. Because type I IFNs are important immune effectors, modulating this pathway may improve disease outcomes. But first, it is critical to understand the context-dependent effects of the type I IFN pathway in intracellular bacterial infections.

Mycobacterium intracellulare Pleurisy Identified on Liquid Cultures of the Pleural Fluid and Pleural Biopsy.

Science.gov (United States)

Lim, Jong Gu; O, Sei Won; Lee, Ki Dong; Suk, Dong Keun; Jung, Tae Young; Shim, Tae Sun; Chon, Gyu Rak

2013-03-01

Pleural effusion is a rare complication in non-tuberculous mycobacterial infection. We report a case of Mycobacterium intracellulare pleuritis with idiopathic pulmonary fibrosis in a 69-year-old man presenting with dyspnea. Pleural effusion revealed lymphocyte dominant exudate. M. intracellulare was identified using a polymerase chain reaction-restriction fragment length polymorphism method and liquid cultures of pleural effusion and pleural biopsy. After combination therapy for M. intracellulare pulmonary disease, the patient was clinically well at a 1-month follow-up.

NAD+-Glycohydrolase Promotes Intracellular Survival of Group A Streptococcus.

Directory of Open Access Journals (Sweden)

Onkar Sharma

2016-03-01

Full Text Available A global increase in invasive infections due to group A Streptococcus (S. pyogenes or GAS has been observed since the 1980s, associated with emergence of a clonal group of strains of the M1T1 serotype. Among other virulence attributes, the M1T1 clone secretes NAD+-glycohydrolase (NADase. When GAS binds to epithelial cells in vitro, NADase is translocated into the cytosol in a process mediated by streptolysin O (SLO, and expression of these two toxins is associated with enhanced GAS intracellular survival. Because SLO is required for NADase translocation, it has been difficult to distinguish pathogenic effects of NADase from those of SLO. To resolve the effects of the two proteins, we made use of anthrax toxin as an alternative means to deliver NADase to host cells, independently of SLO. We developed a novel method for purification of enzymatically active NADase fused to an amino-terminal fragment of anthrax toxin lethal factor (LFn-NADase that exploits the avid, reversible binding of NADase to its endogenous inhibitor. LFn-NADase was translocated across a synthetic lipid bilayer in vitro in the presence of anthrax toxin protective antigen in a pH-dependent manner. Exposure of human oropharyngeal keratinocytes to LFn-NADase in the presence of protective antigen resulted in cytosolic delivery of NADase activity, inhibition of protein synthesis, and cell death, whereas a similar construct of an enzymatically inactive point mutant had no effect. Anthrax toxin-mediated delivery of NADase in an amount comparable to that observed during in vitro infection with live GAS rescued the defective intracellular survival of NADase-deficient GAS and increased the survival of SLO-deficient GAS. Confocal microscopy demonstrated that delivery of LFn-NADase prevented intracellular trafficking of NADase-deficient GAS to lysosomes. We conclude that NADase mediates cytotoxicity and promotes intracellular survival of GAS in host cells.

New intracellular activities of matrix metalloproteinases shine in the moonlight.

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Jobin, Parker G; Butler, Georgina S; Overall, Christopher M

2017-11-01

Adaption of a single protein to perform multiple independent functions facilitates functional plasticity of the proteome allowing a limited number of protein-coding genes to perform a multitude of cellular processes. Multifunctionality is achievable by post-translational modifications and by modulating subcellular localization. Matrix metalloproteinases (MMPs), classically viewed as degraders of the extracellular matrix (ECM) responsible for matrix protein turnover, are more recently recognized as regulators of a range of extracellular bioactive molecules including chemokines, cytokines, and their binders. However, growing evidence has convincingly identified select MMPs in intracellular compartments with unexpected physiological and pathological roles. Intracellular MMPs have both proteolytic and non-proteolytic functions, including signal transduction and transcription factor activity thereby challenging their traditional designation as extracellular proteases. This review highlights current knowledge of subcellular location and activity of these "moonlighting" MMPs. Intracellular roles herald a new era of MMP research, rejuvenating interest in targeting these proteases in therapeutic strategies. This article is part of a Special Issue entitled: Matrix Metalloproteinases edited by Rafael Fridman. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.

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Hannah Karlsson

Full Text Available CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs.

Promotion and Rescue of Intracellular Brucella neotomae Replication during Coinfection with Legionella pneumophila.

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Kang, Yoon-Suk; Kirby, James E

2017-05-01

We established a new Brucella neotomae in vitro model system for study of type IV secretion system-dependent (T4SS) pathogenesis in the Brucella genus. Importantly, B. neotomae is a rodent pathogen, and unlike B. abortus , B. melitensis , and B. suis , B. neotomae has not been observed to infect humans. It therefore can be handled more facilely using biosafety level 2 practices. More particularly, using a series of novel fluorescent protein and lux operon reporter systems to differentially label pathogens and track intracellular replication, we confirmed T4SS-dependent intracellular growth of B. neotomae in macrophage cell lines. Furthermore, B. neotomae exhibited early endosomal (LAMP-1) and late endoplasmic reticulum (calreticulin)-associated phagosome maturation. These findings recapitulate prior observations for human-pathogenic Brucella spp. In addition, during coinfection experiments with Legionella pneumophila , we found that defective intracellular replication of a B. neotomae T4SS virB4 mutant was rescued and baseline levels of intracellular replication of wild-type B. neotomae were significantly stimulated by coinfection with wild-type but not T4SS mutant L. pneumophila Using confocal microscopy, it was determined that intracellular colocalization of B. neotomae and L. pneumophila was required for rescue and that colocalization came at a cost to L. pneumophila fitness. These findings were not completely expected based on known temporal and qualitative differences in the intracellular life cycles of these two pathogens. Taken together, we have developed a new system for studying in vitro Brucella pathogenesis and found a remarkable T4SS-dependent interplay between Brucella and Legionella during macrophage coinfection. Copyright © 2017 American Society for Microbiology.

A charged residue at the subunit interface of PCNA promotes trimer formation by destabilizing alternate subunit interactions

International Nuclear Information System (INIS)

Freudenthal, Bret D.; Gakhar, Lokesh; Ramaswamy, S.; Washington, M. Todd

2009-01-01

Eukaryotic proliferating cell nuclear antigen (PCNA), an essential accessory factor in DNA replication and repair, is a ring-shaped homotrimer. A novel nontrimeric structure of E113G-mutant PCNA protein is reported, which shows that this protein forms alternate subunit interactions. It is concluded that the charged side chain of Glu113 promotes normal trimer formation by destabilizing these alternate subunit interactions. Eukaryotic proliferating cell nuclear antigen (PCNA) is an essential replication accessory factor that interacts with a variety of proteins involved in DNA replication and repair. Each monomer of PCNA has an N-terminal domain A and a C-terminal domain B. In the structure of the wild-type PCNA protein, domain A of one monomer interacts with domain B of a neighboring monomer to form a ring-shaped trimer. Glu113 is a conserved residue at the subunit interface in domain A. Two distinct X-ray crystal structures have been determined of a mutant form of PCNA with a substitution at this position (E113G) that has previously been studied because of its effect on translesion synthesis. The first structure was the expected ring-shaped trimer. The second structure was an unanticipated nontrimeric form of the protein. In this nontrimeric form, domain A of one PCNA monomer interacts with domain A of a neighboring monomer, while domain B of this monomer interacts with domain B of a different neighboring monomer. The B–B interface is stabilized by an antiparallel β-sheet and appears to be structurally similar to the A–B interface observed in the trimeric form of PCNA. The A–A interface, in contrast, is primarily stabilized by hydrophobic interactions. Because the E113G substitution is located on this hydrophobic surface, the A–A interface should be less favorable in the case of the wild-type protein. This suggests that the side chain of Glu113 promotes trimer formation by destabilizing these possible alternate subunit interactions

Destabilized SMC5/6 complex leads to chromosome breakage syndrome with severe lung disease

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van der Crabben, Saskia N.; Hennus, Marije P.; McGregor, Grant A.; Ritter, Deborah I.; Nagamani, Sandesh C.S.; Wells, Owen S.; Harakalova, Magdalena; Chinn, Ivan K.; Alt, Aaron; Vondrova, Lucie; Hochstenbach, Ron; van Montfrans, Joris M.; Terheggen-Lagro, Suzanne W.; van Lieshout, Stef; van Roosmalen, Markus J.; Renkens, Ivo; Duran, Karen; Nijman, Isaac J.; Kloosterman, Wigard P.; Hennekam, Eric; van Hasselt, Peter M.; Wheeler, David A.; Palecek, Jan J.; Lehmann, Alan R.; Oliver, Antony W.; Pearl, Laurence H.; Plon, Sharon E.; Murray, Johanne M.

2016-01-01

The structural maintenance of chromosomes (SMC) family of proteins supports mitotic proliferation, meiosis, and DNA repair to control genomic stability. Impairments in chromosome maintenance are linked to rare chromosome breakage disorders. Here, we have identified a chromosome breakage syndrome associated with severe lung disease in early childhood. Four children from two unrelated kindreds died of severe pulmonary disease during infancy following viral pneumonia with evidence of combined T and B cell immunodeficiency. Whole exome sequencing revealed biallelic missense mutations in the NSMCE3 (also known as NDNL2) gene, which encodes a subunit of the SMC5/6 complex that is essential for DNA damage response and chromosome segregation. The NSMCE3 mutations disrupted interactions within the SMC5/6 complex, leading to destabilization of the complex. Patient cells showed chromosome rearrangements, micronuclei, sensitivity to replication stress and DNA damage, and defective homologous recombination. This work associates missense mutations in NSMCE3 with an autosomal recessive chromosome breakage syndrome that leads to defective T and B cell function and acute respiratory distress syndrome in early childhood. PMID:27427983

Allopregnanolone-induced rise in intracellular calcium in embryonic hippocampal neurons parallels their proliferative potential

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Brinton Roberta

2008-12-01

Full Text Available Abstract Background Factors that regulate intracellular calcium concentration are known to play a critical role in brain function and neural development, including neural plasticity and neurogenesis. We previously demonstrated that the neurosteroid allopregnanolone (APα; 5α-pregnan-3α-ol-20-one promotes neural progenitor proliferation in vitro in cultures of rodent hippocampal and human cortical neural progenitors, and in vivo in triple transgenic Alzheimer's disease mice dentate gyrus. We also found that APα-induced proliferation of neural progenitors is abolished by a calcium channel blocker, nifedipine, indicating a calcium dependent mechanism for the proliferation. Methods In the present study, we investigated the effect of APα on the regulation of intracellular calcium concentration in E18 rat hippocampal neurons using ratiometric Fura2-AM imaging. Results Results indicate that APα rapidly increased intracellular calcium concentration in a dose-dependent and developmentally regulated manner, with an EC50 of 110 ± 15 nM and a maximal response occurring at three days in vitro. The stereoisomers 3β-hydroxy-5α-hydroxy-pregnan-20-one, and 3β-hydroxy-5β-hydroxy-pregnan-20-one, as well as progesterone, were without significant effect. APα-induced intracellular calcium concentration increase was not observed in calcium depleted medium and was blocked in the presence of the broad spectrum calcium channel blocker La3+, or the L-type calcium channel blocker nifedipine. Furthermore, the GABAA receptor blockers bicuculline and picrotoxin abolished APα-induced intracellular calcium concentration rise. Conclusion Collectively, these data indicate that APα promotes a rapid, dose-dependent, stereo-specific, and developmentally regulated increase of intracellular calcium concentration in rat embryonic hippocampal neurons via a mechanism that requires both the GABAA receptor and L-type calcium channel. These data suggest that AP�

Intracellular Bacterial Infections: A Challenge for Developing Cellular Mediated Immunity Vaccines for Farmed Fish

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Hetron Mweemba Munang’andu

2018-04-01

Full Text Available Aquaculture is one of the most rapidly expanding farming systems in the world. Its rapid expansion has brought with it several pathogens infecting different fish species. As a result, there has been a corresponding expansion in vaccine development to cope with the increasing number of infectious diseases in aquaculture. The success of vaccine development for bacterial diseases in aquaculture is largely attributed to empirical vaccine designs based on inactivation of whole cell (WCI bacteria vaccines. However, an upcoming challenge in vaccine design is the increase of intracellular bacterial pathogens that are not responsive to WCI vaccines. Intracellular bacterial vaccines evoke cellular mediated immune (CMI responses that “kill” and eliminate infected cells, unlike WCI vaccines that induce humoral immune responses whose protective mechanism is neutralization of extracellular replicating pathogens by antibodies. In this synopsis, I provide an overview of the intracellular bacterial pathogens infecting different fish species in aquaculture, outlining their mechanisms of invasion, replication, and survival intracellularly based on existing data. I also bring into perspective the current state of CMI understanding in fish together with its potential application in vaccine development. Further, I highlight the immunological pitfalls that have derailed our ability to produce protective vaccines against intracellular pathogens for finfish. Overall, the synopsis put forth herein advocates for a shift in vaccine design to include CMI-based vaccines against intracellular pathogens currently adversely affecting the aquaculture industry.

Semiconductor quantum dots as Förster resonance energy transfer donors for intracellularly-based biosensors

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Field, Lauren D.; Walper, Scott A.; Susumu, Kimihiro; Oh, Eunkeu; Medintz, Igor L.; Delehanty, James B.

2017-02-01

Förster resonance energy transfer (FRET)-based assemblies currently comprise a significant portion of intracellularly based sensors. Although extremely useful, the fluorescent protein pairs typically utilized in such sensors are still plagued by many photophysical issues including significant direct acceptor excitation, small changes in FRET efficiency, and limited photostability. Luminescent semiconductor nanocrystals or quantum dots (QDs) are characterized by many unique optical properties including size-tunable photoluminescence, broad excitation profiles coupled to narrow emission profiles, and resistance to photobleaching, which can cumulatively overcome many of the issues associated with use of fluorescent protein FRET donors. Utilizing QDs for intracellular FRET-based sensing still requires significant development in many areas including materials optimization, bioconjugation, cellular delivery and assay design and implementation. We are currently developing several QD-based FRET sensors for various intracellular applications. These include sensors targeting intracellular proteolytic activity along with those based on theranostic nanodevices for monitoring drug release. The protease sensor is based on a unique design where an intracellularly expressed fluorescent acceptor protein substrate assembles onto a QD donor following microinjection, forming an active complex that can be monitored in live cells over time. In the theranostic configuration, the QD is conjugated to a carrier protein-drug analogue complex to visualize real-time intracellular release of the drug from its carrier in response to an external stimulus. The focus of this talk will be on the design, properties, photophysical characterization and cellular application of these sensor constructs.

Intracellularly Induced Cyclophilins Play an Important Role in Stress Adaptation and Virulence of Brucella abortus

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García Fernández, Lucía; DelVecchio, Vito G.; Briones, Gabriel

2013-01-01

Brucella is an intracellular bacterial pathogen that causes the worldwide zoonotic disease brucellosis. Brucella virulence relies on its ability to transition to an intracellular lifestyle within host cells. Thus, this pathogen must sense its intracellular localization and then reprogram gene expression for survival within the host cell. A comparative proteomic investigation was performed to identify differentially expressed proteins potentially relevant for Brucella intracellular adaptation. Two proteins identified as cyclophilins (CypA and CypB) were overexpressed in the intracellular environment of the host cell in comparison to laboratory-grown Brucella. To define the potential role of cyclophilins in Brucella virulence, a double-deletion mutant was constructed and its resulting phenotype was characterized. The Brucella abortus ΔcypAB mutant displayed increased sensitivity to environmental stressors, such as oxidative stress, pH, and detergents. In addition, the B. abortus ΔcypAB mutant strain had a reduced growth rate at lower temperature, a phenotype associated with defective expression of cyclophilins in other microorganisms. The B. abortus ΔcypAB mutant also displays reduced virulence in BALB/c mice and defective intracellular survival in HeLa cells. These findings suggest that cyclophilins are important for Brucella virulence and survival in the host cells. PMID:23230297

Effects of intracellular iron overload on cell death and identification of potent cell death inhibitors.

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Fang, Shenglin; Yu, Xiaonan; Ding, Haoxuan; Han, Jianan; Feng, Jie

2018-06-11

Iron overload causes many diseases, while the underlying etiologies of these diseases are unclear. Cell death processes including apoptosis, necroptosis, cyclophilin D-(CypD)-dependent necrosis and a recently described additional form of regulated cell death called ferroptosis, are dependent on iron or iron-dependent reactive oxygen species (ROS). However, whether the accumulation of intracellular iron itself induces ferroptosis or other forms of cell death is largely elusive. In present study, we study the role of intracellular iron overload itself-induced cell death mechanisms by using ferric ammonium citrate (FAC) and a membrane-permeable Ferric 8-hydroxyquinoline complex (Fe-8HQ) respectively. We show that FAC-induced intracellular iron overload causes ferroptosis. We also identify 3-phosphoinositide-dependent kinase 1 (PDK1) inhibitor GSK2334470 as a potent ferroptosis inhibitor. Whereas, Fe-8HQ-induced intracellular iron overload causes unregulated necrosis, but partially activates PARP-1 dependent parthanatos. Interestingly, we identify many phenolic compounds as potent inhibitors of Fe-8HQ-induced cell death. In conclusion, intracellular iron overload-induced cell death form might be dependent on the intracellular iron accumulation rate, newly identified cell death inhibitors in our study that target ferroptosis and unregulated oxidative cell death represent potential therapeutic strategies against iron overload related diseases. Copyright © 2018 Elsevier Inc. All rights reserved.

Role of UBIAD1 in Intracellular Cholesterol Metabolism and Vascular Cell Calcification.

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Sha Liu

Full Text Available Vascular calcification is an important risk factor associated with mortality among patients with chronic kidney disease. Intracellular cholesterol metabolism is involved in the process of vascular cell calcification. In this study, we investigated the role of UbiA prenyltransferase domain containing 1 (UBIAD1 in intracellular cholesterol metabolism and vascular cell calcification, and identified its subcellular location. Primary human umbilical vein smooth muscle cells (HUVSMCs were incubated with either growth medium (1.4 mmol/L Pi or calcification medium (CM (3.0 mmol/L Pi. Under treatment with CM, HUVSMCs were further incubated with exogenous cholesterol, or menaquinone-4, a product of UBIAD1. The plasmid and small interfering RNA were transfected in HUVSMCs to alter the expression of UBIAD1. Matrix calcium quantitation, alkaline phosphatase activity, intracellular cholesterol level and menaquinone-4 level were measured. The expression of several genes involved in cholesterol metabolism were analyzed. Using an anti-UBIAD1 antibody, an endoplasmic reticulum marker and a Golgi marker, the subcellular location of UBIAD1 in HUVSMCs was analyzed. CM increased matrix calcium, alkaline phosphatase activity and intracellular cholesterol level, and reduced UBIAD1 expression and menaquinone-4 level. Addition of cholesterol contributed to increased matrix calcification and alkaline phosphatase activity in a dose-dependent manner. Elevated expression of UBIAD1 or menaquinone-4 in HUVSMCs treated with CM significantly reduced intracellular cholesterol level, matrix calcification and alkaline phosphatase activity, but increased menaquinone-4 level. Elevated expression of UBIAD1 or menaquinone-4 reduced the gene expression of sterol regulatory element-binding protein-2, and increased gene expression of ATP binding cassette transporters A1, which are in charge of cholesterol synthesis and efflux. UBIAD1 co-localized with the endoplasmic reticulum marker and

Intracellular bacteria: the origin of dinoflagellate toxicity.

Science.gov (United States)

Silva, E S

1990-01-01

Dinoflagellate blooms of the same species have been registered either as toxic or nontoxic and, in the latter case, toxicity may be of different types. A hypothesis has been formulated according to which the bacteria having in some way taken part in the toxin formation are either inside the dinoflagellate cell or in the nutritive liquid. The presence of intracellular bacteria in those microorganisms has been studied mainly in material from cultures, a few from the sea, and several strains were isolated from different species. Experiments with crossed inoculations have shown that the bacterial strain from Gonyaulax tamarensis caused the cells of some other species to become toxic. From nontoxic clonal cultures of Prorocentrum balticum, Glenodinium foliaceum, and Gyrodinium instriatum, after inoculation of that bacterial strain, cultures were obtained whose cell extracts showed the same kind of toxicity as G. tamarensis. No toxic action could be found in the extracts of the bacterial cells form the assayed strains. The interference of intracellular bacteria in the metabolism of dinoflagellates must be the main cause of their toxicity.

Intracellular Chemistry: Integrating Molecular Inorganic Catalysts with Living Systems.

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Ngo, Anh H; Bose, Sohini; Do, Loi H

2018-03-23

This concept article focuses on the rapid growth of intracellular chemistry dedicated to the integration of small-molecule metal catalysts with living cells and organisms. Although biological systems contain a plethora of biomolecules that can deactivate inorganic species, researchers have shown that small-molecule metal catalysts could be engineered to operate in heterogeneous aqueous environments. Synthetic intracellular reactions have recently been reported for olefin hydrogenation, hydrolysis/oxidative cleavage, azide-alkyne cycloaddition, allylcarbamate cleavage, C-C bond cross coupling, and transfer hydrogenation. Other promising targets for new biocompatible reaction discovery will also be discussed, with a special emphasis on how such innovations could lead to the development of novel technologies and chemical tools. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Streamwise vortices destabilize swimming bluegill sunfish (Lepomis macrochirus).

