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Sample records for skin fibroblast lines

  1. Radiation-induced radioresistance in a normal human skin fibroblast cell line

    International Nuclear Information System (INIS)

    Damage to DNA resulting from ionizing radiation exposure may compromise cellular viability, contribute to malignancy and possibly to senescence. Experimental evidence indicates that several mechanisms exist to repair DNA damage but the molecular events underlying these types of repair in mammalian cells remain obscure. Using the human skin fibroblast line, AG1522, the authors have monitored the end-points of cellular survival, colony growth rate and micronucleus formation to asses adaptation to ionizing radiation. (author). 4 refs., 4 figs

  2. Potentially lethal damage repair in cell lines of radioresistant human tumours and normal skin fibroblasts

    International Nuclear Information System (INIS)

    Radiation cell survival data were obtained in vitro for three cell lines isolated from human tumours traditionally considered to be radioresistant-two melanomas and one osteosarcoma-as well as from a diploid skin fibroblast cell line. One melanoma cell line was much more radioresistant than the other, while the osteosarcoma and fibroblast cell lines were more radiosensitive than either. For cells growing exponentially, little potentially lethal damage repair (PLDR) could be demonstrated by comparing survival data for cells in which subculture was delayed by 6 h with those sub-cultured immediately after treatment. For the malignant cells in plateau phase, which in these cells might be better termed 'slowed growth phase', since an appreciable fraction of the cells are still cycling, a small amount of PLDR was observed, but not as much as reported by other investigators in the literature. The normal fibroblasts, which achieved a truer plateau phase in terms of noncycling cells, showed a significantly larger amount of PLDR than the tumour cells. (author)

  3. Radiosensitivity in vitro of human soft tissue sarcoma cell lines and skin fibroblasts derived from the same patients

    International Nuclear Information System (INIS)

    Skin fibroblast cell strains and tumour cell lines were established from 12 patients with various types of soft tissue neoplasms, and radiation survival curve parameters were measured in vitro. Soft tissue sarcoma cells were consistently more sensitive to X-irradiation than fibroblasts isolated from the same patient, and were also more sensitive as a group than cell lines derived from 34 other human tumours. There was a general correlation in radiosensitivity between fibroblasts and tumour cells derived from the same patient, indicating that some component of tumour cell sensitivity may relate to genetic factors in the host. Such genetic factors, however, do not explain all of the heterogeneity in tumour cell response. The response of soft tissue sarcoma in vivo may be dependent on complex radiomodifying factors other than inherent radiation sensitivity, thus making it difficult to predict clinical outcome by use of assays which use survival of irradiated tumour cell lines in vitro as an endpoint. (author)

  4. Critical role of GSH in Sulfur Mustard-induced Oxidative Stress and Cytotoxicity in Human Skin Fibroblast Cell Line

    Directory of Open Access Journals (Sweden)

    Ali Beman Zaree Mahmoudabad

    2008-01-01

    Full Text Available In this study the role of glutathione (GSH in sulfur mustard -induced oxidative stress and cytotoxicity, in human skin fibroblast cell line (HF2FF was evaluated. Sulfur mustard-induced superoxide radical and hydrogen peroxide formation were evaluated by determination of superoxide dismutase and catalase activity in cell lysate. The cytotoxicity of sulfur mustard was estimated by lactate dehydrogenase leakage. The intracellular GSH content was modulated by N-acetylcysteine (NAC, a GSH precursor, and buthionine sulfoximine (BSO, a specific GSH synthesis inhibitor. It was found that sulfur mustard exposure led to a dose-and time-dependent decrease in GSH content in HF2FF cells. NAC increased intracellular GSH level and protected the cells against sulfur mustard-induced reactive oxygen species formation and lactate dehydrogenase leakage. In contrast, buthionine sulfoximine pretreatment depleted cellular GSH and enhanced the susceptibility of HF2FF to the cytotoxic effects of sulfur mustard. These results indicated that GSH plays a critical role in protecting HF2FF cell line against sulfur mustar-induced cell injury, most probably through its antioxidant activity.

  5. Lymphoblastoid lines and skin fibroblasts from patients with tuberous sclerosis are abnormally sensitive to ionizing radiation and to a radiomimetic chemical

    International Nuclear Information System (INIS)

    Lymphoblastoid lines, derived by transforming peripheral blood lymphocytes with Epstein-Barr virus, and skin fibroblast lines were established from two patients with tuberous sclerosis. The number of viable lymphoblastoid cells was determined by their ability to exclude the vital dye trypan blue after their irradiation with x-rays or 254 nm ultraviolet light. The growth of fibroblasts was determined by their ability to form colonies after treatment with the radiomimetic, DNA-damaging chemical N-methyl-N'-nitro-N-nitrosoguanidine. The tuberous sclerosis lymphoblastoid lines were hypersensitive to x-rays but had normal sensitivity to the ultraviolet radiation. The tuberous sclerosis fibroblast lines were hypersensitive to the N-methyl-N'-nitro-N-nitrosoguanidine. The hypersensitivity of tuberous sclerosis cells to x-rays and to N-methyl-N'-nitro-N-nitrosoguanidine is believed to reflect defective repair of DNA damaged by these agents and may provide the basis for in vitro, including prenatal, diagnostic tests for tuberous sclerosis