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Maia, Anabela; Sheltzer, Alex P; Tytell, Eric D

2015-03-01

In their natural environment, fish must swim stably through unsteady flows and vortices, including vertical vortices, typically shed by posts in a flow, horizontal cross-flow vortices, often produced by a step or a waterfall in a stream, and streamwise vortices, where the axis of rotation is aligned with the direction of the flow. Streamwise vortices are commonly shed by bluff bodies in streams and by ships' propellers and axial turbines, but we know little about their effects on fish. Here, we describe how bluegill sunfish use more energy and are destabilized more often in flow with strong streamwise vorticity. The vortices were created inside a sealed flow tank by an array of four turbines with similar diameter to the experimental fish. We measured oxygen consumption for seven sunfish swimming at 1.5 body lengths (BL) s(-1) with the turbines rotating at 2 Hz and with the turbines off (control). Simultaneously, we filmed the fish ventrally and recorded the fraction of time spent maneuvering side-to-side and accelerating forward. Separately, we also recorded lateral and ventral video for a combination of swimming speeds (0.5, 1.5 and 2.5 BL s(-1)) and turbine speeds (0, 1, 2 and 3 Hz), immediately after turning the turbines on and 10 min later to test for accommodation. Bluegill sunfish are negatively affected by streamwise vorticity. Spills (loss of heading), maneuvers and accelerations were more frequent when the turbines were on than in the control treatment. These unsteady behaviors, particularly acceleration, correlated with an increase in oxygen consumption in the vortex flow. Bluegill sunfish are generally fast to recover from roll perturbations and do so by moving their pectoral fins. The frequency of spills decreased after the turbines had run for 10 min, but was still markedly higher than in the control, showing that fish partially adapt to streamwise vorticity, but not completely. Coping with streamwise vorticity may be an important energetic

Intracellular signaling by diffusion: can waves of hydrogen peroxide transmit intracellular information in plant cells?

DEFF Research Database (Denmark)

Vestergaard, Christian L.; Flyvbjerg, Henrik; Møller, Ian Max

2012-01-01

of the physical and biochemical conditions in plant cells. As model system, we use a H(2)O(2) signal originating at the plasma membrane (PM) and spreading through the cytosol. We consider two maximally simple types of signals, isolated pulses and harmonic oscillations. First we consider the basic limits......Amplitude- and frequency-modulated waves of Ca(2+) ions transmit information inside cells. Reactive Oxygen Species (ROS), specifically hydrogen peroxide, have been proposed to have a similar role in plant cells. We consider the feasibility of such an intracellular communication system in view...

Regulation of dopamine transporter trafficking by intracellular amphetamine

DEFF Research Database (Denmark)

Kahlig, Kristopher M; Lute, Brandon J; Wei, Yuqiang

2006-01-01

-induced cell surface DAT redistribution may result in long-lasting changes in DA homeostasis. The molecular mechanism by which AMPH induces trafficking is not clear. Because AMPH is a substrate, we do not know whether extracellular AMPH stimulates trafficking through its interaction with DAT and subsequent...... alteration in DAT function, thereby triggering intracellular signaling or whether AMPH must be transported and then act intracellularly. In agreement with our previous studies, extracellular AMPH caused cytosolic redistribution of the wild-type human DAT (WT-hDAT). However, AMPH did not induce cytosolic...... redistribution in an uptake-impaired hDAT (Y335A-hDAT) that still binds AMPH. The divalent cation zinc (Zn(2+)) inhibits WT-hDAT activity, but it restores Y335A-hDAT uptake. Coadministration of Zn(2+) and AMPH consistently reduced WT-hDAT trafficking but stimulated cytosolic redistribution of Y335A...

Effect of bauhinia bauhinioides kallikrein inhibitor on endothelial proliferation and intracellular calcium concentration.

Science.gov (United States)

Bilgin, M; Burgazli, K M; Rafiq, A; Mericliler, M; Neuhof, C; Oliva, M L; Parahuleva, M; Soydan, N; Doerr, O; Abdallah, Y; Erdogan, A

2014-01-01

Proteinase inhibitors act as a defensive system against predators e.g. insects, in plants. Bauhinia bauhinioides kallikrein inhibitor (BbKI) is a serine proteinase inhibitor, isolated from seeds of Bauhinia bauhinioides and is structurally similar to plant Kunitz-type inhibitors but lacks disulfide bridges. In this study we evaluated the antiproliferative effect of BbKI on endothelial cells and its impact on changes in membrane potential and intracellular calcium. HUVEC proliferation was significantly reduced by incubation with BbKI 50 and 100 µM 12% and 13%. Furthermore, BbKI (100 µM) exposure caused a significant increase in intracellular Ca2+ concentration by 35% as compared to untreated control. The intracellular rise in calcium was not affected by the absence of extracellular calcium. BBKI also caused a significant change in the cell membrane potential but the antiproliferative effect was independent of changes in membrane potential. BBKI has an antiproliferative effect on HUVEC, which is independent of the changes in membrane potential, and it causes an increase in intracellular Ca2+.

Lansoprazole induces apoptosis of breast cancer cells through inhibition of intracellular proton extrusion

Energy Technology Data Exchange (ETDEWEB)

Zhang, Shangrong; Wang, Yifan; Li, Shu Jie, E-mail: shujieli@nankai.edu.cn

2014-06-13

Highlights: • Lansoprazole (LPZ) induces cell apoptosis in breast cancer cells. • LPZ markedly inhibits intracellular proton extrusion. • LPZ induces an increase in intracellular ATP level, lysosomal alkalinization and ROS accumulation. - Abstract: The increased glycolysis and proton secretion in tumors is proposed to contribute to the proliferation and invasion of cancer cells during the process of tumorigenesis and metastasis. Here, treatment of human breast cancer cells with proton pump inhibitor (PPI) lansoprazole (LPZ) induces cell apoptosis in a dose-dependent manner. In the implantation of the MDA-MB-231 xenografts in nude mice, administration of LPZ significantly inhibits tumorigenesis and induces large-scale apopotosis of tumor cells. LPZ markedly inhibits intracellular proton extrusion, induces an increase in intracellular ATP level, lysosomal alkalinization and accumulation of reactive oxygen species (ROS) in breast cancer cells. The ROS scavenger N-acetyl-L-cysteine (NAC) and diphenyleneiodonium (DPI), a specific pharmacological inhibitor of NADPH oxidases (NOX), significantly abolish LPZ-induced ROS accumulation in breast cancer cells. Our results suggested that LPZ may be used as a new therapeutic drug for breast tumor.

Lansoprazole induces apoptosis of breast cancer cells through inhibition of intracellular proton extrusion

International Nuclear Information System (INIS)

Zhang, Shangrong; Wang, Yifan; Li, Shu Jie

2014-01-01

Highlights: • Lansoprazole (LPZ) induces cell apoptosis in breast cancer cells. • LPZ markedly inhibits intracellular proton extrusion. • LPZ induces an increase in intracellular ATP level, lysosomal alkalinization and ROS accumulation. - Abstract: The increased glycolysis and proton secretion in tumors is proposed to contribute to the proliferation and invasion of cancer cells during the process of tumorigenesis and metastasis. Here, treatment of human breast cancer cells with proton pump inhibitor (PPI) lansoprazole (LPZ) induces cell apoptosis in a dose-dependent manner. In the implantation of the MDA-MB-231 xenografts in nude mice, administration of LPZ significantly inhibits tumorigenesis and induces large-scale apopotosis of tumor cells. LPZ markedly inhibits intracellular proton extrusion, induces an increase in intracellular ATP level, lysosomal alkalinization and accumulation of reactive oxygen species (ROS) in breast cancer cells. The ROS scavenger N-acetyl-L-cysteine (NAC) and diphenyleneiodonium (DPI), a specific pharmacological inhibitor of NADPH oxidases (NOX), significantly abolish LPZ-induced ROS accumulation in breast cancer cells. Our results suggested that LPZ may be used as a new therapeutic drug for breast tumor

Biological macromolecules based targeted nanodrug delivery systems for the treatment of intracellular infections.

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Aparna, V; Shiva, M; Biswas, Raja; Jayakumar, R

2018-04-15

Intracellular infections are tricky to treat, the reason being the poor penetration of antibiotics/antimycotics into the microbial niche (host cell). Macrophages are primary targets of facultative and obligate intracellular bacteria/fungi to be abused as host cells. The need for drugs with better intracellular penetration led to the development of endocytosable drug carriers, which can cross the cell membrane of the host cells (macrophages) by imitating the entry path of the pathogens. Therefore, the drugs can be targeted to macrophages ensuring enhanced therapeutic effect. This review discusses the exploitation of various nanocarriers for targeted delivery of drugs to the macrophages in the last two decades. Copyright © 2018 Elsevier B.V. All rights reserved.

Estimating the biophysical properties of neurons with intracellular calcium dynamics.

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Ye, Jingxin; Rozdeba, Paul J; Morone, Uriel I; Daou, Arij; Abarbanel, Henry D I

2014-06-01

We investigate the dynamics of a conductance-based neuron model coupled to a model of intracellular calcium uptake and release by the endoplasmic reticulum. The intracellular calcium dynamics occur on a time scale that is orders of magnitude slower than voltage spiking behavior. Coupling these mechanisms sets the stage for the appearance of chaotic dynamics, which we observe within certain ranges of model parameter values. We then explore the question of whether one can, using observed voltage data alone, estimate the states and parameters of the voltage plus calcium (V+Ca) dynamics model. We find the answer is negative. Indeed, we show that voltage plus another observed quantity must be known to allow the estimation to be accurate. We show that observing both the voltage time course V(t) and the intracellular Ca time course will permit accurate estimation, and from the estimated model state, accurate prediction after observations are completed. This sets the stage for how one will be able to use a more detailed model of V+Ca dynamics in neuron activity in the analysis of experimental data on individual neurons as well as functional networks in which the nodes (neurons) have these biophysical properties.

Azithromycin effectiveness against intracellular infections of Francisella

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Mann Barbara J

2010-04-01

Full Text Available Abstract Background Macrolide antibiotics are commonly administered for bacterial respiratory illnesses. Azithromycin (Az is especially noted for extremely high intracellular concentrations achieved within macrophages which is far greater than the serum concentration. Clinical strains of Type B Francisella (F. tularensis have been reported to be resistant to Az, however our laboratory Francisella strains were found to be sensitive. We hypothesized that different strains/species of Francisella (including Type A may have different susceptibilities to Az, a widely used and well-tolerated antibiotic. Results In vitro susceptibility testing of Az confirmed that F. tularensis subsp. holarctica Live Vaccine Strain (LVS (Type B was not sensitive while F. philomiragia, F. novicida, and Type A F. tularensis (NIH B38 and Schu S4 strain were susceptible. In J774A.1 mouse macrophage cells infected with F. philomiragia, F. novicida, and F. tularensis LVS, 5 μg/ml Az applied extracellularly eliminated intracellular Francisella infections. A concentration of 25 μg/ml Az was required for Francisella-infected A549 human lung epithelial cells, suggesting that macrophages are more effective at concentrating Az than epithelial cells. Mutants of RND efflux components (tolC and ftlC in F. novicida demonstrated less sensitivity to Az by MIC than the parental strain, but the tolC disc-inhibition assay demonstrated increased sensitivity, indicating a complex role for the outer-membrane transporter. Mutants of acrA and acrB mutants were less sensitive to Az than the parental strain, suggesting that AcrAB is not critical for the efflux of Az in F. novicida. In contrast, F. tularensis Schu S4 mutants ΔacrB and ΔacrA were more sensitive than the parental strain, indicating that the AcrAB may be important for Az efflux in F. tularensis Schu S4. F. novicida LPS O-antigen mutants (wbtN, wbtE, wbtQ and wbtA were found to be less sensitive in vitro to Az compared to the wild

Study on the inactivation of intracellular enzyme molecules by X-ray irradiation

International Nuclear Information System (INIS)

Lee, S.B.

1977-01-01

Inactivation of the glutamic acid dehydrogenase and glucose-6-phosphate dehydrogenase enzyme molecules in the Ehrlich ascites tumor cells of the mouse were studied. The above mentioned intracellular enzyme molecules were irradiated by the X-ray radiation under the condition of 65 kV, 1 Amp under the atmosphere of nitrogen gases and by 4 0 C. Thereby, irradiation doses were 580 KR/min(error: +-3%). After irradiation, the cell homogentes were prepared through liquid air techniques. There after, the activities of the enzymes were measured with photometric method given by O. Warburg and W. Christian. The dose effect curves of the activities of the two enzymes by the X-ray irradiation showed both exponential and the inactivation doses were 6.5x10 6 and 5.0x10 6 R respectively. These results showed one side that the inactivation process of the intracellular enzyme molecules was one hit reaction after target theory, and the other side that this inactivation process could not be the primary causes of the death through X-ray irradiation of the vertebrate animals, because of the high resistance of the intracellular protein molecules against X-ray irradiation. The one hit reaction by the inactivation process of the irradiated intracellular enzyme molecules was discussed. (author)

Intracellular pH-sensing using core/shell silica nanoparticles.

Science.gov (United States)

Korzeniowska, B; Woolley, R; DeCourcey, J; Wencel, D; Loscher, C E; McDonagh, C

2014-07-01

An in-depth understanding of biochemical processes occurring within biological systems is key for early diagnosis of disease and identification of appropriate treatments. Nanobiophotonics offers huge potential benefits for intracellular diagnostics and therapeutics. Intracellular sensing using fluorescent nanoparticles is a potentially useful tool for real-time, in vivo monitoring of important cellular analytes. This work is focused on synthesis of optical chemical nanosensors for the quantitative analysis of pH inside living cells. The structure of the nanosensor comprises a biofriendly silica matrix with co-encapsulated Texas Red, acting as a reference dye, and pH-sensitive fluorescein isothiocyanate enabling ratiometric quantitative environmental detection. In order to obtain silica-based nanoparticles -70 nm in size, a modified sol-gel-based Stöber method was employed. The potential of these nanosensors for intracellular pH monitoring is demonstrated inside a live human embryonic kidney cell line whereby a significant change in fluorescence is observed when the cell pH is switched from acidic to basic. High loading efficiencies of nanoparticles into the cells is seen, with little effect on cell morphology even following extended nanoparticle exposure (up to 72 h). Nanoparticle incubation time and the fast response of the nanosensor (-2 s) make it a very powerful tool in monitoring the processes occurring within the cytosol.

Intracellular Hg(0) Oxidation in Desulfovibrio desulfuricans ND132.

Science.gov (United States)

Wang, Yuwei; Schaefer, Jeffra K; Mishra, Bhoopesh; Yee, Nathan

2016-10-03

The disposal of elemental mercury (Hg(0)) wastes in mining and manufacturing areas has caused serious soil and groundwater contamination issues. Under anoxic conditions, certain anaerobic bacteria can oxidize dissolved elemental mercury and convert the oxidized Hg to neurotoxic methylmercury. In this study, we conducted experiments with the Hg-methylating bacterium Desulfovibrio desulfuricans ND132 to elucidate the role of cellular thiols in anaerobic Hg(0) oxidation. The concentrations of cell-surface and intracellular thiols were measured, and specific fractions of D. desulfuricans ND132 were examined for Hg(0) oxidation activity and analyzed with extended X-ray absorption fine structure (EXAFS) spectroscopy. The experimental data indicate that intracellular thiol concentrations are approximately six times higher than those of the cell wall. Cells reacted with a thiol-blocking reagent were severely impaired in Hg(0) oxidation activity. Spheroplasts lacking cell walls rapidly oxidized Hg(0) to Hg(II), while cell wall fragments exhibited low reactivity toward Hg(0). EXAFS analysis of spheroplast samples revealed that multiple different forms of Hg-thiols are produced by the Hg(0) oxidation reaction and that the local coordination environment of the oxidized Hg changes with reaction time. The results of this study indicate that Hg(0) oxidation in D. desulfuricans ND132 is an intracellular process that occurs by reaction with thiol-containing molecules.

Autophagic clearance of bacterial pathogens: molecular recognition of intracellular microorganisms.

Science.gov (United States)

Pareja, Maria Eugenia Mansilla; Colombo, Maria I

2013-01-01

Autophagy is involved in several physiological and pathological processes. One of the key roles of the autophagic pathway is to participate in the first line of defense against the invasion of pathogens, as part of the innate immune response. Targeting of intracellular bacteria by the autophagic machinery, either in the cytoplasm or within vacuolar compartments, helps to control bacterial proliferation in the host cell, controlling also the spreading of the infection. In this review we will describe the means used by diverse bacterial pathogens to survive intracellularly and how they are recognized by the autophagic molecular machinery, as well as the mechanisms used to avoid autophagic clearance.

Salmonella Intracellular Lifestyles and Their Impact on Host-to-Host Transmission.

Science.gov (United States)

Pucciarelli, M Graciela; García-Del Portillo, Francisco

2017-07-01

More than a century ago, infections by Salmonella were already associated with foodborne enteric diseases with high morbidity in humans and cattle. Intestinal inflammation and diarrhea are hallmarks of infections caused by nontyphoidal Salmonella serovars, and these pathologies facilitate pathogen transmission to the environment. In those early times, physicians and microbiologists also realized that typhoid and paratyphoid fever caused by some Salmonella serovars could be transmitted by "carriers," individuals outwardly healthy or at most suffering from some minor chronic complaint. In his pioneering study of the nontyphoidal serovar Typhimurium in 1967, Takeuchi published the first images of intracellular bacteria enclosed by membrane-bound vacuoles in the initial stages of the intestinal epithelium penetration. These compartments, called Salmonella -containing vacuoles, are highly dynamic phagosomes with differing biogenesis depending on the host cell type. Single-cell studies involving real-time imaging and gene expression profiling, together with new approaches based on genetic reporters sensitive to growth rate, have uncovered unprecedented heterogeneous responses in intracellular bacteria. Subpopulations of intracellular bacteria displaying fast, reduced, or no growth, as well as cytosolic and intravacuolar bacteria, have been reported in both in vitro and in vivo infection models. Recent investigations, most of them focused on the serovar Typhimurium, point to the selection of persisting bacteria inside macrophages or following an autophagy attack in fibroblasts. Here, we discuss these heterogeneous intracellular lifestyles and speculate on how these disparate behaviors may impact host-to-host transmissibility of Salmonella serovars.

Experimental Branch Retinal Vein Occlusion Induces Upstream Pericyte Loss and Vascular Destabilization.

Directory of Open Access Journals (Sweden)

Elisa Dominguez

Full Text Available Branch retinal vein occlusion (BRVO leads to extensive vascular remodeling and is important cause of visual impairment. Although the vascular morphological changes following experimental vein occlusion have been described in a variety of models using angiography, the underlying cellular events are ill defined.We here show that laser-induced experimental BRVO in mice leads to a wave of TUNEL-positive endothelial cell (EC apoptosis in the upstream vascular network associated with a transient edema and hemorrhages. Subsequently, we observe an induction of EC proliferation within the dilated vein and capillaries, detected by EdU incorporation, and the edema resolves. However, the pericytes of the upstream capillaries are severely reduced, which was associated with continuing EC apoptosis and proliferation. The vascular remodeling was associated with increased expression of TGFβ, TSP-1, but also FGF2 expression. Exposure of the experimental animals to hypoxia, when pericyte (PC dropout had occurred, led to a dramatic increase in endothelial cell proliferation, confirming the vascular instability induced by the experimental BRVO.Experimental BRVO leads to acute endothelial cells apoptosis and increased permeability. Subsequently the upstream vascular network remains destabilized, characterized by pericyte dropout, un-physiologically high endothelial cells turnover and sensitivity to hypoxia. These early changes might pave the way for capillary loss and subsequent chronic ischemia and edema that characterize the late stage disease.

Tissue- and Cell-Specific Co-localization of Intracellular Gelatinolytic Activity and Matrix Metalloproteinase 2

Science.gov (United States)

Solli, Ann Iren; Fadnes, Bodil; Winberg, Jan-Olof; Uhlin-Hansen, Lars

2013-01-01

Matrix metalloproteinase 2 (MMP-2) is a proteolytic enzyme that degrades extracellular matrix proteins. Recent studies indicate that MMP-2 also has a role in intracellular proteolysis during various pathological conditions, such as ischemic injuries in heart and brain and in tumor growth. The present study was performed to map the distribution of intracellular MMP-2 activity in various mouse tissues and cells under physiological conditions. Samples from normal brain, heart, lung, liver, spleen, pancreas, kidney, adrenal gland, thyroid gland, gonads, oral mucosa, salivary glands, esophagus, intestines, and skin were subjected to high-resolution in situ gelatin zymography and immunohistochemical staining. In hepatocytes, cardiac myocytes, kidney tubuli cells, epithelial cells in the oral mucosa as well as in excretory ducts of salivary glands, and adrenal cortical cells, we found strong intracellular gelatinolytic activity that was significantly reduced by the metalloprotease inhibitor EDTA but not by the cysteine protease inhibitor E-64. Furthermore, the gelatinolytic activity was co-localized with MMP-2. Western blotting and electron microscopy combined with immunogold labeling revealed the presence of MMP-2 in different intracellular compartments of isolated hepatocytes. Our results indicate that MMP-2 takes part in intracellular proteolysis in specific tissues and cells during physiological conditions. PMID:23482328

Intracellular serpins, firewalls and tissue necrosis.

Science.gov (United States)

Marciniak, Stefan J; Lomas, David A

2008-02-01

Luke and colleagues have recently attributed a new role to a member of the serpin superfamily of serine proteinase inhibitors. They have used Caenorhabditis elegans to show that an intracellular serpin is crucial for maintaining lysosomal integrity. We examine the role of this firewall in preventing necrosis and attempt to integrate this with current theories of stress-induced protein degradation. We discuss how mutant serpins cause disease either through polymerization or now, perhaps, by unleashing necrosis.