  6. Alteration of Skin Properties with Autologous Dermal Fibroblasts

    OpenAIRE

    Rajesh L. Thangapazham; Thomas N. Darling; Jon Meyerle

    2014-01-01

    Dermal fibroblasts are mesenchymal cells found between the skin epidermis and subcutaneous tissue. They are primarily responsible for synthesizing collagen and glycosaminoglycans; components of extracellular matrix supporting the structural integrity of the skin. Dermal fibroblasts play a pivotal role in cutaneous wound healing and skin repair. Preclinical studies suggest wider applications of dermal fibroblasts ranging from skin based indications to non-skin tissue regeneration in tendon re...

  7. Alteration of Skin Properties with Autologous Dermal Fibroblasts

    Directory of Open Access Journals (Sweden)

    Rajesh L. Thangapazham

    2014-05-01

    Full Text Available Dermal fibroblasts are mesenchymal cells found between the skin epidermis and subcutaneous tissue. They are primarily responsible for synthesizing collagen and glycosaminoglycans; components of extracellular matrix supporting the structural integrity of the skin. Dermal fibroblasts play a pivotal role in cutaneous wound healing and skin repair. Preclinical studies suggest wider applications of dermal fibroblasts ranging from skin based indications to non-skin tissue regeneration in tendon repair. One clinical application for autologous dermal fibroblasts has been approved by the Food and Drug Administration (FDA while others are in preclinical development or various stages of regulatory approval. In this context, we outline the role of fibroblasts in wound healing and discuss recent advances and the current development pipeline for cellular therapies using autologous dermal fibroblasts. The microanatomic and phenotypic differences of fibroblasts occupying particular locations within the skin are reviewed, emphasizing the therapeutic relevance of attributes exhibited by subpopulations of fibroblasts. Special focus is provided to fibroblast characteristics that define regional differences in skin, including the thick and hairless skin of the palms and soles as compared to hair-bearing skin. This regional specificity and functional identity of fibroblasts provides another platform for developing regional skin applications such as the induction of hair follicles in bald scalp or alteration of the phenotype of stump skin in amputees to better support their prosthetic devices.

  8. Cultures of cancer patient's skin tissue fibroblast and radiosensitivity assay

    International Nuclear Information System (INIS)

    In order to test the radiosensitivity of normal skin tissue, the authors cultured cancer patient's skin tissue fibroblast, surviving fraction experiment was employed to provide data for understanding of the different radiosensitivity among the cancer patients, Method: cancer patient's skin tissue fibroblast were cultured in vitro by the way of tar's attachment, cells were irradiated by graded doses of γ-ray , cell dose response experiment was used to test the radiosensitivity of cell. Result: Cancer patient's skin fibroblast could be propagated and passaged by the method of culture in vitro. Radiosensitivity are different among the various cancer patient's skin tissue fibroblasts

  9. Protective and restorative effects of a Commiphora mukul gum resin and triheptanoin preparation on the CCL-110 skin fibroblast cell line.

    Science.gov (United States)

    Ramachandran, C; Quirin, K-W; Resek, A; Melnick, S J

    2012-04-01

    Coenzyme Q10 (CoQ10) is a major ingredient in skin care products because of its anti-wrinkle effects, although it has some side effects especially at higher amounts. In this study, we compare the anti-wrinkle related properties of CoQ10 and a proprietary Commiphora mukul gum resin (guggul) and triheptanoin preparation (GU-TC7). GU-TC7 is prepared with a supercritical CO₂-co-solvent extraction with ethanol, standardized to 2% guggulsterones and triheptanoin, a triglyceride composed of three 7-carbon fatty acids. Treatment of CCL-110 skin fibroblasts with GU-TC7 demonstrates a mild proliferative effect compared to CoQ10 and increased type I collagen synthesis. Additionally, GU-TC7 inhibited matrix metalloproteinase-1 (MMP-1) expression in a dose-dependent manner at 20-100 μg mL⁻¹ and inhibited human elastase expression by more than 50% as compared to no elastase inhibition with CoQ10 treatment. These results suggest that GU-TC7 possesses properties that are applicable to the treatment of wrinkles and may be considered for its further evaluation in skin care products. PMID:22084831

  10. Differences in motility pattern between human buccal fibroblasts and periodontal and skin fibroblasts

    DEFF Research Database (Denmark)

    Lepekhin, Eugene; Grøn, Birgitte; Berezin, Vladimir; Bock, Elisabeth; Dabelsteen, Erik

    2002-01-01

    Migration of fibroblasts from surrounding normal tissue into the wound bed is an important requirement for successful wound healing. This study investigated the motility pattern of buccal, periodontal and skin fibroblasts to determine whether differences in the wound healing efficiency at these s...... and motility pattern amongst the three fibroblast types could not be explained by differences in secretion of extracellular matrix components and are therefore believed to reflect phenotypic differences amongst fibroblast subpopulations....