Cytoplasmic tail of coronavirus spike protein has intracellular

Indian Academy of Sciences (India)

https://www.ias.ac.in/article/fulltext/jbsc/042/02/0231-0244. Keywords. Coronavirus spike protein trafficking; cytoplasmic tail signal; endoplasmic reticulum–Golgi intermediate complex; lysosome. Abstract. Intracellular trafficking and localization studies of spike protein from SARS and OC43 showed that SARS spikeprotein is ...

Facilitating Intracellular Drug Delivery by Ultrasound-Activated Microbubbles

NARCIS (Netherlands)

Lammertink, BHA

2017-01-01

The goal of this thesis was to investigate the combination of ultrasound and microbubbles (USMB) for intracellular delivery of (model) drugs in vitro. We have focused on clinically approved drugs, i.e. cisplatin, and microbubbles, i.e. SonoVue™, to facilitate clinical translation. In addition, model

Heavy metals toxicity after acute exposure of cultured renal cells. Intracellular accumulation and repartition

International Nuclear Information System (INIS)

Khodja, Hicham; Carriere, Marie; Avoscan, Laure; Gouget, Barbara

2005-01-01

Lead (Pb), cadmium (Cd) and uranium (U) present no known biological function but are toxic in various concentration ranges. Pb and Cd lead generally to nephrotoxicity consisting in proximal renal tubular dysfunction and accumulation while U has been reported to induce chemical kidney toxicity, functional and histological damages being as well mainly observed in proximal tubule cells. This work address the question of Cd, Pb, and U cytotoxicity, intracellular accumulation and repartition after acute intoxication of renal proximal tubule epithelial cells. After cells exposure to different concentrations of metals for various times, morphological changes were observed and intracellular concentrations and distributions of toxic metals were specified by PIXE coupled to RBS. Cell viability, measured by biochemical tests, was used as toxicity indicator. A direct correlation between cytotoxicity and intracellular accumulation in renal epithelial cells have been established. Finally, intracellular Pb and U localizations were detected while Cd was found to be uniformly distributed in renal cells. (author)

Detection of Intracellular Factor VIII Protein in Peripheral Blood Mononuclear Cells by Flow Cytometry

Directory of Open Access Journals (Sweden)

Gouri Shankar Pandey

2013-01-01

Full Text Available Flow cytometry is widely used in cancer research for diagnosis, detection of minimal residual disease, as well as immune monitoring and profiling following immunotherapy. Detection of specific host proteins for diagnosis predominantly uses quantitative PCR and western blotting assays. In this study, we optimized a flow cytometry-based detection assay for Factor VIII protein in peripheral blood mononuclear cells (PBMCs. An indirect intracellular staining (ICS method was standardized using monoclonal antibodies to different domains of human Factor VIII protein. The FVIII protein expression level was estimated by calculating the mean and median fluorescence intensities (MFI values for each monoclonal antibody. ICS staining of transiently transfected cell lines supported the method's specificity. Intracellular FVIII protein expression was also detected by the monoclonal antibodies used in the study in PBMCs of five blood donors. In summary, our data suggest that intracellular FVIII detection in PBMCs of hemophilia A patients can be a rapid and reliable method to detect intracellular FVIII levels.

A general mechanism for intracellular toxicity of metal-containing nanoparticles

Science.gov (United States)

Sabella, Stefania; Carney, Randy P.; Brunetti, Virgilio; Malvindi, Maria Ada; Al-Juffali, Noura; Vecchio, Giuseppe; Janes, Sam M.; Bakr, Osman M.; Cingolani, Roberto; Stellacci, Francesco; Pompa, Pier Paolo

2014-05-01

The assessment of the risks exerted by nanoparticles is a key challenge for academic, industrial, and regulatory communities worldwide. Experimental evidence points towards significant toxicity for a range of nanoparticles both in vitro and in vivo. Worldwide efforts aim at uncovering the underlying mechanisms for this toxicity. Here, we show that the intracellular ion release elicited by the acidic conditions of the lysosomal cellular compartment - where particles are abundantly internalized - is responsible for the cascading events associated with nanoparticles-induced intracellular toxicity. We call this mechanism a ``lysosome-enhanced Trojan horse effect'' since, in the case of nanoparticles, the protective cellular machinery designed to degrade foreign objects is actually responsible for their toxicity. To test our hypothesis, we compare the toxicity of similar gold particles whose main difference is in the internalization pathways. We show that particles known to pass directly through cell membranes become more toxic when modified so as to be mostly internalized by endocytosis. Furthermore, using experiments with chelating and lysosomotropic agents, we found that the toxicity mechanism for different metal containing NPs (such as metallic, metal oxide, and semiconductor NPs) is mainly associated with the release of the corresponding toxic ions. Finally, we show that particles unable to release toxic ions (such as stably coated NPs, or diamond and silica NPs) are not harmful to intracellular environments.The assessment of the risks exerted by nanoparticles is a key challenge for academic, industrial, and regulatory communities worldwide. Experimental evidence points towards significant toxicity for a range of nanoparticles both in vitro and in vivo. Worldwide efforts aim at uncovering the underlying mechanisms for this toxicity. Here, we show that the intracellular ion release elicited by the acidic conditions of the lysosomal cellular compartment - where

ORIGINAL ARTICLES

African Journals Online (AJOL)

Hypothyroidism is easily controlled by amiodarone withdrawal and a short course of potassium perchlorate if the condition is transient, or by L-thyroxine replacement in permanent forms. In contrast, amiodarone-induced hyperthyroidism is a more complex therapeutic challenge. Proposed treatment regimens have not been ...

Driving gradual endogenous c-myc overexpression by flow-sorting: intracellular signaling and tumor cell phenotype correlate with oncogene expression

DEFF Research Database (Denmark)

Knudsen, Kasper Jermiin; Holm, G.M.N.; Krabbe, J.S.

2009-01-01

Insulin-exposed rat mammary cancer cells were flow sorted based on a c-myc reporter plasmid encoding a destabilized green fluorescent protein. Sorted cells exhibited gradual increases in c-myc levels. Cells overexpressing c-myc by only 10% exhibited phenotypic changes attributable to c-myc overex...

Medication Review and Transitions of Care: A Case Report of a Decade-Old Medication Error.

Science.gov (United States)

Comer, Rachel; Lizer, Mitsi

2017-10-01

A 69-year-old Caucasian male with a 25-year history of paranoid schizophrenia was brought to the emergency department because of violence toward the staff in his nursing facility. He was diagnosed with a urinary tract infection and was admitted to the behavioral health unit for medication stabilization. History included a five-year state psychiatric hospital admission and nursing facility placement. Because of poor cognitive function, the patient was unable to corroborate medication history, so the pharmacy student on rotation performed an in-depth chart review. The review revealed a transcription error in 2003 deleting amantadine 100 mg twice daily and adding amiodarone 100 mg twice daily. Subsequent hospitalization resulted in another transcription error increasing the amiodarone to 200 mg twice daily. All electrocardiograms conducted were negative for atrial fibrillation. Once detected, the consulted cardiologist discontinued the amiodarone, and the primary care provider was notified via letter and discharge papers. An admission four months later revealed that the nursing facility restarted the amiodarone. Amiodarone was discontinued and the facility was again notified. This case reviews how a 10-year-old medication error went undetected in the electronic medical records through numerous medication reconciliations, but was uncovered when a single comprehensive medication review was conducted.

Preparation and determination of desethylamiodarone in dog lung by HPLC-MS.

Science.gov (United States)

Hong, Liu Xue; Ying, Yang Chuan; Lei, Zheng; Chen, Guo Rui

2011-11-01

A novel method was developed for the preparation and determination of desethylamiodarone in dog lung by high-performance liquid chromatography (HPLC) with amiodarone as a standard. The selected dog was orally given amiodarone, then executed and the active metabolite desethylamiodirone in lung tissue was isolated, concentrated and purified by Waters C18 column (25 mm x 250 mm, 10 microm) with mobile phase of acetonitrile-100 mmol/L acetic acid containing 15 mmol/L diethylamine (55:45 v/v) at a flow rate of 10.0 mL/min. Hypersil ODS2 column (4.6 mm x 250 mm 5 microm) was used to analyze amiodarone and desethylamiodarone, with the same mobile phase at a flow rate of 1.0 mL/min, the detection wavelength was 237.5 nm. Atmosperic pressure electronic spray ionization (AP-ESI) and ion mass spectral (m/z) of 618.1(M + H) were selected to validate desethylamiodarone. The f(i) of desethylamiodarone and amiodarone were 1.04 +/- 0.02 and 1.020 +/- 0.01, respectively. It indicated that desethylamiodarone can be separated and purified by preparational HPLC after Mass Spectrometry (MS) validation and quantified according to f(i) of amiodarone indirectly. The proposed method enables the preparation and determination of desethylamiodarone in dog lung successfully.

Highly Efficient Intracellular Protein Delivery by Cationic Polyethyleneimine-Modified Gelatin Nanoparticles

Directory of Open Access Journals (Sweden)

Ming-Ju Chou

2018-02-01

Full Text Available Intracellular protein delivery may provide a safe and non-genome integrated strategy for targeting abnormal or specific cells for applications in cell reprogramming therapy. Thus, highly efficient intracellular functional protein delivery would be beneficial for protein drug discovery. In this study, we generated a cationic polyethyleneimine (PEI-modified gelatin nanoparticle and evaluated its intracellular protein delivery ability in vitro and in vivo. The experimental results showed that the PEI-modified gelatin nanoparticle had a zeta potential of approximately +60 mV and the particle size was approximately 135 nm. The particle was stable at different biological pH values and temperatures and high protein loading efficiency was observed. The fluorescent image results revealed that large numbers of particles were taken up into the mammalian cells and escaped from the endosomes into the cytoplasm. In a mouse C26 cell-xenograft cancer model, particles accumulated in cancer cells. In conclusion, the PEI-modified gelatin particle may provide a biodegradable and highly efficient protein delivery system for use in regenerative medicine and cancer therapy.

Mechanisms of cellular invasion by intracellular parasites.

Science.gov (United States)

Walker, Dawn M; Oghumu, Steve; Gupta, Gaurav; McGwire, Bradford S; Drew, Mark E; Satoskar, Abhay R

2014-04-01

Numerous disease-causing parasites must invade host cells in order to prosper. Collectively, such pathogens are responsible for a staggering amount of human sickness and death throughout the world. Leishmaniasis, Chagas disease, toxoplasmosis, and malaria are neglected diseases and therefore are linked to socio-economical and geographical factors, affecting well-over half the world's population. Such obligate intracellular parasites have co-evolved with humans to establish a complexity of specific molecular parasite-host cell interactions, forming the basis of the parasite's cellular tropism. They make use of such interactions to invade host cells as a means to migrate through various tissues, to evade the host immune system, and to undergo intracellular replication. These cellular migration and invasion events are absolutely essential for the completion of the lifecycles of these parasites and lead to their for disease pathogenesis. This review is an overview of the molecular mechanisms of protozoan parasite invasion of host cells and discussion of therapeutic strategies, which could be developed by targeting these invasion pathways. Specifically, we focus on four species of protozoan parasites Leishmania, Trypanosoma cruzi, Plasmodium, and Toxoplasma, which are responsible for significant morbidity and mortality.

Intracellular thiol levels and radioresistance: Studies with glutathione and glutathione mono ethyl ester

International Nuclear Information System (INIS)

Astor, M.B.; Meister, A.; Anderson, M.E.

1987-01-01

Intracellular thiols such as glutathione (GSH) protect cells against free radicals formed during oxidative metabolism or from exposure to drugs or ionizing radiation. The role of intracellular GSH in the repair of radiation induced free radical damage was studied using GSH or its analog glutathione mono ethyl ester (GEE), which readily penetrates into the cell. Chinese hamster V79 cells with normal GSH levels were afforded equal protection under aerated and hypoxic conditions (DMF = 1.2 OER = 3.7) by both 10 mM GSH and GEE although GEE had raised interacellular GSH levels three-fold. Growth of V79 cells in cysteine free media resulted in undetectable levels of GSH and OER of 2.2 with no change in aerated survival. Restoration of intracellular GSH by 10 mM GEE resulted in an increase of the OER from 2.2. to 3.8 (DMF = 1.7). Only 14% of the intracellular GSH needs to be repleted to give an OER of 3.0. These experiments provide evidence that thiols do play a role in the oxygen effect and are present at levels in excess of what is necessary for maximal radioprotection

Conditions With High Intracellular Glucose Inhibit Sensing Through Glucose Sensor Snf3 in Saccharomyces cerevisiae

DEFF Research Database (Denmark)

Karhumaa, Kaisa; Wu, B.Q.; Kielland-Brandt, Morten

2010-01-01

as for amino acids. An alternating-access model of the function of transporter-like sensors has been previously suggested based on amino acid sensing, where intracellular ligand inhibits binding of extracellular ligand. Here we studied the effect of intracellular glucose on sensing of extracellular glucose...... through the transporter-like sensor Snf3 in yeast. Sensing through Snf3 was determined by measuring degradation of Mth1 protein. High intracellular glucose concentrations were achieved by using yeast strains lacking monohexose transporters which were grown on maltose. The apparent affinity...... of extracellular glucose to Snf3 was measured for cells grown in non-fermentative medium or on maltose. The apparent affinity for glucose was lowest when the intracellular glucose concentration was high. The results conform to an alternating-access model for transporter-like sensors. J. Cell. Biochem. 110: 920...

Palatal Actinomycosis and Kaposi Sarcoma in an HIV-Infected Subject with Disseminated Mycobacterium avium-intracellulare Infection

Directory of Open Access Journals (Sweden)

Yuria Ablanedo-Terrazas

2012-01-01

Full Text Available Actinomyces and Mycobacterium avium-intracellulare are facultative intracellular organisms, members of the bacterial order actinomycetales. Although Actinomyces can behave as copathogen when anatomic barriers are compromised, its coinfection with Mycobacterium avium-intracellulare has not previously been reported. We present the first reported case of palatal actinomycosis co-infection with disseminated MAC, in an HIV-infected subject with Kaposi sarcoma and diabetes. We discuss the pathogenesis of the complex condition of this subject.

Intracellular nitrate in sediments of an oxygen-deficient marine basin is linked to pelagic diatoms

DEFF Research Database (Denmark)

Kamp, Anja; Petro, Caitlin; Røy, Hans

2018-01-01

Intracellular nitrate is an important electron acceptor in oxygen-deficient aquatic environments, either for the nitrate-storing microbes themselves, or for ambient microbial communities through nitrate leakage. This study links the spatial distribution of intracellular nitrate with the abundance...... and identity of nitrate-storing microbes in sediments of the Bornholm Basin, an environmental showcase for severe hypoxia. Intracellular nitrate (up to 270 nmol cm−3 sediment) was detected at all 18 stations along a 35-km transect through the basin and typically extended as deep as 1.6 cm into the sediment...

Activation of Host IRE1α-Dependent Signaling Axis Contributes the Intracellular Parasitism of Brucella melitensis

Directory of Open Access Journals (Sweden)

Aseem Pandey

2018-04-01

Full Text Available Brucella spp. are intracellular vacuolar pathogens that causes brucellosis, a worldwide zoonosis of profound importance. We previously demonstrated that the activity of host unfolded protein response (UPR sensor IRE1α (inositol-requiring enzyme 1 and ER-associated autophagy confer susceptibility to Brucella melitensis and Brucella abortus intracellular replication. However, the mechanism by which host IRE1α regulates the pathogen intracellular lifestyle remains elusive. In this study, by employing a diverse array of molecular approaches, including biochemical analyses, fluorescence microscopy imaging, and infection assays using primary cells derived from Ern1 (encoding IRE1 conditional knockout mice, we address this gap in our understanding by demonstrating that a novel IRE1α to ULK1, an important component for autophagy initiation, signaling axis confers susceptibility to Brucella intracellular parasitism. Importantly, deletion or inactivation of key signaling components along this axis, including IRE1α, BAK/BAX, ASK1, and JNK as well as components of the host autophagy system ULK1, Atg9a, and Beclin 1, resulted in striking disruption of Brucella intracellular trafficking and replication. Host kinases in the IRE1α-ULK1 axis, including IRE1α, ASK1, JNK1, and/or AMPKα as well as ULK1, were also coordinately phosphorylated in an IRE1α-dependent fashion upon the pathogen infection. Taken together, our findings demonstrate that the IRE1α-ULK1 signaling axis is subverted by the bacterium to promote intracellular parasitism, and provide new insight into our understanding of the molecular mechanisms of intracellular lifestyle of Brucella.

Insight into destabilization mechanism of Mg-based hydrides interstitially co-doped with nonmetals: a DFT study

Science.gov (United States)

Wu, Zhen; Zhu, Luying; Yang, Fusheng; Zhang, Zaoxiao; Nyamsi, Serge N.

2018-04-01

Mg-based metal hydride is one of the most promising materials for hydrogen energy storage. However, the high thermal stability due to strong bonding effects between the atoms limits its practical application. In order to reduce the thermal stability, a method of doping double nonmetals into Mg-based system was proposed in this study. The density functional theory (DFT) calculation results showed that the thermal stabilities of both the B-N co-doped Mg-based alloy and its hydride are reduced compared with pure Mg-based system. The relative formation enthalpies of the alloy and its hydride are 0.323 and 0.595 eV atom-1, respectively. The values are much higher than those for either singly B- or N-doped Mg-based system. The more significant destabilization by doping double nonmetal elements than single element is mainly attributed to a dual effect in weakening Mg-Ni/NiH4 bonds, caused by criss-cross interactions between B-Ni and N-Mg bonds.

Intracellular Zn(2+) signaling in the dentate gyrus is required for object recognition memory.

Science.gov (United States)

Takeda, Atsushi; Tamano, Haruna; Ogawa, Taisuke; Takada, Shunsuke; Nakamura, Masatoshi; Fujii, Hiroaki; Ando, Masaki

2014-11-01

The role of perforant pathway-dentate granule cell synapses in cognitive behavior was examined focusing on synaptic Zn(2+) signaling in the dentate gyrus. Object recognition memory was transiently impaired when extracellular Zn(2+) levels were decreased by injection of clioquinol and N,N,N',N'-tetrakis-(2-pyridylmethyl) ethylendediamine. To pursue the effect of the loss and/or blockade of Zn(2+) signaling in dentate granule cells, ZnAF-2DA (100 pmol, 0.1 mM/1 µl), an intracellular Zn(2+) chelator, was locally injected into the dentate molecular layer of rats. ZnAF-2DA injection, which was estimated to chelate intracellular Zn(2+) signaling only in the dentate gyrus, affected object recognition memory 1 h after training without affecting intracellular Ca(2+) signaling in the dentate molecular layer. In vivo dentate gyrus long-term potentiation (LTP) was affected under the local perfusion of the recording region (the dentate granule cell layer) with 0.1 mM ZnAF-2DA, but not with 1-10 mM CaEDTA, an extracellular Zn(2+) chelator, suggesting that the blockade of intracellular Zn(2+) signaling in dentate granule cells affects dentate gyrus LTP. The present study demonstrates that intracellular Zn(2+) signaling in the dentate gyrus is required for object recognition memory, probably via dentate gyrus LTP expression. Copyright © 2014 Wiley Periodicals, Inc.

Resolution of intracellular calcium metabolism in intact segments of rabbit aorta

International Nuclear Information System (INIS)

Phair, R.D.; Hai, C.M.

1986-01-01

A new method, based on computer-assisted kinetic analysis of 45 Ca efflux data, was used to measure calcium contents and fluxes for extracellular and intracellular compartments in intact segments of rabbit aorta. After a 1-hour loading period, efflux data were collected for 8 hours using a flow-through tissue chamber. These long-term effluxes were necessary because information on intracellular calcium metabolism was concentrated in the slow components of the efflux curves while earlier components appeared to be dominated by washout of extracellular calcium. Intracellular compartments were identified as those whose calcium contents were altered by 10 microM phenylephrine. This method complements previous approaches by providing simultaneous estimates of compartmental calcium contents and fluxes without requiring the assumption of isotopic equilibrium and without recourse to standard wash techniques for removal of extracellular calcium. In normal, calcium-containing, bicarbonate-buffered physiological salt solution these compartments contained a total of approximately 300 nmol Ca/g wet aorta. Of this total, 55 nmol/g were associated with the slowest resolvable compartment whose turnover time was 170 minutes and whose exchange flux was 0.32 nmol min-1g-1. Two other intracellular compartments had turnover times of 30 minutes. One of these was phenylephrine releasable and contained 145 nmol/g; it exchanged calcium at 4.9 nmol min-1g-1. In normal physiological salt solution the plasma membrane was, surprisingly, not rate limiting for Ca efflux; and in 10 microM phenylephrine the membrane Ca flux was even greater, increasing 3.5-fold compared to control

Model-based control of the temporal patterns of intracellular signaling in silico

Science.gov (United States)

Murakami, Yohei; Koyama, Masanori; Oba, Shigeyuki; Kuroda, Shinya; Ishii, Shin

2017-01-01

The functions of intracellular signal transduction systems are determined by the temporal behavior of intracellular molecules and their interactions. Of the many dynamical properties of the system, the relationship between the dynamics of upstream molecules and downstream molecules is particularly important. A useful tool in understanding this relationship is a methodology to control the dynamics of intracellular molecules with an extracellular stimulus. However, this is a difficult task because the relationship between the levels of upstream molecules and those of downstream molecules is often not only stochastic, but also time-inhomogeneous, nonlinear, and not one-to-one. In this paper, we present an easy-to-implement model-based control method that makes the target downstream molecule to trace a desired time course by changing the concentration of a controllable upstream molecule. Our method uses predictions from Monte Carlo simulations of the model to decide the strength of the stimulus, while using a particle-based approach to make inferences regarding unobservable states. We applied our method to in silico control problems of insulin-dependent AKT pathway model and EGF-dependent Akt pathway model with system noise. We show that our method can robustly control the dynamics of the intracellular molecules against unknown system noise of various strengths, even in the absence of complete knowledge of the true model of the target system. PMID:28275530

Trade-Offs of Escherichia coli Adaptation to an Intracellular Lifestyle in Macrophages.

Directory of Open Access Journals (Sweden)

M Azevedo

Full Text Available The bacterium Escherichia coli exhibits remarkable genomic and phenotypic variation, with some pathogenic strains having evolved to survive and even replicate in the harsh intra-macrophage environment. The rate and effects of mutations that can cause pathoadaptation are key determinants of the pace at which E. coli can colonize such niches and become pathogenic. We used experimental evolution to determine the speed and evolutionary paths undertaken by a commensal strain of E. coli when adapting to intracellular life. We estimated the acquisition of pathoadaptive mutations at a rate of 10-6 per genome per generation, resulting in the fixation of more virulent strains in less than a hundred generations. Whole genome sequencing of independently evolved clones showed that the main targets of intracellular adaptation involved loss of function mutations in genes implicated in the assembly of the lipopolysaccharide core, iron metabolism and di- and tri-peptide transport, namely rfaI, fhuA and tppB, respectively. We found a substantial amount of antagonistic pleiotropy in evolved populations, as well as metabolic trade-offs, commonly found in intracellular bacteria with reduced genome sizes. Overall, the low levels of clonal interference detected indicate that the first steps of the transition of a commensal E. coli into intracellular pathogens are dominated by a few pathoadaptive mutations with very strong effects.

The destabilization of the French electricity supply industry nascent competition in an open environment

International Nuclear Information System (INIS)

Finon, D.

2001-06-01

In February 2000 France, compelled by the 1996 European Directive 96/92, undertook a minimal reform of the organisation of its electricity industry, while preserving the boundaries of the incumbent company. The aim of this paper is to analyse the conditions of destabilization of the industrial organisation of the French ESI, by identifying the economic factors of endogenous and exogenous erosion. Firstly, after setting out the main elements of the French reform, which is aimed at making the electricity market contestable, the effectiveness of the ''contestability'' of the French power market is discussed. Secondly in order to test the stability of the new institutional arrangements, an institutional prospect is developed, on the basis of economic factors of instability and resistance, to produce two contrasting scenarios: one in which the particularly French model is retained (limited contestability market scenario); another in which there is movement towards a de-integrated competitive model (contamination by competition scenario). Thirdly the author concludes on the basis of recent elements, that the future would be a mix of these two trajectories which will come within in the progressive integration of the national markets in continental Europe. (A.L.B.)

The destabilization of the French electricity supply industry nascent competition in an open environment

Energy Technology Data Exchange (ETDEWEB)

Finon, D

2001-06-01

In February 2000 France, compelled by the 1996 European Directive 96/92, undertook a minimal reform of the organisation of its electricity industry, while preserving the boundaries of the incumbent company. The aim of this paper is to analyse the conditions of destabilization of the industrial organisation of the French ESI, by identifying the economic factors of endogenous and exogenous erosion. Firstly, after setting out the main elements of the French reform, which is aimed at making the electricity market contestable, the effectiveness of the ''contestability'' of the French power market is discussed. Secondly in order to test the stability of the new institutional arrangements, an institutional prospect is developed, on the basis of economic factors of instability and resistance, to produce two contrasting scenarios: one in which the particularly French model is retained (limited contestability market scenario); another in which there is movement towards a de-integrated competitive model (contamination by competition scenario). Thirdly the author concludes on the basis of recent elements, that the future would be a mix of these two trajectories which will come within in the progressive integration of the national markets in continental Europe. (A.L.B.)

Molecular and biochemical characterization of a novel intracellular invertase from Aspergillus niger with transfructosylating activity

NARCIS (Netherlands)

Goosen, C.; Yuan, X.L.; Munster, J.M. van; Ram, A.F.J.; Maarel, M.J.E.C. van der; Dijkhuizen, L.

2007-01-01

A novel subfamily of putative intracellular invertase enzymes (glycoside hydrolase family 32) has previously been identified in fungal genomes. Here, we report phylogenetic, molecular, and biochemical characteristics of SucB, one of two novel intracellular invertases identified in Aspergillus niger.

Extracellular and Intracellular Mechanisms Mediating Metastatic Activity of Exogenous Osteopontin

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Mandelin, Jami; Lin, Emme C. K.; Hu, Dana D.; Knowles, Susan K.; Do, Kim-Anh; Wang, Xuemei; Sage, E. Helene; Smith, Jeffrey W.; Arap, Wadih; Pasqualini, Renata

2009-01-01

BACKGROUND Osteopontin affects several steps of the metastatic cascade. Despite direct correlation with metastasis in experimental systems and in patient studies, the extracellular and intracellular basis for these observations remains unsolved. We used human melanoma and sarcoma cell lines to evaluate the effects of soluble osteopontin on metastasis. METHODS Exogenous osteopontin or negative controls, including a site-directed mutant osteopontin, were used in functional assays in vitro, ex vivo, and in vivo designed to test extracellular and intracellular mechanisms involved in experimental metastasis. RESULTS In the extracellular environment, we confirm that soluble osteopontin is required for its pro-metastatic effects; this phenomenon is specific, RGD-dependent, and evident in experimental models of metastasis. In the intracellular environment, osteopontin initially induces rapid Tyr-418 dephosphorylation of c-Src, with decreases in actin stress fibers and increased binding to the vascular endothelium. This heretofore undescribed Tyr dephosphorylation is followed by a tandem c-Src phosphorylation after tumor cell attachment to the metastatic site. CONCLUSION Our results reveal a complex molecular interaction as well as a dual role for osteopontin in metastasis that is dependent on whether tumor cells are in circulation or attached. Such context-dependent functional insights may contribute to anti-metastasis strategies. PMID:19224553

Dihydroceramide biology - Structure-specific metabolism and intracellular localization

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Kok, JW; NikolovaKarakashian, M; Klappe, K; Alexander, C; Merrill, AH

1997-01-01

This study utilized fluorescent analogs to characterize the intracellular transport and metabolism of dihydroceramide (DN-Cer), an intermediate in de novo sphingolipid biosynthesis, When 6-[N-(7-nitro-2,1,3-benzoxadiazol-4-yl) amino]hexanoyl-DH-Cer (C-6-NBD-DH-Cer) was incubated with HT29, NRK, BHK,

Intracellular Trafficking Modulation by Ginsenoside Rg3 Inhibits Brucella abortus Uptake and Intracellular Survival within RAW 264.7 Cells.

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Huy, Tran Xuan Ngoc; Reyes, Alisha Wehdnesday Bernardo; Hop, Huynh Tan; Arayan, Lauren Togonon; Min, WonGi; Lee, Hu Jang; Rhee, Man Hee; Chang, Hong Hee; Kim, Suk

2017-03-28

Ginsenoside Rg3, a saponin extracted from ginseng, has various pharmacological and biological activities; however, its effects against Brucella infection are still unclear. Herein, the inhibitory effects of ginsenoside Rg3 against intracellular parasitic Brucella infection were evaluated through bacterial infection, adherence assays, and LAMP-1 colocalization, as well as immunoblotting and FACS for detecting MAPK signaling proteins and F-actin polymerization, respectively. The internalization, intracellular growth, and adherence of Brucella abortus in Rg3-treated RAW 264.7 cells were significantly decreased compared with the Rg3-untreated control. Furthermore, an apparent reduction of F-actin content and intensity of F-actin fluorescence in Rg3-treated cells was observed compared with B. abortus -infected cells without treatment by flow cytometry analysis and confocal microscopy, respectively. In addition, treating cells with Rg3 decreased the phosphorylation of MAPK signaling proteins such as ERK 1/2 and p38 compared with untreated cells. Moreover, the colocalization of B. abortus -containing phagosomes with LAMP-1 was markedly increased in Rg3-treated cells. These findings suggest that ginsenoside Rg3 inhibits B. abortus infection in mammalian cells and can be used as an alternative approach in the treatment of brucellosis.

Pico gauges for minimally invasive intracellular hydrostatic pressure measurements

DEFF Research Database (Denmark)

Knoblauch, Jan; Mullendore, Daniel L.; Jensen, Kaare Hartvig

2014-01-01

Intracellular pressure has a multitude of functions in cells surrounded by a cell wall or similar matrix in all kingdoms of life. The functions include cell growth, nastic movements, and penetration of tissue by parasites. The precise measurement of intracellular pressure in the majority of cells......, however, remains difficult or impossible due to their small size and/or sensitivity to manipulation. Here, we report on a method that allows precise measurements in basically any cell type over all ranges of pressure. It is based on the compression of nanoliter and picoliter volumes of oil entrapped...... in the tip of microcapillaries, which we call pico gauges. The production of pico gauges can be accomplished with standard laboratory equipment, and measurements are comparably easy to conduct. Example pressure measurements are performed on cells that are difficult or impossible to measure with other methods....

Clinical significance and epidemiologic analyses of Mycobacterium avium and Mycobacterium intracellulare lung disease from post-marketing surveillance.

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Suzuki, Katsuhiro; Kurashima, Atsuyuki; Tatsuno, Kinji; Kadota, Jun-Ichi

2018-01-01

In Japan, nontuberculous mycobacterial lung disease is mostly attributable to Mycobacterium avium complex (MAC), i.e., M. avium or M. intracellulare. However, clinical features of the disease caused by these two pathogens have not been studied sufficiently yet. A post-marketing survey of clarithromycin was performed at 130 facilities across Japan. The data on patients with M. avium infection and patients with M. intracellulare infection were selected from this survey for comparison of background variables and clinical features of the two pathogens. Among the patients analyzed (n = 368), 67.4% had M. avium infection and 32.6% had M. intracellulare infection. Stratified analysis revealed no significant differences between the ratio of the two pathogens based on gender, disease type, complication, past medical history, or smoking history. However, the percentage of patients with M. intracellulare infection was significantly higher among those with underlying lung disease than among those without lung disease (p = 0.0217). The percentage of patients with M. intracellulare infection rose significantly with age (p = 0.0296). This age-related change was more significant in women (p = 0.0018). When district-wise analysis was performed for Japan, the percentage of M. intracellulare infection was higher in the Chugoku/Shikoku and Kyushu districts whereas the percentage of M. avium infection was higher in the other districts. This survey revealed some differences in the clinical and epidemiologic features of M. avium and M. intracellulare infection. The significant predominance of M. avium infection among relatively young women is suggestive of an increase in the M. avium/M. intracellulare infection ratio among women in the future. Copyright © 2017 The Japanese Respiratory Society. Published by Elsevier B.V. All rights reserved.

EVIDENCE FOR THE MACROPHAGE INDUCING GENE IN MYCOBACTERIUM INTRACELLULARE

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Background: The Mycobacterium avium Complex (MAC) includes the species M. avium (MA), M. intracellulare (MI), and possibly others. Organisms belonging to the MAC are phylogenetically closely related, opportunistic pathogens. The macrophage inducing gene (mig) is the only well-des...

BI-2 destabilizes HIV-1 cores during infection and Prevents Binding of CPSF6 to the HIV-1 Capsid.

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Fricke, Thomas; Buffone, Cindy; Opp, Silvana; Valle-Casuso, Jose; Diaz-Griffero, Felipe

2014-12-11

The recently discovered small-molecule BI-2 potently blocks HIV-1 infection. BI-2 binds to the N-terminal domain of HIV-1 capsid. BI-2 utilizes the same capsid pocket used by the small molecule PF74. Although both drugs bind to the same pocket, it has been proposed that BI-2 uses a different mechanism to block HIV-1 infection when compared to PF74. This work demonstrates that BI-2 destabilizes the HIV-1 core during infection, and prevents the binding of the cellular factor CPSF6 to the HIV-1 core. Overall this short-form paper suggests that BI-2 is using a similar mechanism to the one used by PF74 to block HIV-1 infection.

Self-Assembly of Multi-nanozymes to Mimic an Intracellular Antioxidant Defense System.

Science.gov (United States)

Huang, Yanyan; Liu, Zhen; Liu, Chaoqun; Ju, Enguo; Zhang, Yan; Ren, Jinsong; Qu, Xiaogang

2016-06-01

In this work, for the first time, we constructed a novel multi-nanozymes cooperative platform to mimic intracellular antioxidant enzyme-based defense system. V2 O5 nanowire served as a glutathione peroxidase (GPx) mimic while MnO2 nanoparticle was used to mimic superoxide dismutase (SOD) and catalase (CAT). Dopamine was used as a linker to achieve the assembling of the nanomaterials. The obtained V2 O5 @pDA@MnO2 nanocomposite could serve as one multi-nanozyme model to mimic intracellular antioxidant enzyme-based defense procedure in which, for example SOD, CAT, and GPx co-participate. In addition, through assembling with dopamine, the hybrid nanocomposites provided synergistic antioxidative effect. Importantly, both in vitro and in vivo experiments demonstrated that our biocompatible system exhibited excellent intracellular reactive oxygen species (ROS) removal ability to protect cell components against oxidative stress, showing its potential application in inflammation therapy. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Vitamin D is positively associated with sperm motility and increases intracellular calcium in human spermatozoa

DEFF Research Database (Denmark)

Blomberg Jensen, Martin; Bjerrum, Poul J; Jessen, Torben E

2011-01-01

BACKGROUND The vitamin D receptor (VDR) is expressed in human spermatozoa, and VDR-knockout mice and vitamin D (VD) deficiency in rodents results in impaired fertility, low sperm counts and a low number of motile spermatozoa. We investigated the role of activated VD (1,25(OH)(2)D(3)) in human...... spermatozoa and whether VD serum levels are associated with semen quality. METHODS Cross-sectional association study of semen quality and VD serum level in 300 men from the general population, and in vitro studies on spermatozoa from 40 men to investigate the effects of VD on intracellular calcium, sperm......M). 1,25(OH)(2)D(3) increased intracellular calcium concentration in human spermatozoa through VDR-mediated calcium release from an intracellular calcium storage, increased sperm motility and induced the acrosome reaction in vitro. CONCLUSIONS 1,25(OH)(2)D(3) increased intracellular calcium...

Engineering of obligate intracellular bacteria: progress, challenges and paradigms

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Over twenty years have passed since the first report of genetic manipulation of an obligate intracellular bacterium. Through progress interspersed by bouts of stagnation, microbiologists and geneticists have developed approaches to genetically manipulate obligates. A brief overview of the current ge...

CRISPR/Cas9 allows efficient and complete knock-in of a destabilization domain-tagged essential protein in a human cell line, allowing rapid knockdown of protein function.

Science.gov (United States)

Park, Arnold; Won, Sohui T; Pentecost, Mickey; Bartkowski, Wojciech; Lee, Benhur

2014-01-01

Although modulation of protein levels is an important tool for study of protein function, it is difficult or impossible to knockdown or knockout genes that are critical for cell growth or viability. For such genes, a conditional knockdown approach would be valuable. The FKBP protein-based destabilization domain (DD)-tagging approach, which confers instability to the tagged protein in the absence of the compound Shield-1, has been shown to provide rapid control of protein levels determined by Shield-1 concentration. Although a strategy to knock-in DD-tagged protein at the endogenous loci has been employed in certain parasite studies, partly due to the relative ease of knock-in as a result of their mostly haploid lifecycles, this strategy has not been demonstrated in diploid or hyperploid mammalian cells due to the relative difficulty of achieving complete knock-in in all alleles. The recent advent of CRISPR/Cas9 homing endonuclease-mediated targeted genome cleavage has been shown to allow highly efficient homologous recombination at the targeted locus. We therefore assessed the feasibility of using CRISPR/Cas9 to achieve complete knock-in to DD-tag the essential gene Treacher Collins-Franceschetti syndrome 1 (TCOF1) in human 293T cells. Using a double antibiotic selection strategy to select clones with at least two knock-in alleles, we obtained numerous complete knock-in clones within three weeks of initial transfection. DD-TCOF1 expression in the knock-in cells was Shield-1 concentration-dependent, and removal of Shield-1 resulted in destabilization of DD-TCOF1 over the course of hours. We further confirmed that the tagged TCOF1 retained the nucleolar localization of the wild-type untagged protein, and that destabilization of DD-TCOF1 resulted in impaired cell growth, as expected for a gene implicated in ribosome biogenesis. CRISPR/Cas9-mediated homologous recombination to completely knock-in a DD tag likely represents a generalizable and efficient strategy to

Monitoring changes in the intracellular calcium concentration and synaptic efficacy in the mollusc Aplysia.

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Ludwar, Bjoern Ch; Evans, Colin G; Cropper, Elizabeth C

2012-07-15

It has been suggested that changes in intracellular calcium mediate the induction of a number of important forms of synaptic plasticity (e.g., homosynaptic facilitation). These hypotheses can be tested by simultaneously monitoring changes in intracellular calcium and alterations in synaptic efficacy. We demonstrate how this can be accomplished by combining calcium imaging with intracellular recording techniques. Our experiments are conducted in a buccal ganglion of the mollusc Aplysia californica. This preparation has a number of experimentally advantageous features: Ganglia can be easily removed from Aplysia and experiments use adult neurons that make normal synaptic connections and have a normal ion channel distribution. Due to the low metabolic rate of the animal and the relatively low temperatures (14-16 °C) that are natural for Aplysia, preparations are stable for long periods of time. To detect changes in intracellular free calcium we will use the cell impermeant version of Calcium Orange which is easily 'loaded' into a neuron via iontophoresis. When this long wavelength fluorescent dye binds to calcium, fluorescence intensity increases. Calcium Orange has fast kinetic properties and, unlike ratiometric dyes (e.g., Fura 2), requires no filter wheel for imaging. It is fairly photo stable and less phototoxic than other dyes (e.g., fluo-3). Like all non-ratiometric dyes, Calcium Orange indicates relative changes in calcium concentration. But, because it is not possible to account for changes in dye concentration due to loading and diffusion, it can not be calibrated to provide absolute calcium concentrations. An upright, fixed stage, compound microscope was used to image neurons with a CCD camera capable of recording around 30 frames per second. In Aplysia this temporal resolution is more than adequate to detect even a single spike induced alteration in the intracellular calcium concentration. Sharp electrodes are simultaneously used to induce and record

Intracellular distribution of nontargeted quantum dots after natural uptake and microinjection

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Damalakiene, Leona; Karabanovas, Vitalijus; Bagdonas, Saulius; Valius, Mindaugas; Rotomskis, Ricardas

2013-01-01

Background: The purpose of this study was to elucidate the mechanism of natural uptake of nonfunctionalized quantum dots in comparison with microinjected quantum dots by focusing on their time-dependent accumulation and intracellular localization in different cell lines. Methods: The accumulation dynamics of nontargeted CdSe/ZnS carboxyl-coated quantum dots (emission peak 625 nm) was analyzed in NIH3T3, MCF-7, and HepG2 cells by applying the methods of confocal and steady-state fluorescence spectroscopy. Intracellular colocalization of the quantum dots was investigated by staining with Lysotracker®. Results: The uptake of quantum dots into cells was dramatically reduced at a low temperature (4°C), indicating that the process is energy-dependent. The uptake kinetics and imaging of intracellular localization of quantum dots revealed three accumulation stages of carboxyl-coated quantum dots at 37°C, ie, a plateau stage, growth stage, and a saturation stage, which comprised four morphological phases: adherence to the cell membrane; formation of granulated clusters spread throughout the cytoplasm; localization of granulated clusters in the perinuclear region; and formation of multivesicular body-like structures and their redistribution in the cytoplasm. Diverse quantum dots containing intracellular vesicles in the range of approximately 0.5–8 μm in diameter were observed in the cytoplasm, but none were found in the nucleus. Vesicles containing quantum dots formed multivesicular body-like structures in NIH3T3 cells after 24 hours of incubation, which were Lysotracker-negative in serum-free medium and Lysotracker-positive in complete medium. The microinjected quantum dots remained uniformly distributed in the cytosol for at least 24 hours. Conclusion: Natural uptake of quantum dots in cells occurs through three accumulation stages via a mechanism requiring energy. The sharp contrast of the intracellular distribution after microinjection of quantum dots in comparison

[Intracellular signaling mechanisms in thyroid cancer].

Science.gov (United States)

Mondragón-Terán, Paul; López-Hernández, Luz Berenice; Gutiérrez-Salinas, José; Suárez-Cuenca, Juan Antonio; Luna-Ceballos, Rosa Isela; Erazo Valle-Solís, Aura

2016-01-01

Thyroid cancer is the most common malignancy of the endocrine system, the papillary variant accounts for 80-90% of all diagnosed cases. In the development of papillary thyroid cancer, BRAF and RAS genes are mainly affected, resulting in a modification of the system of intracellular signaling proteins known as «protein kinase mitogen-activated» (MAPK) which consist of «modules» of internal signaling proteins (Receptor/Ras/Raf/MEK/ERK) from the cell membrane to the nucleus. In thyroid cancer, these signanling proteins regulate diverse cellular processes such as differentiation, growth, development and apoptosis. MAPK play an important role in the pathogenesis of thyroid cancer as they are used as molecular biomarkers for diagnostic, prognostic and as possible therapeutic molecular targets. Mutations in BRAF gene have been correlated with poor response to treatment with traditional chemotherapy and as an indicator of poor prognosis. To review the molecular mechanisms involved in intracellular signaling of BRAF and RAS genes in thyroid cancer. Molecular therapy research is in progress for this type of cancer as new molecules have been developed in order to inhibit any of the components of the signaling pathway (RET/PTC)/Ras/Raf/MEK/ERK; with special emphasis on the (RET/PTC)/Ras/Raf section, which is a major effector of ERK pathway. Copyright © 2016 Academia Mexicana de Cirugía A.C. Publicado por Masson Doyma México S.A. All rights reserved.

Curcumin Mitigates the Intracellular Lipid Deposit Induced by Antipsychotics In Vitro.

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Alberto Canfrán-Duque

Full Text Available First- and second-generation antipsychotics (FGAs and SGAs, respectively, both inhibit cholesterol biosynthesis and impair the intracellular cholesterol trafficking, leading to lipid accumulation in the late endosome/lysosome compartment. In this study we examined if curcumin, a plant polyphenol that stimulates exosome release, can alleviate antipsychotic-induced intracellular lipid accumulation.HepG2 hepatocarcinoma cells were treated with antipsychotics or placebo and DiI-labelled LDL for 18 h and then exposed to curcumin for the last 2 h. Cells and media were collected separately and used for biochemical analyses, electron microscopy and immunocytochemistry. Exosomes were isolated from the incubation medium by ultracentrifugation.Curcumin treatment reduced the number of heterolysosomes and shifted their subcellular localization to the periphery, as revealed by electron microscopy, and stimulated the release of lysosomal β-hexosaminidase and exosome markers flotillin-2 and CD63 into the media. The presence of DiI in exosomes released by cells preloaded with DiI-LDL demonstrated the endolysosomal origin of the microvesicles. Furthermore, curcumin increased the secretion of cholesterol as well as LDL-derived DiI and [3H]-cholesterol, in association with a decrease of intracellular lipids. Thus, the disruption of lipid trafficking induced by FGAs or SGAs can be relieved by curcumin treatment. This polyphenol, however, did not mitigate the reduction of cholesterol esterification induced by antipsychotics.Curcumin stimulates exosome release to remove cholesterol (and presumably other lipids accumulated within the endolysosomal compartment, thereby normalizing intracellular lipid homeostasis. This action may help minimize the adverse metabolic effects of antipsychotic treatment, which should now be evaluated in clinical trials.

Monitoring intracellular oxidative events using dynamic spectral unmixing microscopy

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There is increasing interest in using live-cell imaging to monitor not just individual intracellular endpoints, but to investigate the interplay between multiple molecular events as they unfold in real time within the cell. A major impediment to simultaneous acquisition of multip...

Gold-carbon dots for the intracellular imaging of cancer-derived exosomes

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Jiang, Xiaoyue; Zong, Shenfei; Chen, Chen; Zhang, Yizhi; Wang, Zhuyuan; Cui, Yiping

2018-04-01

As a novel fluorescent nanomaterial, gold-carbon quantum dots (GCDs) possess high biocompatibility and can be easily synthesized by a microwave-assisted method. Owing to their small sizes and unique optical properties, GCDs can be applied to imaging of biological targets, such as cells, exosomes and other organelles. In this study, GCDs were used for fluorescence imaging of exosomes. Tumor-specific antibodies are attached to the GCDs, forming exosome specific nanoprobes. The nanoprobes can label exosomes via immuno-reactions and thus facilitate fluorescent imaging of exosomes. When incubated with live cells, exosomes labeled with the nanoprobes can be taken up by the cells. The intracellular experiments confirmed that the majority of exosomes were endocytosed by cells and transported to lysosomes. The manner by which exosomes were taken up and the intracellular distribution of exosomes are unaffected by the GCDs. The experimental results successfully demonstrated that the presented nanoprobe can be used to study the intrinsic intracellular behavior of tumor derived exosomes. We believe that the GCDs based nanoprobe holds a great promise in the study of exosome related cellular events, such as cancer metastasis.

Flow cytometric analysis of cell-surface and intracellular antigens in leukemia diagnosis.

Science.gov (United States)

Knapp, W; Strobl, H; Majdic, O

1994-12-15

New technology allows highly sensitive flow cytometric detection and quantitative analysis of intracellular antigens in normal and malignant hemopoietic cells. With this technology, the earliest stages of myeloid and lymphoid differentiation can easily and reliably be identified using antibodies directed against (pro-)myeloperoxidase/MPO, CD22 and CD3 antigens, respectively. Particularly for the analysis of undifferentiated acute myeloblastic leukemia (AML) cells, the immunological demonstration of intracellular MPO or its enzymatically inactive proforms is highly relevant, since other myeloid marker molecules such as CD33, CD13, or CDw65 are either not restricted to the granulomonocytic lineage or appear later in differentiation. By combining MPO staining with staining for lactoferrin (LF), undifferentiated cells can be distinguished from the granulomonocytic maturation compartment in bone marrow, since LF is selectively expressed from the myelocyte stage of differentiation onward. The list of informative intracellular antigens to be used in leukemia cell analysis will certainly expand in the near future. One candidate, intracellular CD68, has already been tested by us, and results are presented. Also dealt within this article are surface marker molecules not (as yet) widely used in leukemia cell analysis but with the potential to provide important additional information. Among them are the surface structures CD15, CD15s, CDw65, CD79a (MB-1), CD79b (B29), CD87 (uPA-R), and CD117 (c-kit).

MicroRNA-155 promotes autophagy to eliminate intracellular mycobacteria by targeting Rheb.

Science.gov (United States)

Wang, Jinli; Yang, Kun; Zhou, Lin; Minhaowu; Wu, Yongjian; Zhu, Min; Lai, Xiaomin; Chen, Tao; Feng, Lianqiang; Li, Meiyu; Huang, Chunyu; Zhong, Qiu; Huang, Xi

2013-01-01

Mycobacterium tuberculosis is a hard-to-eradicate intracellular pathogen that infects one-third of the global population. It can live within macrophages owning to its ability to arrest phagolysosome biogenesis. Autophagy has recently been identified as an effective way to control the intracellular mycobacteria by enhancing phagosome maturation. In the present study, we demonstrate a novel role of miR-155 in regulating the autophagy-mediated anti-mycobacterial response. Both in vivo and in vitro studies showed that miR-155 expression was significantly enhanced after mycobacterial infection. Forced expression of miR-155 accelerated the autophagic response in macrophages, thus promoting the maturation of mycobacterial phagosomes and decreasing the survival rate of intracellular mycobacteria, while transfection with miR-155 inhibitor increased mycobacterial survival. However, macrophage-mediated mycobacterial phagocytosis was not affected after miR-155 overexpression or inhibition. Furthermore, blocking autophagy with specific inhibitor 3-methyladenine or silencing of autophagy related gene 7 (Atg7) reduced the ability of miR-155 to promote autophagy and mycobacterial elimination. More importantly, our study demonstrated that miR-155 bound to the 3'-untranslated region of Ras homologue enriched in brain (Rheb), a negative regulator of autophagy, accelerated the process of autophagy and sequential killing of intracellular mycobacteria by suppressing Rheb expression. Our results reveal a novel role of miR-155 in regulating autophagy-mediated mycobacterial elimination by targeting Rheb, and provide potential targets for clinical treatment.

MicroRNA-155 promotes autophagy to eliminate intracellular mycobacteria by targeting Rheb.

Directory of Open Access Journals (Sweden)

Jinli Wang

Full Text Available Mycobacterium tuberculosis is a hard-to-eradicate intracellular pathogen that infects one-third of the global population. It can live within macrophages owning to its ability to arrest phagolysosome biogenesis. Autophagy has recently been identified as an effective way to control the intracellular mycobacteria by enhancing phagosome maturation. In the present study, we demonstrate a novel role of miR-155 in regulating the autophagy-mediated anti-mycobacterial response. Both in vivo and in vitro studies showed that miR-155 expression was significantly enhanced after mycobacterial infection. Forced expression of miR-155 accelerated the autophagic response in macrophages, thus promoting the maturation of mycobacterial phagosomes and decreasing the survival rate of intracellular mycobacteria, while transfection with miR-155 inhibitor increased mycobacterial survival. However, macrophage-mediated mycobacterial phagocytosis was not affected after miR-155 overexpression or inhibition. Furthermore, blocking autophagy with specific inhibitor 3-methyladenine or silencing of autophagy related gene 7 (Atg7 reduced the ability of miR-155 to promote autophagy and mycobacterial elimination. More importantly, our study demonstrated that miR-155 bound to the 3'-untranslated region of Ras homologue enriched in brain (Rheb, a negative regulator of autophagy, accelerated the process of autophagy and sequential killing of intracellular mycobacteria by suppressing Rheb expression. Our results reveal a novel role of miR-155 in regulating autophagy-mediated mycobacterial elimination by targeting Rheb, and provide potential targets for clinical treatment.

Identification of lactoferricin B intracellular targets using an Escherichia coli proteome chip.

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Tu, Yu-Hsuan; Ho, Yu-Hsuan; Chuang, Ying-Chih; Chen, Po-Chung; Chen, Chien-Sheng

2011-01-01

Lactoferricin B (LfcinB) is a well-known antimicrobial peptide. Several studies have indicated that it can inhibit bacteria by affecting intracellular activities, but the intracellular targets of this antimicrobial peptide have not been identified. Therefore, we used E. coli proteome chips to identify the intracellular target proteins of LfcinB in a high-throughput manner. We probed LfcinB with E. coli proteome chips and further conducted normalization and Gene Ontology (GO) analyses. The results of the GO analyses showed that the identified proteins were associated with metabolic processes. Moreover, we validated the interactions between LfcinB and chip assay-identified proteins with fluorescence polarization (FP) assays. Sixteen proteins were identified, and an E. coli interaction database (EcID) analysis revealed that the majority of the proteins that interact with these 16 proteins affected the tricarboxylic acid (TCA) cycle. Knockout assays were conducted to further validate the FP assay results. These results showed that phosphoenolpyruvate carboxylase was a target of LfcinB, indicating that one of its mechanisms of action may be associated with pyruvate metabolism. Thus, we used pyruvate assays to conduct an in vivo validation of the relationship between LfcinB and pyruvate level in E. coli. These results showed that E. coli exposed to LfcinB had abnormal pyruvate amounts, indicating that LfcinB caused an accumulation of pyruvate. In conclusion, this study successfully revealed the intracellular targets of LfcinB using an E. coli proteome chip approach.

Identification of lactoferricin B intracellular targets using an Escherichia coli proteome chip.

Directory of Open Access Journals (Sweden)

Yu-Hsuan Tu

Full Text Available Lactoferricin B (LfcinB is a well-known antimicrobial peptide. Several studies have indicated that it can inhibit bacteria by affecting intracellular activities, but the intracellular targets of this antimicrobial peptide have not been identified. Therefore, we used E. coli proteome chips to identify the intracellular target proteins of LfcinB in a high-throughput manner. We probed LfcinB with E. coli proteome chips and further conducted normalization and Gene Ontology (GO analyses. The results of the GO analyses showed that the identified proteins were associated with metabolic processes. Moreover, we validated the interactions between LfcinB and chip assay-identified proteins with fluorescence polarization (FP assays. Sixteen proteins were identified, and an E. coli interaction database (EcID analysis revealed that the majority of the proteins that interact with these 16 proteins affected the tricarboxylic acid (TCA cycle. Knockout assays were conducted to further validate the FP assay results. These results showed that phosphoenolpyruvate carboxylase was a target of LfcinB, indicating that one of its mechanisms of action may be associated with pyruvate metabolism. Thus, we used pyruvate assays to conduct an in vivo validation of the relationship between LfcinB and pyruvate level in E. coli. These results showed that E. coli exposed to LfcinB had abnormal pyruvate amounts, indicating that LfcinB caused an accumulation of pyruvate. In conclusion, this study successfully revealed the intracellular targets of LfcinB using an E. coli proteome chip approach.

Intracellular vesicles as reproduction elements in cell wall-deficient L-form bacteria.

Directory of Open Access Journals (Sweden)

Yves Briers

Full Text Available Cell wall-deficient bacteria, or L-forms, represent an extreme example of bacterial plasticity. Stable L-forms can multiply and propagate indefinitely in the absence of a cell wall. Data presented here are consistent with the model that intracellular vesicles in Listeria monocytogenes L-form cells represent the actual viable reproductive elements. First, small intracellular vesicles are formed along the mother cell cytoplasmic membrane, originating from local phospholipid accumulation. During growth, daughter vesicles incorporate a small volume of the cellular cytoplasm, and accumulate within volume-expanding mother cells. Confocal Raman microspectroscopy demonstrated the presence of nucleic acids and proteins in all intracellular vesicles, but only a fraction of which reveals metabolic activity. Following collapse of the mother cell and release of the daughter vesicles, they can establish their own membrane potential required for respiratory and metabolic processes. Premature depolarization of the surrounding membrane promotes activation of daughter cell metabolism prior to release. Based on genome resequencing of L-forms and comparison to the parental strain, we found no evidence for predisposing mutations that might be required for L-form transition. Further investigations revealed that propagation by intracellular budding not only occurs in Listeria species, but also in L-form cells generated from different Enterococcus species. From a more general viewpoint, this type of multiplication mechanism seems reminiscent of the physicochemical self-reproducing properties of abiotic lipid vesicles used to study the primordial reproduction pathways of putative prokaryotic precursor cells.

Intracellular vesicles as reproduction elements in cell wall-deficient L-form bacteria.

Science.gov (United States)

Briers, Yves; Staubli, Titu; Schmid, Markus C; Wagner, Michael; Schuppler, Markus; Loessner, Martin J

2012-01-01

Cell wall-deficient bacteria, or L-forms, represent an extreme example of bacterial plasticity. Stable L-forms can multiply and propagate indefinitely in the absence of a cell wall. Data presented here are consistent with the model that intracellular vesicles in Listeria monocytogenes L-form cells represent the actual viable reproductive elements. First, small intracellular vesicles are formed along the mother cell cytoplasmic membrane, originating from local phospholipid accumulation. During growth, daughter vesicles incorporate a small volume of the cellular cytoplasm, and accumulate within volume-expanding mother cells. Confocal Raman microspectroscopy demonstrated the presence of nucleic acids and proteins in all intracellular vesicles, but only a fraction of which reveals metabolic activity. Following collapse of the mother cell and release of the daughter vesicles, they can establish their own membrane potential required for respiratory and metabolic processes. Premature depolarization of the surrounding membrane promotes activation of daughter cell metabolism prior to release. Based on genome resequencing of L-forms and comparison to the parental strain, we found no evidence for predisposing mutations that might be required for L-form transition. Further investigations revealed that propagation by intracellular budding not only occurs in Listeria species, but also in L-form cells generated from different Enterococcus species. From a more general viewpoint, this type of multiplication mechanism seems reminiscent of the physicochemical self-reproducing properties of abiotic lipid vesicles used to study the primordial reproduction pathways of putative prokaryotic precursor cells.

Measurement of Intracellular Ionized Calcium in a Free-living Soil Nematode, Caenorhabditis elegans.

Science.gov (United States)

Kawaii, S; Yoshizawa, Y; Mizutani, J

1993-01-01

A calcium chelating fluorescence indicator, fura-2, was used to measure intracellular ionized calcium in Caenorhabditis elegans. The indicator loading process was harmless to the nematode, and completed within 2-3 h. Fura-2 was loaded mainly at its intestinal tract. The effects of DOPA on locomotion and the level of intracellular calcium were investigated and measured by using a microfluorometer. The addition of DOPA temporarily increased [Ca(2+)]i for several minutes.

Diagnosing MJO Destabilization and Propagation with the Moisture and MSE Budgets

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Maloney, Eric; Wolding, Brandon

2015-04-01

Novel diagnostics obtained as an extension of empirical orthogonal function analysis are used as a composting basis to gain insight into MJO dynamics through examination of reanalysis moisture and moist static energy budgets. The net effect of vertical moisture advection and cloud processes was found to be a modest positive feedback to column moisture anomalies during both enhanced and suppressed phases of the MJO. This positive feedback is regionally strengthened by anomalous surface fluxes of latent heat. The modulation of horizontal synoptic scale eddy mixing acts as a negative feedback to column moisture anomalies, while anomalous winds acting against the mean state moisture gradient aid in eastward propagation. These processes act in a systematic fashion across the Indian Ocean and oceanic regions of the Maritime Continent. The ability to approximately close the MSE budget serves an important role in constraining the moisture budget, whose residual is several times larger than the total and horizontal advective moisture tendencies. Comparison with TRMM precipitation anomalies suggests that the moisture budget residual results from an underestimation by ERAi of variations in both total precipitation and vertical moisture advection associated with the MJO. The results of this study support the concept of the MJO as a moisture-mode. This analysis is extended to examine the impact of boundary layer convergence driven by MJO SST anomalies on the vertically-integrated moisture budget. Results from a coupled version of the SP-CAM suggest that SST-driven moisture convergence anomalies are of a sufficient amplitude to be important for MJO propagation and destabilization, and may help explain why coupled models produce better simulations of the MJO than uncoupled models.

Fluorescent nanoparticles for intracellular sensing: a review.

Science.gov (United States)

Ruedas-Rama, Maria J; Walters, Jamie D; Orte, Angel; Hall, Elizabeth A H

2012-11-02

Fluorescent nanoparticles (NPs), including semiconductor NPs (Quantum Dots), metal NPs, silica NPs, polymer NPs, etc., have been a major focus of research and development during the past decade. The fluorescent nanoparticles show unique chemical and optical properties, such as brighter fluorescence, higher photostability and higher biocompatibility, compared to classical fluorescent organic dyes. Moreover, the nanoparticles can also act as multivalent scaffolds for the realization of supramolecular assemblies, since their high surface to volume ratio allow distinct spatial domains to be functionalized, which can provide a versatile synthetic platform for the implementation of different sensing schemes. Their excellent properties make them one of the most useful tools that chemistry has supplied to biomedical research, enabling the intracellular monitoring of many different species for medical and biological purposes. In this review, we focus on the developments and analytical applications of fluorescent nanoparticles in chemical and biological sensing within the intracellular environment. The review also points out the great potential of fluorescent NPs for fluorescence lifetime imaging microscopy (FLIM). Finally, we also give an overview of the current methods for delivering of fluorescent NPs into cells, where critically examine the benefits and liabilities of each strategy. Copyright © 2012 Elsevier B.V. All rights reserved.

Antimycobacterial Efficacy of Andrographis paniculata Leaf Extracts Under Intracellular and Hypoxic Conditions.

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Bhatter, Purva; Gupta, Pooja; Daswani, Poonam; Tetali, Pundarikakshudu; Birdi, Tannaz

2015-01-01

The inhibition of the growth of Mycobacterium tuberculosis by the extracts of Andrographis paniculata has been studied using intracellular and axenic hypoxic conditions. The inhibition (confirmed using the gold standard colony forming unit assay) was found to increase with "double stimuli" or higher concentration of the extract. Organic solvent extracts were found to inhibit bacterial growth more than the aqueous extracts under microaerophilic conditions mimicked through axenic and intracellular assays. This could be further explored to evaluate the potential of the plant to be used against nonreplicating/dormant bacilli. © The Author(s) 2014.

A general mechanism for intracellular toxicity of metal-containing nanoparticles

KAUST Repository

Sabella, Stefania

2014-04-09

The assessment of the risks exerted by nanoparticles is a key challenge for academic, industrial, and regulatory communities worldwide. Experimental evidence points towards significant toxicity for a range of nanoparticles both in vitro and in vivo. Worldwide efforts aim at uncovering the underlying mechanisms for this toxicity. Here, we show that the intracellular ion release elicited by the acidic conditions of the lysosomal cellular compartment-where particles are abundantly internalized-is responsible for the cascading events associated with nanoparticles-induced intracellular toxicity. We call this mechanism a "lysosome-enhanced Trojan horse effect" since, in the case of nanoparticles, the protective cellular machinery designed to degrade foreign objects is actually responsible for their toxicity. To test our hypothesis, we compare the toxicity of similar gold particles whose main difference is in the internalization pathways. We show that particles known to pass directly through cell membranes become more toxic when modified so as to be mostly internalized by endocytosis. Furthermore, using experiments with chelating and lysosomotropic agents, we found that the toxicity mechanism for different metal containing NPs (such as metallic, metal oxide, and semiconductor NPs) is mainly associated with the release of the corresponding toxic ions. Finally, we show that particles unable to release toxic ions (such as stably coated NPs, or diamond and silica NPs) are not harmful to intracellular environments. The Royal Society of Chemistry 2014.

A general mechanism for intracellular toxicity of metal-containing nanoparticles

KAUST Repository

Sabella, Stefania; Carney, Randy P.; Brunetti, Virgilio; Malvindi, Maria Ada; Al-Juffali, Noura; Vecchio, Giuseppe; Janes, Sam M.; Bakr, Osman; Cingolani, Roberto; Stellacci, Francesco; Pompa, Pier Paolo

2014-01-01

The assessment of the risks exerted by nanoparticles is a key challenge for academic, industrial, and regulatory communities worldwide. Experimental evidence points towards significant toxicity for a range of nanoparticles both in vitro and in vivo. Worldwide efforts aim at uncovering the underlying mechanisms for this toxicity. Here, we show that the intracellular ion release elicited by the acidic conditions of the lysosomal cellular compartment-where particles are abundantly internalized-is responsible for the cascading events associated with nanoparticles-induced intracellular toxicity. We call this mechanism a "lysosome-enhanced Trojan horse effect" since, in the case of nanoparticles, the protective cellular machinery designed to degrade foreign objects is actually responsible for their toxicity. To test our hypothesis, we compare the toxicity of similar gold particles whose main difference is in the internalization pathways. We show that particles known to pass directly through cell membranes become more toxic when modified so as to be mostly internalized by endocytosis. Furthermore, using experiments with chelating and lysosomotropic agents, we found that the toxicity mechanism for different metal containing NPs (such as metallic, metal oxide, and semiconductor NPs) is mainly associated with the release of the corresponding toxic ions. Finally, we show that particles unable to release toxic ions (such as stably coated NPs, or diamond and silica NPs) are not harmful to intracellular environments. The Royal Society of Chemistry 2014.

Dynamics of postirradiation intracellular cysteine and aspartic proteinases profiles in proliferating and nonproliferating mammalian cells

International Nuclear Information System (INIS)

Korbelik, M.; Osmak, M.; Suhar, A.; Turk, V.; Skrk, J.

1990-01-01

Dynamics of postirradiation intracellular cysteine and aspartic proteinases profiles were examined in proliferating and nonproliferating Chinese hamster fibroblasts (V 79). The results show that there are significant alterations in cysteine and aspartic intracellular proteinases activity already in the early postirradiation period, which are different in proliferating and nonproliferating cells. Irradiation of the cells examined to low doses and up to 15 Gy induced an increase in cysteine proteinases activity in the early postexposure period, while at higher irradiation doses applied, the activity of these proteinases was decreased. These observations suggest that intracellular proteinases are actively participating in process involving recovery from radiation injury or cell killing. (orig.) [de

Bovine lactoferrin and lactoferricin interfere with intracellular trafficking of Herpes simplex virus-1.

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Marr, A K; Jenssen, H; Moniri, M Roshan; Hancock, R E W; Panté, N

2009-01-01

Although both lactoferrin (Lf), a component of the innate immune system of living organisms, and its N-terminal pepsin cleavage product lactoferricin (Lfcin) have anti-herpes activity, the precise mechanisms by which Lf and Lfcin bring about inhibition of herpes infections are not fully understood. In the present study, experiments were carried out to characterize the activity of bovine Lf and Lfcin (BLf and BLfcin) against the Herpes simplex virus-1 (HSV-1). HSV-1 cellular uptake and intracellular trafficking were studied by immunofluorescence microscopy. In comparison to the untreated infected control cells, both the BLf- and BLfcin-treated cells showed a significant reduction in HSV-1 cellular uptake. The few virus particles that were internalized appeared to have a delayed intracellular trafficking. Thus, in addition to their interference with the uptake of the virus into host cells, Lf and Lfcin also exert their antiviral effect intracellularly.

Single quantum dot tracking reveals the impact of nanoparticle surface on intracellular state.

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Zahid, Mohammad U; Ma, Liang; Lim, Sung Jun; Smith, Andrew M

2018-05-08

Inefficient delivery of macromolecules and nanoparticles to intracellular targets is a major bottleneck in drug delivery, genetic engineering, and molecular imaging. Here we apply live-cell single-quantum-dot imaging and tracking to analyze and classify nanoparticle states after intracellular delivery. By merging trajectory diffusion parameters with brightness measurements, multidimensional analysis reveals distinct and heterogeneous populations that are indistinguishable using single parameters alone. We derive new quantitative metrics of particle loading, cluster distribution, and vesicular release in single cells, and evaluate intracellular nanoparticles with diverse surfaces following osmotic delivery. Surface properties have a major impact on cell uptake, but little impact on the absolute cytoplasmic numbers. A key outcome is that stable zwitterionic surfaces yield uniform cytosolic behavior, ideal for imaging agents. We anticipate that this combination of quantum dots and single-particle tracking can be widely applied to design and optimize next-generation imaging probes, nanoparticle therapeutics, and biologics.

Regulation of intracellular pH in cnidarians: response to acidosis in Anemonia viridis.

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Laurent, Julien; Venn, Alexander; Tambutté, Éric; Ganot, Philippe; Allemand, Denis; Tambutté, Sylvie

2014-02-01

The regulation of intracellular pH (pHi) is a fundamental aspect of cell physiology that has received little attention in studies of the phylum Cnidaria, which includes ecologically important sea anemones and reef-building corals. Like all organisms, cnidarians must maintain pH homeostasis to counterbalance reductions in pHi, which can arise because of changes in either intrinsic or extrinsic parameters. Corals and sea anemones face natural daily changes in internal fluids, where the extracellular pH can range from 8.9 during the day to 7.4 at night. Furthermore, cnidarians are likely to experience future CO₂-driven declines in seawater pH, a process known as ocean acidification. Here, we carried out the first mechanistic investigation to determine how cnidarian pHi regulation responds to decreases in extracellular and intracellular pH. Using the anemone Anemonia viridis, we employed confocal live cell imaging and a pH-sensitive dye to track the dynamics of pHi after intracellular acidosis induced by acute exposure to decreases in seawater pH and NH₄Cl prepulses. The investigation was conducted on cells that contained intracellular symbiotic algae (Symbiodinium sp.) and on symbiont-free endoderm cells. Experiments using inhibitors and Na⁺-free seawater indicate a potential role of Na⁺/H⁺ plasma membrane exchangers (NHEs) in mediating pHi recovery following intracellular acidosis in both cell types. We also measured the buffering capacity of cells, and obtained values between 20.8 and 43.8 mM per pH unit, which are comparable to those in other invertebrates. Our findings provide the first steps towards a better understanding of acid-base regulation in these basal metazoans, for which information on cell physiology is extremely limited. © 2013 FEBS.

Intracellular compartmentalization of skeletal muscle glycogen metabolism and insulin signalling

DEFF Research Database (Denmark)

Prats Gavalda, Clara; Gomez-Cabello, Alba; Vigelsø Hansen, Andreas

2011-01-01

The interest in skeletal muscle metabolism and insulin signalling has increased exponentially in recent years as a consequence of their role in the development of type 2 diabetes mellitus. Despite this, the exact mechanisms involved in the regulation of skeletal muscle glycogen metabolism...... and insulin signalling transduction remain elusive. We believe that one of the reasons is that the role of intracellular compartmentalization as a regulator of metabolic pathways and signalling transduction has been rather ignored. This paper briefly reviews the literature to discuss the role of intracellular...... compartmentalization in the regulation of skeletal muscle glycogen metabolism and insulin signalling. As a result, a hypothetical regulatory mechanism is proposed by which cells could direct glycogen resynthesis towards different pools of glycogen particles depending on the metabolic needs. Furthermore, we discuss...

Rapid extra-/intracellular biosynthesis of gold nanoparticles by the fungus Penicillium sp.

Science.gov (United States)

Du, Liangwei; Xian, Liang; Feng, Jia-Xun

2011-03-01

In this work, the fungus Penicillium was used for rapid extra-/intracellular biosynthesis of gold nanoparticles. AuCl4 - ions reacted with the cell filtrate of Penicillium sp. resulting in extracellular biosynthesis of gold nanoparticles within 1 min. Intracellular biosynthesis of gold nanoparticles was obtained by incubating AuCl4 - solution with fungal biomass for 8 h. The gold nanoparticles were characterized by means of visual observation, UV-Vis absorption spectroscopy, X-ray diffraction (XRD), transmission electron microscopy (TEM), scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDX). The extracellular nanoparticles exhibited maximum absorbance at 545 nm in UV-Vis spectroscopy. The XRD spectrum showed Bragg reflections corresponding to the gold nanocrystals. TEM exhibited the formed spherical gold nanoparticles in the size range from 30 to 50 nm with an average size of 45 nm. SEM and TEM revealed that the intracellular gold nanoparticles were well dispersed on the cell wall and within the cell, and they are mostly spherical in shape with an average diameter of 50 nm. The presence of gold was confirmed by EDX analysis.

FLIPR assays of intracellular calcium in GPCR drug discovery

DEFF Research Database (Denmark)

Hansen, Kasper Bø; Bräuner-Osborne, Hans

2009-01-01

Fluorescent dyes sensitive to changes in intracellular calcium have become increasingly popular in G protein-coupled receptor (GPCR) drug discovery for several reasons. First of all, the assays using the dyes are easy to perform and are of low cost compared to other assays. Second, most non...

Mycobacterium intracellulare Pleurisy Identified on Liquid Cultures of the Pleural Fluid and Pleural Biopsy

OpenAIRE

Lim, Jong Gu; O, Sei Won; Lee, Ki Dong; Suk, Dong Keun; Jung, Tae Young; Shim, Tae Sun; Chon, Gyu Rak

2013-01-01

Pleural effusion is a rare complication in non-tuberculous mycobacterial infection. We report a case of Mycobacterium intracellulare pleuritis with idiopathic pulmonary fibrosis in a 69-year-old man presenting with dyspnea. Pleural effusion revealed lymphocyte dominant exudate. M. intracellulare was identified using a polymerase chain reaction-restriction fragment length polymorphism method and liquid cultures of pleural effusion and pleural biopsy. After combination therapy for M. intracellu...

Molecular characterization of a novel intracellular ADP-ribosyl cyclase.

Directory of Open Access Journals (Sweden)

Dev Churamani

2007-08-01

Full Text Available ADP-ribosyl cyclases are remarkable enzymes capable of catalyzing multiple reactions including the synthesis of the novel and potent intracellular calcium mobilizing messengers, cyclic ADP-ribose and NAADP. Not all ADP-ribosyl cyclases however have been characterized at the molecular level. Moreover, those that have are located predominately at the outer cell surface and thus away from their cytosolic substrates.Here we report the molecular cloning of a novel expanded family of ADP-ribosyl cyclases from the sea urchin, an extensively used model organism for the study of inositol trisphosphate-independent calcium mobilization. We provide evidence that one of the isoforms (SpARC1 is a soluble protein that is targeted exclusively to the endoplasmic reticulum lumen when heterologously expressed. Catalytic activity of the recombinant protein was readily demonstrable in crude cell homogenates, even under conditions where luminal continuity was maintained.Our data reveal a new intracellular location for ADP-ribosyl cyclases and suggest that production of calcium mobilizing messengers may be compartmentalized.

Factors influencing intracellular uptake and radiosensitization by 2-nitroimidazoles in vitro

International Nuclear Information System (INIS)

Brown, D.M.; Gonzalez-Mendez, R.; Brown, J.M.

1983-01-01

In this study it is shown that the radiosensitization of hypoxic Chinese hamster ovary (HA-1) cells in vitro by misonidazole (MIS) and other 1-substituted 2-nitroimidazoles depends on the rate and extent of intracellular uptake of these radiosensitizers, which in turn is governed by their lipophilicity [expressed as the octanol:water partition coefficient (P)]. As the lipophilicity of the compounds decreased, the rate of drug entry into the cells was slower, and below P values of approximately 0.05, peak intracellular drug concentrations were found to be lower than that of MIS (P=0.43). In addition, the number of hydroxyl groups on the side chain of the nitroimidazole molecule influenced the uptake of drug into the cells. For compounds of similar P, but differing in the number of side-chain hydroxyl groups, the addition of a single hydroxyl group to the molecule decreased the amount of drug entering the cell by a factor of approximately 2. These compounds enter the cell by nonmediated passive diffusion since altering the energy (ATP) capacity of the cell by 2-deoxyglucose did not affect uptake. It is also shown that increases in temperature or decreases in pH can increase the intracellular uptake of MIS. For example, equal intracellular and extracellular concentrations (100% uptake) of MIS were obtained if cells were heated to 44-45 0 C for 15 min compared to 20-40% uptake at 37 0 C. Increases in MIS uptake by factors of 2 to 3 could be demonstrated within 30 min when cells were incubated in Hanks' balanced salt solution at pH between 6.0 and 6.3 without loss of cell viability. In addition, MIS uptake in aerobic cultured cells varied from 15 to 60% depending on the cell line and culure conditions used

Impact of intracellular metallothionein on metal biouptake and partitioning dynamics at bacterial interfaces.

Science.gov (United States)

Présent, Romain M; Rotureau, Elise; Billard, Patrick; Pagnout, Christophe; Sohm, Bénédicte; Flayac, Justine; Gley, Renaud; Pinheiro, José P; Duval, Jérôme F L

2017-11-08

Genetically engineered microorganisms are alternatives to physicochemical methods for remediation of metal-contaminated aquifers due to their remarkable bioaccumulation capacities. The design of such biosystems would benefit from the elaboration of a sound quantitative connection between performance in terms of metal removal from aqueous solution and dynamics of the multiscale processes leading to metal biouptake. In this work, this elaboration is reported for Escherichia coli cells modified to overexpress intracellular metallothionein (MTc), a strong proteinaceous metal chelator. Depletion kinetics of Cd(ii) from bulk solution following biouptake and intracellular accumulation is addressed as a function of cell volume fraction using electroanalytical probes and ligand exchange-based analyses. It is shown that metal biouptake in the absence and presence of MTc is successfully interpreted on the basis of a formalism recently developed for metal partitioning dynamics at biointerfaces with integration of intracellular metal speciation. The analysis demonstrates how fast sequestration of metals by intracellular MTc bypasses metal excretion (efflux) and enhances the rate of metal depletion to an extent such that complete removal is achieved at sufficiently large cell volume fractions. The magnitude of the stability constant of nanoparticulate metal-MTc complexes, as derived from refined analysis of macroscopic bulk metal depletion data, is further confirmed by independent electrochemical measurement of metal binding by purified MTc extracts.

A new type of intracellular retention signal identified in a pestivirus structural glycoprotein.

Science.gov (United States)

Burrack, Sandra; Aberle, Daniel; Bürck, Jochen; Ulrich, Anne S; Meyers, Gregor

2012-08-01

Sorting of membrane proteins into intracellular organelles is crucial for cell function. Viruses exploit intracellular transport and retention systems to concentrate envelope proteins at the site of virus budding. In pestiviruses, a group of important pathogens of pigs and ruminants closely related to human hepatitis C virus, the E(rns) protein translated from the viral RNA is secreted from the infected cells and found in the serum of infected animals. Secretion of the protein is regarded as crucial for its function as a viral virulence factor associated with its RNase activity. However, ∼95% of the E(rns) molecules are retained within the infected cell. Fusion of different E(rns) fragments to the C terminus of CD72 allowed identification of a retention signal within the C-terminal 65 aa of the viral protein. This C-terminal sequence represents its membrane anchor and folds into an amphipathic helix binding in-plane to the membrane surface. Residues L183, I190, and L208 are important for intracellular location of E(rns). Presentation of the retention signal on the cytoplasmic instead of the luminal face of the ER membrane in CD8α fusion proteins still led to retention. Thus, E(rns) contains in its C-terminal amphipathic helix an intracellular retention signal that is active on both faces of the membrane.

Reduced intracellular c-di-GMP content increases expression of quorum sensing-regulated genes in Pseudomonas aeruginosa

DEFF Research Database (Denmark)

Chua, Song Lin; Liu, Yang; Li, Yingying

2017-01-01

Cyclic-di-GMP (c-di-GMP) is an intracellular secondary messenger which controls the biofilm life cycle in many bacterial species. High intracellular c-di-GMP content enhances biofilm formation via the reduction of motility and production of biofilm matrix, while low c-di-GMP content in biofilm...... cells leads to increased motility and biofilm dispersal. While the effect of high c-di-GMP levels on bacterial lifestyles is well studied, the physiology of cells at low c-di-GMP levels remains unclear. Here, we showed that Pseudomonas aeruginosa cells with high and low intracellular c-di-GMP contents...... possessed distinct transcriptome profiles. There were 535 genes being upregulated and 432 genes downregulated in cells with low c-di-GMP, as compared to cells with high c-di-GMP. Interestingly, both rhl and pqs quorum-sensing (QS) operons were expressed at higher levels in cells with low intracellular c-di-GMP...

Intracellular ion monitoring using a gold-core polymer-shell nanosensor architecture

Energy Technology Data Exchange (ETDEWEB)

Stanca, S E; Cranfield, C G; Biskup, C [Biomolecular Photonics Group, University Hospital Jena, Teichgraben 8, 07743 Jena (Germany); Nietzsche, S [Centre for Electron Microscopy, University Hospital Jena, Ziegel-muehlenweg 1, 07743 Jena (Germany); Fritzsche, W, E-mail: sarmiza.stanca@mti.uni-jena.de, E-mail: charles.cranfield@mti.uni-jena.de, E-mail: christoph.biskup@mti.uni-jena.de [Institute of Photonic Technology, Albert-Einstein-Strasse 9, 07745 Jena (Germany)

2010-02-05

In this study, we describe the design of new ratiometric fluorescent nanosensors, whose architecture is based on a gold core surrounded by a poly(vinyl alcohol)-polyacetal shell. To the gold core, indicator dyes and reference dyes are attached via a cysteine linker. This nanosensor architecture is flexible with regards to the number and types of fluorophore linkages possible. The robust poly(vinyl alcohol)-polyacetal shell protects the fluorophores linked to the core from non-specific interactions with intracellular proteins. The nanosensors developed in this way are biocompatible and can be easily incorporated into mammalian cells without the use of transfection agents. Here, we show the application of these nanosensors for intracellular pH and sodium ion measurements.

Intracellular Hyper-Acidification Potentiated by Hydrogen Sulfide Mediates Invasive and Therapy Resistant Cancer Cell Death

Directory of Open Access Journals (Sweden)

Zheng-Wei Lee

2017-10-01

Full Text Available Slow and continuous release of H2S by GYY4137 has previously been demonstrated to kill cancer cells by increasing glycolysis and impairing anion exchanger and sodium/proton exchanger activity. This action is specific for cancer cells. The resulting lactate overproduction and defective pH homeostasis bring about intracellular acidification-induced cancer cell death. The present study investigated the potency of H2S released by GYY4137 against invasive and radio- as well as chemo-resistant cancers, known to be glycolytically active. We characterized and utilized cancer cell line pairs of various organ origins, based on their aggressive behaviors, and assessed their response to GYY4137. We compared glycolytic activity, via lactate production, and intracellular pH of each cancer cell line pair after exposure to H2S. Invasive and therapy resistant cancers, collectively termed aggressive cancers, are receptive to H2S-mediated cytotoxicity, albeit at a higher concentration of GYY4137 donor. While lactate production was enhanced, intracellular pH of aggressive cancers was only modestly decreased. Inherently, the magnitude of intracellular pH decrease is a key determinant for cancer cell sensitivity to H2S. We demonstrated the utility of coupling GYY4137 with either simvastatin, known to inhibit monocarboxylate transporter 4 (MCT4, or metformin, to further boost glycolysis, in bringing about cell death for aggressive cancers. Simvastatin inhibiting lactate extrusion thence contained excess lactate induced by GYY4137 within intracellular compartment. In contrast, the combined exposure to both GYY4137 and metformin overwhelms cancer cells with lactate over-production exceeding its expulsion rate. Together, GYY4137 and simvastatin or metformin synergize to induce intracellular hyper-acidification-mediated cancer cell death.

Development of an in vitro photosafety evaluation method utilizing intracellular ROS production in THP-1 cells.

Science.gov (United States)

Toyoda, Akemi; Itagaki, Hiroshi

2018-01-01

Photoreactive compounds that may experience exposure to ultraviolet (UV) radiation can lead to the intracellular production of reactive oxygen species (ROS), which may cause phototoxic and photoallergenic responses. Here, we developed a novel in vitro photosafety assay and investigated whether it could be used to predict phototoxicity and photosensitivity by measuring changes in intracellular ROS production. THP-1 cells that had previously taken up 5-(and-6)-carboxy-2',7'-difluorodihydrofluorescein diacetate (carboxy-H 2 DFFDA), a ROS-sensitive fluorescent reagent, were exposed to photoreactive substances such as phototoxic and photoallergenic materials and then subjected to with UV-A irradiation (5 J/cm 2 ). The fluorescence intensity was subsequently measured using a flow cytometer, and the intracellular ROS production was calculated. A statistically significant increase in ROS following treatment with photoreactive substances was observed in cells irradiated with UV-A. In contrast, no significant increase was observed for non-photoreactive substances in comparison to the control solution. Next, to confirm the impact of intracellular ROS on the photosensitive response, changes in CD86 and CD54 expression were measured following quencher addition during the photo human cell line activation test (photo h-CLAT). The results confirmed the reduction of CD86 and CD54 expression in response to photoallergenic substances following quencher addition. Together, these findings suggest that intracellular ROS production is involved in photosensitizing reactions. Therefore, we suggest that the developed method utilizing intracellular ROS production as an index may be useful as a novel in vitro evaluation tool for photoreactive substances.

Microtubule Destabilizer KIF2A Undergoes Distinct Site-Specific Phosphorylation Cascades that Differentially Affect Neuronal Morphogenesis

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Tadayuki Ogawa

2015-09-01

Full Text Available Neurons exhibit dynamic structural changes in response to extracellular stimuli. Microtubules (MTs provide rapid and dramatic cytoskeletal changes within the structural framework. However, the molecular mechanisms and signaling networks underlying MT dynamics remain unknown. Here, we have applied a comprehensive and quantitative phospho-analysis of the MT destabilizer KIF2A to elucidate the regulatory mechanisms of MT dynamics within neurons in response to extracellular signals. Interestingly, we identified two different sets of KIF2A phosphorylation profiles that accelerate (A-type and brake (B-type the MT depolymerization activity of KIF2A. Brain-derived neurotrophic factor (BDNF stimulates PAK1 and CDK5 kinases, which decrease the MT depolymerizing activity of KIF2A through B-type phosphorylation, resulting in enhanced outgrowth of neural processes. In contrast, lysophosphatidic acid (LPA induces ROCK2 kinase, which suppresses neurite outgrowth from round cells via A-type phosphorylation. We propose that these two mutually exclusive forms of KIF2A phosphorylation differentially regulate neuronal morphogenesis during development.

SLAP displays tumour suppressor functions in colorectal cancer via destabilization of the SRC substrate EPHA2

Science.gov (United States)

Naudin, Cécile; Sirvent, Audrey; Leroy, Cédric; Larive, Romain; Simon, Valérie; Pannequin, Julie; Bourgaux, Jean-François; Pierre, Josiane; Robert, Bruno; Hollande, Frédéric; Roche, Serge

2014-01-01

The adaptor SLAP is a negative regulator of receptor signalling in immune cells but its role in human cancer is ill defined. Here we report that SLAP is abundantly expressed in healthy epithelial intestine but strongly downregulated in 50% of colorectal cancer. SLAP overexpression suppresses cell tumorigenicity and invasiveness while SLAP silencing enhances these transforming properties. Mechanistically, SLAP controls SRC/EPHA2/AKT signalling via destabilization of the SRC substrate and receptor tyrosine kinase EPHA2. This activity is independent from CBL but requires SLAP SH3 interaction with the ubiquitination factor UBE4A and SLAP SH2 interaction with pTyr594-EPHA2. SRC phosphorylates EPHA2 on Tyr594, thus creating a feedback loop that promotes EPHA2 destruction and thereby self-regulates its transforming potential. SLAP silencing enhances SRC oncogenicity and sensitizes colorectal tumour cells to SRC inhibitors. Collectively, these data establish a tumour-suppressive role for SLAP in colorectal cancer and a mechanism of SRC oncogenic induction through stabilization of its cognate substrates.

Intracellular localization of Na + /H + antiporter from Malus zumi ...

African Journals Online (AJOL)

In this study, we examined the intracellular localization of the product of Na+/H+ antiporter gene (MzNHX1) cloned from Malus zumi. Analysis using yeast cells expressing a fusion protein of MzNHX1 and green fluorescent protein confirmed the localization of MzNHX1 on the tonoplast.

Destabilization of low mode number Alfven modes in a tokamak by energetic or alpha particles

International Nuclear Information System (INIS)

Tsang, K.T.; Sigmar, D.J.; Whitson, J.C.

1980-12-01

With the inclusion of finite Larmor radius effects in the shear Alfven eigenmode equation, the continuous Alfven spectrum, which has been extensively discussed in ideal magnetohydrodynamics, is removed. Neutrally stable, discrete radial eigenmodes appear in the absence of sources of free energy and dissipation. Alpha (or energetic) particle toroidal drifts destabilize these modes, provided the particles are faster than the Alfven speed. Although the electron Landu resonance contributes to damping, a stability study of the parametric variation of the energy and the density scale length of the energetic particles shows that modes with low radial mode numbers remain unstable in most cases. Since the alpha particles are concentrated in the center of the plasma, this drift-type instability suggests anomalous helium ash diffusion. Indeed, it is shown that stochasticity of alpha orbits due to the overlapping of radially neighboring Alfven resonances is induced at low amplitudes, e/sub i//sup approx./phi/T/sub i/ greater than or equal to 0.05, implying a diffusion coefficient D/sub r//sup α/ greater than or equal to 4.4 x 10 3 cm 2 /s

Destabilizing effect of time-dependent oblique magnetic field on magnetic fluids streaming in porous media.

Science.gov (United States)

El-Dib, Yusry O; Ghaly, Ahmed Y

2004-01-01

The present work studies Kelvin-Helmholtz waves propagating between two magnetic fluids. The system is composed of two semi-infinite magnetic fluids streaming throughout porous media. The system is influenced by an oblique magnetic field. The solution of the linearized equations of motion under the boundary conditions leads to deriving the Mathieu equation governing the interfacial displacement and having complex coefficients. The stability criteria are discussed theoretically and numerically, from which stability diagrams are obtained. Regions of stability and instability are identified for the magnetic fields versus the wavenumber. It is found that the increase of the fluid density ratio, the fluid velocity ratio, the upper viscosity, and the lower porous permeability play a stabilizing role in the stability behavior in the presence of an oscillating vertical magnetic field or in the presence of an oscillating tangential magnetic field. The increase of the fluid viscosity plays a stabilizing role and can be used to retard the destabilizing influence for the vertical magnetic field. Dual roles are observed for the fluid velocity in the stability criteria. It is found that the field frequency plays against the constant part for the magnetic field.

Monitoring Intracellular Redox Changes in Ozone-exposed airway epithelial cells

Science.gov (United States)

Background: The toxicity of many compounds involves oxidative injury to cells. Direct assessment of mechanistic events involved in xenobiotic-induced oxidative stress is not easily achievable. Development of genetically-encoded probes designed for monitoring intracellular redox s...

A Bombesin-Shepherdin Radioconjugate Designed for Combined Extra- and Intracellular Targeting

Directory of Open Access Journals (Sweden)

Christiane A. Fischer

2014-05-01

Full Text Available Radiolabeled peptides which target tumor-specific membrane structures of cancer cells represent a promising class of targeted radiopharmaceuticals for the diagnosis and therapy of cancer. A potential drawback of a number of reported radiopeptides is the rapid washout of a substantial fraction of the initially delivered radioactivity from cancer cells and tumors. This renders the initial targeting effort in part futile and results in a lower imaging quality and efficacy of the radiotracer than achievable. We are investigating the combination of internalizing radiopeptides with molecular entities specific for an intracellular target. By enabling intracellular interactions of the radioconjugate, we aim at reducing/decelerating the externalization of radioactivity from cancer cells. Using the “click-to-chelate” approach, the 99mTc-tricarbonyl core as a reporter probe for single-photon emission computed tomography (SPECT was combined with the binding sequence of bombesin for extracellular targeting of the gastrin-releasing peptide receptor (GRP-r and peptidic inhibitors of the cytosolic heat shock 90 protein (Hsp90 for intracellular targeting. Receptor-specific uptake of the multifunctional radioconjugate could be confirmed, however, the cellular washout of radioactivity was not improved. We assume that either endosomal trapping or lysosomal degradation of the radioconjugate is accountable for these observations.

Transient fluctuations of intracellular zinc ions in cell proliferation

Energy Technology Data Exchange (ETDEWEB)

Li, Yuan [Division of Human Nutrition, Department of Preventive Medicine and Community Health, The University of Texas Medical Branch, Galveston, TX 77555 (United States); Maret, Wolfgang, E-mail: womaret@utmb.edu [Division of Human Nutrition, Department of Preventive Medicine and Community Health, The University of Texas Medical Branch, Galveston, TX 77555 (United States); Department of Anesthesiology, The University of Texas Medical Branch, Galveston, TX 77555 (United States)

2009-08-15

Zinc is essential for cell proliferation, differentiation, and viability. When zinc becomes limited for cultured cells, DNA synthesis ceases and the cell cycle is arrested. The molecular mechanisms of actions of zinc are believed to involve changes in the availability of zinc(II) ions (Zn{sup 2+}). By employing a fluorescent Zn{sup 2+} probe, FluoZin-3 acetoxymethyl ester, intracellular Zn{sup 2+} concentrations were measured in undifferentiated and in nerve growth factor (NGF)-differentiated rat pheochromocytoma (PC12) cells. Intracellular Zn{sup 2+} concentrations are pico- to nanomolar in PC12 cells and are higher in the differentiated than in the undifferentiated cells. When following cellular Zn{sup 2+} concentrations for 48 h after the removal of serum, a condition that is known to cause cell cycle arrest, Zn{sup 2+} concentrations decrease after 30 min but, remarkably, increase after 1 h, and then decrease again to about one half of the initial concentration. Cell proliferation, measured by an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, decreases after both serum starvation and zinc chelation. Two peaks of Zn{sup 2+} concentrations occur within one cell cycle: one early in the G1 phase and the other in the late G1/S phase. Thus, fluctuations of intracellular Zn{sup 2+} concentrations and established modulation of phosphorylation signaling, via an inhibition of protein tyrosine phosphatases at commensurately low Zn{sup 2+} concentrations, suggest a role for Zn{sup 2+} in the control of the cell cycle. Interventions targeted at these picomolar Zn{sup 2+} fluctuations may be a way of controlling cell growth in hyperplasia, neoplasia, and diseases associated with aberrant differentiation.

Intracellular Retention of ABL Kinase Inhibitors Determines Commitment to Apoptosis in CML Cells

Science.gov (United States)

Dziadosz, Marek; Schnöder, Tina; Heidel, Florian; Schemionek, Mirle; Melo, Junia V.; Kindler, Thomas; Müller-Tidow, Carsten; Koschmieder, Steffen; Fischer, Thomas

2012-01-01

Clinical development of imatinib in CML established continuous target inhibition as a paradigm for successful tyrosine kinase inhibitor (TKI) therapy. However, recent reports suggested that transient potent target inhibition of BCR-ABL by high-dose TKI (HD-TKI) pulse-exposure is sufficient to irreversibly commit cells to apoptosis. Here, we report a novel mechanism of prolonged intracellular TKI activity upon HD-TKI pulse-exposure (imatinib, dasatinib) in BCR-ABL-positive cells. Comprehensive mechanistic exploration revealed dramatic intracellular accumulation of TKIs which closely correlated with induction of apoptosis. Cells were rescued from apoptosis upon HD-TKI pulse either by repetitive drug wash-out or by overexpression of ABC-family drug transporters. Inhibition of ABCB1 restored sensitivity to HD-TKI pulse-exposure. Thus, our data provide evidence that intracellular drug retention crucially determines biological activity of imatinib and dasatinib. These studies may refine our current thinking on critical requirements of TKI dose and duration of target inhibition for biological activity of TKIs. PMID:22815843

Intracellular trafficking of superparamagnetic iron oxide nanoparticles conjugated with TAT peptide: 3-dimensional electron tomography analysis

International Nuclear Information System (INIS)

Nair, Baiju G.; Fukuda, Takahiro; Mizuki, Toru; Hanajiri, Tatsuro; Maekawa, Toru

2012-01-01

Highlights: ► We study the intracellular localisation of TAT-SPIONs using 3-D electron tomography. ► 3-D images of TAT-SPIONs in a cell are clearly shown. ► Release of TAT-SPIONs from endocytic vesicles into the cytoplasm is clearly shown. -- Abstract: Internalisation of nanoparticles conjugated with cell penetrating peptides is a promising approach to various drug delivery applications. Cell penetrating peptides such as transactivating transcriptional activator (TAT) peptides derived from HIV-1 proteins are effective intracellular delivery vectors for a wide range of nanoparticles and pharmaceutical agents thanks to their amicable ability to enter cells and minimum cytotoxicity. Although different mechanisms of intracellular uptake and localisation have been proposed for TAT conjugated nanoparticles, it is necessary to visualise the particles on a 3-D plane in order to investigate the actual intracellular uptake and localisation. Here, we study the intracellular localisation and trafficking of TAT peptide conjugated superparamagnetic ion oxide nanoparticles (TAT-SPIONs) using 3-D electron tomography. 3-D tomograms clearly show the location of TAT-SPIONs in a cell and their slow release from the endocytic vesicles into the cytoplasm. The present methodology may well be utilised for further investigations of the behaviours of nanoparticles in cells and eventually for the development of nano drug delivery systems.

Allyl Isothiocyanate Inhibits Actin-Dependent Intracellular Transport in Arabidopsis thaliana

Directory of Open Access Journals (Sweden)

Bjørnar Sporsheim

2015-12-01

Full Text Available Volatile allyl isothiocyanate (AITC derives from the biodegradation of the glucosinolate sinigrin and has been associated with growth inhibition in several plants, including the model plant Arabidopsis thaliana. However, the underlying cellular mechanisms of this feature remain scarcely investigated in plants. In this study, we present evidence of an AITC-induced inhibition of actin-dependent intracellular transport in A. thaliana. A transgenic line of A. thaliana expressing yellow fluorescent protein (YFP-tagged actin filaments was used to show attenuation of actin filament movement by AITC. This appeared gradually in a time- and dose-dependent manner and resulted in actin filaments appearing close to static. Further, we employed four transgenic lines with YFP-fusion proteins labeling the Golgi apparatus, endoplasmic reticulum (ER, vacuoles and peroxisomes to demonstrate an AITC-induced inhibition of actin-dependent intracellular transport of or, in these structures, consistent with the decline in actin filament movement. Furthermore, the morphologies of actin filaments, ER and vacuoles appeared aberrant following AITC-exposure. However, AITC-treated seedlings of all transgenic lines tested displayed morphologies and intracellular movements similar to that of the corresponding untreated and control-treated plants, following overnight incubation in an AITC-absent environment, indicating that AITC-induced decline in actin-related movements is a reversible process. These findings provide novel insights into the cellular events in plant cells following exposure to AITC, which may further expose clues to the physiological significance of the glucosinolate-myrosinase system.

Horizontal Transmission of Intracellular Insect Symbionts via Plants

Directory of Open Access Journals (Sweden)

Ewa Chrostek

2017-11-01

Full Text Available Experimental evidence is accumulating that endosymbionts of phytophagous insects may transmit horizontally via plants. Intracellular symbionts known for manipulating insect reproduction and altering fitness (Rickettsia, Cardinium, Wolbachia, and bacterial parasite of the leafhopper Euscelidius variegatus have been found to travel from infected insects into plants. Other insects, either of the same or different species can acquire the symbiont from the plant through feeding, and in some cases transfer it to their progeny. These reports prompt many questions regarding how intracellular insect symbionts are delivered to plants and how they affect them. Are symbionts passively transported along the insect-plant-insect path, or do they actively participate in the process? How widespread are these interactions? How does symbiont presence influence the plant? And what conditions are required for the new infection to establish in an insect? From an ecological, evolutionary, and applied perspective, this mode of horizontal transmission could have profound implications if occurring frequently enough or if new stable symbiont infections are established. Transmission of symbionts through plants likely represents an underappreciated means of infection, both in terms of symbiont epidemiology and the movement of symbionts to new host species.

Exceptional recurrence of flank destabilizations in the recent activity of the Colima volcanic complex, Mexico; Recurrence exceptionnelle de destabilisations de flanc dans l`activite recente du complexe volcanique du Colima, Mexique

Energy Technology Data Exchange (ETDEWEB)

Komorowski, J C [IPGP, (Mexico); Siebe, C [Institut de Geofisica, UNAM (Mexico); Rodriguez, S [Institut de Geologia, UNAM (Mexico); Cortes, A; Navarro, C; Gavilanes, J C

1997-12-31

This short paper reports on new {sup 14}C datings of debris flow units from the Nevado de Colima and Fuego de Colima volcanoes in Mexico. These new datings in connection with a detailed stratigraphic study in the deep canyons around the volcanoes has revealed an exceptional recurrence of flank destabilizations of the Fuego de Colima during the last 45000 years. The cumulated volume of debris in the whole Colima massif is estimated to 60-100 km{sup 3}. The correlation between Landsat satellite pictures and the distribution and age of the debris flows shows that both volcanoes are made of several post-destabilization remaining structures, and that both volcanoes were active and simultaneously collapsed 18500 years ago. The numerous fluvial-lacustrine sequences intercalated between the successive flows indicate that the debris flow were partially sedimented under water and could have led to catastrophic tsunamis towards the Pacific coast. Implications of this work are important because a population of more than 200000 inhabitants is living in a zone covered by several debris flows. (J.S.).

Exceptional recurrence of flank destabilizations in the recent activity of the Colima volcanic complex, Mexico; Recurrence exceptionnelle de destabilisations de flanc dans l`activite recente du complexe volcanique du Colima, Mexique

Energy Technology Data Exchange (ETDEWEB)

Komorowski, J.C. [IPGP, (Mexico); Siebe, C. [Institut de Geofisica, UNAM (Mexico); Rodriguez, S. [Institut de Geologia, UNAM (Mexico); Cortes, A.; Navarro, C.; Gavilanes, J.C.

1996-12-31

This short paper reports on new {sup 14}C datings of debris flow units from the Nevado de Colima and Fuego de Colima volcanoes in Mexico. These new datings in connection with a detailed stratigraphic study in the deep canyons around the volcanoes has revealed an exceptional recurrence of flank destabilizations of the Fuego de Colima during the last 45000 years. The cumulated volume of debris in the whole Colima massif is estimated to 60-100 km{sup 3}. The correlation between Landsat satellite pictures and the distribution and age of the debris flows shows that both volcanoes are made of several post-destabilization remaining structures, and that both volcanoes were active and simultaneously collapsed 18500 years ago. The numerous fluvial-lacustrine sequences intercalated between the successive flows indicate that the debris flow were partially sedimented under water and could have led to catastrophic tsunamis towards the Pacific coast. Implications of this work are important because a population of more than 200000 inhabitants is living in a zone covered by several debris flows. (J.S.).

Intracellular disposition of chitosan nanoparticles in macrophages: intracellular uptake, exocytosis, and intercellular transport

Directory of Open Access Journals (Sweden)

Jiang LQ

2017-08-01

Full Text Available Li Qun Jiang,1 Ting Yu Wang,1 Thomas J Webster,2 Hua-Jian Duan,1 Jing Ying Qiu,1 Zi Ming Zhao,1 Xiao Xing Yin,1,* Chun Li Zheng3,* 1Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, School of Pharmacy, Xuzhou Medical University, Xuzhou, People’s Republic of China; 2Department of Chemical Engineering, Northeastern University, Boston, MA, USA; 3School of Pharmacy, China Pharmaceutical University, Nanjing, People’s Republic of China *These authors contributed equally to this work Abstract: Biodegradable nanomaterials have been widely used in numerous medical fields. To further improve such efforts, this study focused on the intracellular disposition of chitosan nanoparticles (CsNPs in macrophages, a primary cell of the mononuclear phagocyte system (MPS. Such interactions with the MPS determine the nanoparticle retention time in the body and consequently play a significant role in their own clinical safety. In this study, various dye-labeled CsNPs (about 250 nm were prepared, and a murine macrophage cell line (RAW 264.7 was selected as a model macrophage. The results showed two mechanisms of macrophage incorporation of CsNPs, ie, a clathrin-mediated endocytosis pathway (the primary and phagocytosis. Following internalization, the particles partly dissociated in the cells, indicating cellular digestion of the nanoparticles. It was proved that, after intracellular uptake, a large proportion of CsNPs were exocytosed within 24 h; this excretion induced a decrease in fluorescence intensity in cells by 69%, with the remaining particles possessing difficulty being cleared. Exocytosis could be inhibited by both wortmannin and vacuolin-1, indicating that CsNP uptake was mediated by lysosomal and multivesicular body pathways, and after exocytosis, the reuptake of CsNPs by neighboring cells was verified by further experiments. This study, thus, elucidated the fate of CsNPs in macrophages as well as identified cellular disposition

Trapped fast particle destabilization of internal kink mode for the locally flattened q-profile with an inflection point

Energy Technology Data Exchange (ETDEWEB)

Wang, Xian-Qu [Institute of Fusion Science, School of Physical Science and Technology, Southwest Jiaotong University, Chengdu, Sichuan 610031 (China); Zhang, Rui-Bin; Meng, Guo [State Key Lab of Nuclear Physics and Technology, School of Physics, Peking University, Beijing 100871 (China)

2016-07-15

The destabilization of ideal internal kink modes by trapped fast particles in tokamak plasmas with a “shoulder”-like equilibrium current is investigated. It is found that energetic particle branch of the mode is unstable with the driving of fast-particle precession drifts and corresponds to a precessional fishbone. The mode with a low stability threshold is also more easily excited than the conventional precessional fishbone. This is different from earlier studies for the same equilibrium in which the magnetohydrodynamic (MHD) branch of the mode is stable. Furthermore, the stability and characteristic frequency of the mode are analyzed by solving the dispersion relation and comparing with the conventional fishbone. The results suggest that an equilibrium with a locally flattened q-profile, may be modified by localized current drive (or bootstrap current, etc.), is prone to the onset of the precessional fishbone branch of the mode.

The developmental outcomes of P0-mediated ARGONAUTE destabilization in tomato.

Science.gov (United States)

Hendelman, Anat; Kravchik, Michael; Stav, Ran; Zik, Moriyah; Lugassi, Nitsan; Arazi, Tzahi

2013-01-01

The plant protein ARGONAUTE1 (AGO1) functions in multiple RNA-silencing pathways, including those of microRNAs, key regulators of growth and development. Genetic analysis of ago1 mutants with informative defects has provided valuable insights into AGO1's biological functions. Tomato encodes two AGO1 homologs (SlAGO1s), but mutants have not been described to date. To analyze SlAGO1s' involvement in development, we confirmed that both undergo decay in the presence of the Polerovirus silencing suppressor P0 and produce a transgenic responder line (OP:P0HA) that, upon transactivation, expresses P0 C-terminally fused to a hemagglutinin (HA) tag (P0HA) and destabilizes SlAGO1s at the site of expression. By crossing OP:P0HA with a battery of driver lines, constitutive as well as organ- and stage-specific SlAGO1 downregulation was induced in the F1 progeny. Activated plants exhibited various developmental phenotypes that partially overlapped with those of Arabidopsis ago1 mutants. Plants that constitutively expressed P0HA had reduced SlAGO1 levels and increased accumulation of miRNA targets, indicating compromised SlAGO1-mediated silencing. Consistent with this, they exhibited pleiotropic morphological defects and their growth was arrested post-germination. Transactivation of P0HA in young leaf and floral organ primordia dramatically modified corresponding organ morphology, including the radialization of leaflets, petals and anthers, suggesting that SlAGO1s' activities are required for normal lateral organ development and polarity. Overall, our results suggest that the OP:P0HA responder line can serve as a valuable tool to suppress SlAGO1 silencing pathways in tomato. The suppression of additional SlAGOs by P0HA and its contribution to the observed phenotypes awaits investigation.

Fully glutathione degradable waterborne polyurethane nanocarriers: Preparation, redox-sensitivity, and triggered intracellular drug release

Energy Technology Data Exchange (ETDEWEB)

Omrani, Ismail; Babanejad, Niloofar; Shendi, Hasan Kashef; Nabid, Mohammad Reza, E-mail: m-nabid@sbu.ac.ir

2017-01-01

Polyurethanes are important class of biomaterials that are extensively used in medical devices. In spite of their easy synthesis, polyurethanes that are fully degradable in response to the intracellular reducing environment are less explored for controlled drug delivery. Herein, a novel glutathione degradable waterborne polyurethane (WPU) nanocarrier for redox triggered intracellular delivery of a model lipophilic anticancer drug, doxorubicin (DOX) is reported. The WPU was prepared from polyaddition reaction of isophorone diisocyanate (IPDI) and a novel linear polyester polyol involving disulfide linkage, disulfide labeled chain extender, dimethylolpropionic acid (DMPA) using dibutyltin dilaurate (DBTDL) as a catalyst. The resulting polyurethane self-assembles into nanocarrier in water. The dynamic light scattering (DLS) measurements and scanning electron microscope (SEM) revealed fast swelling and disruption of nanocarriers under an intracellular reduction-mimicking environment. The in vitro release studies showed that DOX was released in a controlled and redox-dependent manner. MTT assays showed that DOX-loaded WPU had a high in vitro antitumor activity in both HDF noncancer cells and MCF- 7 cancer cells. In addition, it is found that the blank WPU nanocarriers are nontoxic to HDF and MCF-7 cells even at a high concentration of 2 mg/mL. Hence, nanocarriers based on disulfide labeled WPU have appeared as a new class of biocompatible and redox-degradable nanovehicle for efficient intracellular drug delivery. - Highlights: • A novel fully glutathione degradable waterborne polyurethane was developed. • The waterborne nanocarrier with disulfide bonds in both hard and soft segment were developed for redox-triggered intracellular delivery of DOX. • The polyester diol bearing disulfide bonds in the backbone was prepared by a polycondensation polymerization reaction.

Cathepsin-Mediated Cleavage of Peptides from Peptide Amphiphiles Leads to Enhanced Intracellular Peptide Accumulation

Energy Technology Data Exchange (ETDEWEB)

Acar, Handan [Institute; Department; Samaeekia, Ravand [Institute; Department; Schnorenberg, Mathew R. [Institute; Department; Medical; Sasmal, Dibyendu K. [Institute; Huang, Jun [Institute; Tirrell, Matthew V. [Institute; Institute; LaBelle, James L. [Department

2017-08-24

Peptides synthesized in the likeness of their native interaction domain(s) are natural choices to target protein protein interactions (PPIs) due to their fidelity of orthostatic contact points between binding partners. Despite therapeutic promise, intracellular delivery of biofunctional peptides at concentrations necessary for efficacy remains a formidable challenge. Peptide amphiphiles (PAs) provide a facile method of intracellular delivery and stabilization of bioactive peptides. PAs consisting of biofunctional peptide headgroups linked to hydrophobic alkyl lipid-like tails prevent peptide hydrolysis and proteolysis in circulation, and PA monomers are internalized via endocytosis. However, endocytotic sequestration and steric hindrance from the lipid tail are two major mechanisms that limit PA efficacy to target intracellular PPIs. To address these problems, we have constructed a PA platform consisting of cathepsin-B cleavable PAs in which a selective p53-based inhibitory peptide is cleaved from its lipid tail within endosomes, allowing for intracellular peptide accumulation and extracellular recycling of the lipid moiety. We monitor for cleavage and follow individual PA components in real time using a resonance energy transfer (FRET)-based tracking system. Using this platform, components in real time using a Forster we provide a better understanding and quantification of cellular internalization, trafficking, and endosomal cleavage of PAs and of the ultimate fates of each component.

A simple model for solvation in mixed solvents. Applications to the stabilization and destabilization of macromolecular structures.

Science.gov (United States)

Schellman, J A

1990-08-31

. Examination of the few studies which have been reported in the literature shows that most of the qualitative features of the stabilization of proteins by sugars and their destabilization by urea and guanidinium chloride are faithfully represented with the model. This includes maxima in the free energy of stabilization and destabilization, decreased and zero selective interaction at high concentrations, etc. These phenomena had no prior explanation. Deficiencies in the model as a representation of solvation in aqueous solution are discussed in the appendix.

LDL Receptors as Gateways for Intracellular Porphyrin Uptake

International Nuclear Information System (INIS)

Novick, S.; Laster, B.; Quastel, M.

2004-01-01

Boronated compounds are currently being studied for possible use in Boron Neutron Capture Therapy (BNCT). We found that one of these agents, BOPP (tetrakis-carborane-carboxylate, esters of 2,4-bis (a,b- dihydroxyethyl) deuteroporphyrin IX), could also be labeled with indium (In-BOPP) and, therefore, could also be used potentially to transport high Z atoms into tumor cell DNA for AET (Auger Electron Therapy). In order to assess the uptake of these agents into cells, the role of the LDL receptor in the intracellular accumulation of BOPP and In-BOPP was investigated. Pre-incubation of V-79 Chinese hamster cells in medium containing delipidized fetal bovine serum (FBS) markedly increased the subsequent uptake of intracellular boron transported by both BOPP and In-BOPP when compared with cells that had been pre-incubated with medium containing 10% normal FBS (lipidized). The increased uptake was characterized by elevated levels of receptor, and greater affinity was shown for both BOPP and In-BOPP, although less marked with the latter. Positive cooperativity was demonstrated by sigmoid saturation curves, Scatchard analysis and Hill plots. Increasing the amount of LDL in the incubation medium had a relatively small effect on the total accumulation of either indium or boron atoms inside the cell. Furthermore, chemical acetylation of LDL did not decrease the intracellular uptake of either boron or indium transported by BOPP or In-BOPP. It is thus concluded that BOPP and In-BOPP preferentially enter the cells directly by way of the LDL receptor and that only a small fraction of these molecules are transported into the cells indirectly using serum LDLs as their carriers. These data suggest a novel way of bringing greater amounts of boron and indium (and perhaps other agents) into tissues. Porphyrins can be used to transport different agents into tumor cells because they are tumor affinic molecules. Tumors express a higher number of LDL receptors than do most normal tissues

Galectin-3 guides intracellular trafficking of some human serotransferrin glycoforms

DEFF Research Database (Denmark)

Carlsson, Carl Michael; Bengtson, Per; Cucak, Helena

2013-01-01

these transferrin glycoforms differently after preloading with exogenously added galectin-3. In all, this study provides the first evidence of a functional role for transferrin glycans, in intracellular trafficking after uptake. Moreover, the galectin-3 bound glycoform increased in cancer, suggesting...

Weakened Intracellular Zn2+-Buffering in the Aged Dentate Gyrus and Its Involvement in Erasure of Maintained LTP.

Science.gov (United States)

Takeda, Atsushi; Tamano, Haruna; Murakami, Taku; Nakada, Hiroyuki; Minamino, Tatsuya; Koike, Yuta

2018-05-01

Memory is lost by the increased influx of extracellular Zn 2+ into neurons. It is possible that intracellular Zn 2+ dynamics is modified even at non-zincergic medial perforant pathway-dentate granule cell synapses along with aging and that vulnerability to the modification is linked to age-related cognitive decline. To examine these possibilities, vulnerability of long-term potentiation (LTP) maintenance, which underlies memory retention, to modification of synaptic Zn 2+ dynamics was compared between young and aged rats. The influx of extracellular Zn 2+ into dentate granule cells was increased in aged rats after injection of high K + into the dentate gyrus, but not in young rats. This increase impaired maintained LTP in aged rats. However, the impairment was rescued by co-injection of CaEDTA, an extracellular Zn 2+ chelator, or CNQX, an AMPA receptor antagonist, which suppressed the Zn 2+ influx. Maintained LTP was also impaired in aged rats after injection of ZnAF-2DA into the dentate gyrus that chelates intracellular Zn 2+ , but not in young rats. Interestingly, the capacity of chelating intracellular Zn 2+ with intracellular ZnAF-2 was almost lost in the aged dentate gyrus 2 h after injection of ZnAF-2DA into the dentate gyrus, suggesting that intracellular Zn 2+ -buffering is weakened in the aged dentate gyrus, compared to the young dentate gyrus. In the dentate gyrus of aged rats, maintained LTP is more vulnerable to modification of intracellular Zn 2+ dynamics than in young rats, probably due to weakened intracellular Zn 2+ -buffering.

Mesurements of intracellular ATP provide new insight into the regulation of glycolysis in the yeast Saccharomyces cerevisiae

DEFF Research Database (Denmark)

Ytting, Cecilie Karkov; Fuglsang, Anja Thoe; Hiltunen, J. Kalervo

2012-01-01

Glycolysis in the yeast Saccharomyces cerevisiae exhibits temporal oscillation under anaerobic or semianaerobic conditions. Previous evidence indicated that at least two membrane-bound ATPases, the mitochondrial F0F1 ATPase and the plasma membrane P-type ATPase (Pma1p), were important in regulating...... of the temporal behaviour of intracellular ATP in a yeast strain with oscillating glycolysis showed that, in addition to oscillation in intracellular ATP, there is an overall slow decrease in intracellular ATP because the ATP consumption rate exceeds the ATP production in glycolysis. Measurements of the temporal...... activity is under strict control. In the absence of glucose ATPase activity is switched off, and the intracellular ATP concentration is high. When glucose is added to the cells the ATP concentration starts to decrease, because ATP consumption exceeds ATP production by glycolysis. Finally, when glucose...

Gold-coated polydimethylsiloxane microwells for high-throughput electrochemiluminescence analysis of intracellular glucose at single cells.

Science.gov (United States)

Xia, Juan; Zhou, Junyu; Zhang, Ronggui; Jiang, Dechen; Jiang, Depeng

2018-06-04

In this communication, a gold-coated polydimethylsiloxane (PDMS) chip with cell-sized microwells was prepared through a stamping and spraying process that was applied directly for high-throughput electrochemiluminescence (ECL) analysis of intracellular glucose at single cells. As compared with the previous multiple-step fabrication of photoresist-based microwells on the electrode, the preparation process is simple and offers fresh electrode surface for higher luminescence intensity. More luminescence intensity was recorded from cell-retained microwells than that at the planar region among the microwells that was correlated with the content of intracellular glucose. The successful monitoring of intracellular glucose at single cells using this PDMS chip will provide an alternative strategy for high-throughput single-cell analysis. Graphical abstract ᅟ.

Intracellular trafficking of the β-secretase and processing of amyloid precursor protein.

Science.gov (United States)

Zhi, Pei; Chia, Pei Zhi Cheryl; Chia, Cheryl; Gleeson, Paul A

2011-09-01

The main component of the amyloid plaques found in the brains of those with Alzheimer's disease (AD) is a polymerized form of the β-amyloid peptide (Aβ) and is considered to play a central role in the pathogenesis of this neurodegenerative disorder. Aβ is derived from the proteolytic processing of the amyloid precursor protein (APP). Beta site APP-cleaving enzyme, BACE1 (also known as β-secretase) is a membrane-bound aspartyl protease responsible for the initial step in the generation of Aβ peptide and is thus a prime target for therapeutic intervention. Substantive evidence now indicates that the processing of APP by BACE1 is regulated by the intracellular sorting of the enzyme and, moreover, perturbations in these intracellular trafficking pathways have been linked to late-onset AD. In this review, we highlight the recent advances in the understanding of the regulation of the intracellular sorting of BACE1 and APP and illustrate why the trafficking of these cargos represent a key issue for understanding the membrane-mediated events associated with the generation of the neurotoxic Aβ products in AD. Copyright © 2011 International Union of Biochemistry and Molecular Biology, Inc.

Activation of a Neospora caninum EGFR-Like Kinase Facilitates Intracellular Parasite Proliferation

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Xiaoxia Jin

2017-10-01

Full Text Available The Apicomplexan parasite Neospora caninum, an obligate intracellular protozoan, causes serious diseases in a number of mammalian species, especially in cattle. Infection with N. caninum is associated with abortions in both dairy and beef cattle worldwide which have a major economic impact on the cattle industry. However, the mechanism by which N. caninum proliferates within host cells is poorly understood. Epidermal growth factor receptor (EGFR is a protein kinase ubiquitously expressed, present on cell surfaces in numerous species, which has been confirmed to be essential in signal transduction involved in cell growth, proliferation, survival, and many other intracellular processes. However, the presence of EGFR in N. caninum and its role in N. caninum proliferation remain unclear. In the present study, we identified a putative EGFR-like kinase in N. caninum, which could be activated in tachyzoites by infection or treatment with rNcMIC3 [containing four epidermal growth factor (EGF domains] or human EGF. Blockade of EGFR-like in tachyzoites by AG1478 significantly reduced parasite proliferation in host cells. Our data suggested that the activation of tachyzoite EGFR-like might facilitate the intracellular proliferation of N. caninum.

Intracellular Delivery of Nanobodies for Imaging of Target Proteins in Live Cells.

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Röder, Ruth; Helma, Jonas; Preiß, Tobias; Rädler, Joachim O; Leonhardt, Heinrich; Wagner, Ernst

2017-01-01

Cytosolic delivery of nanobodies for molecular target binding and fluorescent labeling in living cells. Fluorescently labeled nanobodies were formulated with sixteen different sequence-defined oligoaminoamides. The delivery of formulated anti-GFP nanobodies into different target protein-containing HeLa cell lines was investigated by flow cytometry and fluorescence microscopy. Nanoparticle formation was analyzed by fluorescence correlation spectroscopy. The initial oligomer screen identified two cationizable four-arm structured oligomers (734, 735) which mediate intracellular nanobody delivery in a receptor-independent (734) or folate receptor facilitated (735) process. The presence of disulfide-forming cysteines in the oligomers was found critical for the formation of stable protein nanoparticles of around 20 nm diameter. Delivery of labeled GFP nanobodies or lamin nanobodies to their cellular targets was demonstrated by fluorescence microscopy including time lapse studies. Two sequence-defined oligoaminoamides with or without folate for receptor targeting were identified as effective carriers for intracellular nanobody delivery, as exemplified by GFP or lamin binding in living cells. Due to the conserved nanobody core structure, the methods should be applicable for a broad range of nanobodies directed to different intracellular targets.

Intracellular localisation of proteins to specific cellular areas by nanocapsule mediated delivery.

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Wang, Huabin; Chen, Ligang; Sun, Xianchao; Fu, Ailing

2017-09-01

Nanocapsules are promising carriers with great potential for intracellular protein transport. Although many studies have intended to improve cell uptake efficacy, there is an increasing interest in understanding of subcellular distribution of cargoes inside cells, which is essential for purposeful delivery of biomolecules into specific sites within cells. Herein, we interrogate the intracellular localisation of exogenous proteins, including fluorescein isothiocyanate (FITC)-labelled bovine serum albumin (BSA) and green fluorescent protein (GFP), mediated by specially designed nanocapsules. The results show that the designed nanocapsules can deliver the two types of fluorescent proteins into different cellular destinations (cytosol, nucleus or the whole cell), depending on the composition of nanocapsules. Meanwhile, several impact factors that influence the distribution of proteins in cells have also been investigated, and the results suggest that the localisation of capsule-mediated proteins in cells is strongly affected by the surface properties of nanocapsules, the types of stabilisers and proteins, and environmental temperatures. The rational control of intracellular localised delivery of exogenous proteins as we demonstrated in this study might open new avenues to obtain desired magnitude of drug effects for modulating cell activity.

Accumulation of intra-cellular polyphosphate in Chlorella vulgaris cells is related to indole-3-acetic acid produced by Azospirillum brasilense.

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Meza, Beatriz; de-Bashan, Luz E; Hernandez, Juan-Pablo; Bashan, Yoav

2015-06-01

Accumulation of intra-cellular phosphate, as polyphosphate, was measured when the microalga Chlorella vulgaris was immobilized in alginate with either of two wild-type strains of the microalgae growth-promoting bacterium Azospirillum brasilense or their corresponding IAA-attenuated mutants. Wild type strains of A. brasilense induced higher amounts of intra-cellular phosphate in Chlorella than their respective mutants. Calculations comparing intra-cellular phosphate accumulation by culture or net accumulation by the cell and the amount of IAA that was produced by each of these strains revealed that higher IAA was linked to higher accumulations of intra-cellular phosphate. Application of four levels of exogenous IAA reported for A. brasilense and their IAA-attenuated mutants to cultures of C. vulgaris enhanced accumulation of intra-cellular phosphate; the higher the content of IAA per culture or per single cell, the higher was the amount of accumulated phosphate. When an IAA-attenuated mutant was complemented with exogenous IAA, accumulation of intra-cellular phosphate at the culture level was even higher than phosphate accumulation with the respective wild type strains. When calculating the net accumulation of intra-cellular phosphate in the complementation experiment, net intra-cellular phosphate induced by the IAA-attenuated mutant was completely restored and was similar to the wild strains. We propose that IAA produced by A. brasilense is linked to polyphosphate accumulation in C. vulgaris. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Intracellular trafficking of superparamagnetic iron oxide nanoparticles conjugated with TAT peptide: 3-dimensional electron tomography analysis

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Nair, Baiju G.; Fukuda, Takahiro; Mizuki, Toru; Hanajiri, Tatsuro [Bio-Nano Electronics Research Centre, Toyo University, Saitama 350-8585 (Japan); Maekawa, Toru, E-mail: maekawa@toyo.jp [Bio-Nano Electronics Research Centre, Toyo University, Saitama 350-8585 (Japan)

2012-05-18

Highlights: Black-Right-Pointing-Pointer We study the intracellular localisation of TAT-SPIONs using 3-D electron tomography. Black-Right-Pointing-Pointer 3-D images of TAT-SPIONs in a cell are clearly shown. Black-Right-Pointing-Pointer Release of TAT-SPIONs from endocytic vesicles into the cytoplasm is clearly shown. -- Abstract: Internalisation of nanoparticles conjugated with cell penetrating peptides is a promising approach to various drug delivery applications. Cell penetrating peptides such as transactivating transcriptional activator (TAT) peptides derived from HIV-1 proteins are effective intracellular delivery vectors for a wide range of nanoparticles and pharmaceutical agents thanks to their amicable ability to enter cells and minimum cytotoxicity. Although different mechanisms of intracellular uptake and localisation have been proposed for TAT conjugated nanoparticles, it is necessary to visualise the particles on a 3-D plane in order to investigate the actual intracellular uptake and localisation. Here, we study the intracellular localisation and trafficking of TAT peptide conjugated superparamagnetic ion oxide nanoparticles (TAT-SPIONs) using 3-D electron tomography. 3-D tomograms clearly show the location of TAT-SPIONs in a cell and their slow release from the endocytic vesicles into the cytoplasm. The present methodology may well be utilised for further investigations of the behaviours of nanoparticles in cells and eventually for the development of nano drug delivery systems.

Involvement of intracellular Zn2+ signaling in LTP at perforant pathway-CA1 pyramidal cell synapse.

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Tamano, Haruna; Nishio, Ryusuke; Takeda, Atsushi

2017-07-01

Physiological significance of synaptic Zn 2+ signaling was examined at perforant pathway-CA1 pyramidal cell synapses. In vivo long-term potentiation (LTP) at perforant pathway-CA1 pyramidal cell synapses was induced using a recording electrode attached to a microdialysis probe and the recording region was locally perfused with artificial cerebrospinal fluid (ACSF) via the microdialysis probe. Perforant pathway LTP was not attenuated under perfusion with CaEDTA (10 mM), an extracellular Zn 2+ chelator, but attenuated under perfusion with ZnAF-2DA (50 μM), an intracellular Zn 2+ chelator, suggesting that intracellular Zn 2+ signaling is required for perforant pathway LTP. Even in rat brain slices bathed in CaEDTA in ACSF, intracellular Zn 2+ level, which was measured with intracellular ZnAF-2, was increased in the stratum lacunosum-moleculare where perforant pathway-CA1 pyramidal cell synapses were contained after tetanic stimulation. These results suggest that intracellular Zn 2+ signaling, which originates in internal stores/proteins, is involved in LTP at perforant pathway-CA1 pyramidal cell synapses. Because the influx of extracellular Zn 2+ , which originates in presynaptic Zn 2+ release, is involved in LTP at Schaffer collateral-CA1 pyramidal cell synapses, synapse-dependent Zn 2+ dynamics may be involved in plasticity of postsynaptic CA1 pyramidal cells. © 2017 Wiley Periodicals, Inc.

Brucella abortus Induces a Warburg Shift in Host Metabolism That Is Linked to Enhanced Intracellular Survival of the Pathogen.

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Czyż, Daniel M; Willett, Jonathan W; Crosson, Sean

2017-08-01

Intracellular bacterial pathogens exploit host cell resources to replicate and survive inside the host. Targeting these host systems is one promising approach to developing novel antimicrobials to treat intracellular infections. We show that human macrophage-like cells infected with Brucella abortus undergo a metabolic shift characterized by attenuated tricarboxylic acid cycle metabolism, reduced amino acid consumption, altered mitochondrial localization, and increased lactate production. This shift to an aerobic glycolytic state resembles the Warburg effect, a change in energy production that is well described in cancer cells and also occurs in activated inflammatory cells. B. abortus efficiently uses lactic acid as its sole carbon and energy source and requires the ability to metabolize lactate for normal survival in human macrophage-like cells. We demonstrate that chemical inhibitors of host glycolysis and lactate production do not affect in vitro growth of B. abortus in axenic culture but decrease its survival in the intracellular niche. Our data support a model in which infection shifts host metabolism to a Warburg-like state, and B. abortus uses this change in metabolism to promote intracellular survival. Pharmacological perturbation of these features of host cell metabolism may be a useful strategy to inhibit infection by intracellular pathogens. IMPORTANCE Brucella spp. are intracellular bacterial pathogens that cause disease in a range of mammals, including livestock. Transmission from livestock to humans is common and can lead to chronic human disease. Human macrophage-like cells infected with Brucella abortus undergo a Warburg-like metabolic shift to an aerobic glycolytic state where the host cells produce lactic acid and have reduced amino acid catabolism. We provide evidence that the pathogen can exploit this change in host metabolism to support growth and survival in the intracellular niche. Drugs that inhibit this shift in host cell metabolism