  11. Development of a full-thickness human skin equivalent in vitro model derived from TERT-immortalized keratinocytes and fibroblasts

    OpenAIRE

    Reijnders, C.M.A.; Lier, van, R.; Roffel, S.; Kramer, D.; Scheper, R. J.; Gibbs, S.

    2015-01-01

    Currently, human skin equivalents (HSEs) used for in vitro assays (e.g., for wound healing) make use of primary human skin cells. Limitations of primary keratinocytes and fibroblasts include availability of donor skin and donor variation. The use of physiologically relevant cell lines could solve these limitations. The aim was to develop a fully differentiated HSE constructed entirely from human skin cell lines, which could be applied for in vitro wound-healing assays. Skin equivalents were c...

  12. Development of a Full-Thickness Human Skin Equivalent In Vitro Model Derived from TERT-Immortalized Keratinocytes and Fibroblasts

    OpenAIRE

    Reijnders, Christianne M.A.; van Lier, Amanda; Roffel, Sanne; Kramer, Duco; Scheper, Rik J; Gibbs, Susan

    2015-01-01

    Currently, human skin equivalents (HSEs) used for in vitro assays (e.g., for wound healing) make use of primary human skin cells. Limitations of primary keratinocytes and fibroblasts include availability of donor skin and donor variation. The use of physiologically relevant cell lines could solve these limitations. The aim was to develop a fully differentiated HSE constructed entirely from human skin cell lines, which could be applied for in vitro wound-healing assays. Skin equivalents were c...

  13. The effects of human skin fibroblast monolayers on human sperm motility and mouse zygote development.

    Science.gov (United States)

    Wetzels, A M; Punt-Van der Zalm, A P; Bastiaans, B A; Janssen, B A; Goverde, H J; Rolland, R

    1992-07-01

    A new system for co-culture in human in-vitro fertilization (IVF), using human skin fibroblasts, is described and tested pre-clinically. The first test involved the development of 1-cell mouse embryos which exhibit the 2-cell developmental block in vitro. Passage through this block (pb1-ratio) was determined by the ratio of compacted morula stages on day 4 of incubation. For nine human skin cell lines tested (fetal, neonatal and adult), the pb1-ratio was approximately 0.45 (0.07 in culture medium alone; P less than 0.0005). At the compacted morula stage, a second developmental block was observed. The ratio of passing this block (pb2-ratio) was 0.70 +/- 0.09 on skin fibroblasts obtained from fetal or neonatal tissue. On fibroblasts from adult patients the pb2-ratio was 0.30 +/- 0.04 (P less than 0.0005). The second test examined the influence of skin fibroblasts from fetal or neonatal tissue on human sperm motility. After 24 h of incubation, all skin cell lines had a positive influence (P less than 0.01) on the percentage motility compared to culture medium alone. The curvilinear velocity was not significantly increased. From the results we conclude that (i) human skin fibroblasts (especially from fetal tissue) have a positive influence on the development of mouse embryos in vitro, (ii) there is a positive influence of human skin fibroblasts on the percentage motility of human spermatozoa, and (iii) a clinical trial of co-culture with human skin fibroblasts can be justified. PMID:1500485

  14. Treatment of Skin Avulsion Injuries with Basic Fibroblast Growth Factor

    OpenAIRE

    Hajime Matsumine, MD, PhD

    2015-01-01

    Summary: This report describes favorable outcomes in 9 patients with skin avulsion injuries of the extremities who underwent full-thickness skin grafting and basic fibroblast growth factor (bFGF) application. Following removal of contaminated subcutaneous fat tissue on the inside of skin, the avulsed skin was processed into a full-thickness skin graft, with as much of the skin used as possible irrespective of damage. Several drainage holes (5–10 mm in diameter) were made on the graft for drai...

  15. Testosterone metabolism of fibroblasts grown from prostatic carcinoma, benign prostatic hyperplasia and skin fibroblasts

    International Nuclear Information System (INIS)

    The metabolism of [1,2,6,7-3H]testosterone was assessed in fibroblast monolayers derived from tissue of 5 prostates with benign hyperplasia (BPH), 4 prostates with carcinoma (PC), and 3 biopsy samples of skin, 2 nongenital skin (NG) and 1 genital skin. The following metabolites could be identified: androstanedione androstenedione, dihydrotestosterone, androsterone, epiandrosterone, androstane-3 alpha, 17 beta-diol and androstane-3 beta, 17 beta-diol. Testosterone was metabolized much more rapidly in fibroblasts originating from prostatic tissue than in fibroblasts derived from NG. A significantly higher formation of 5 alpha-androstanes and 3 alpha-hydroxysteroids could be observed in fibroblasts from BPH as compared to PC. 17-ketosteroid formation exceeded 5 alpha-androstane formation in BPH, whereas 5 alpha-reduction was the predominant pathway in fibroblasts grown from PC and NG. Since testosterone metabolism in fibroblasts of prostatic origin therefore resembles in many aspects that in whole prostatic tissue, fibroblasts grown from prostatic tissues might be a valuable tool for further investigation of the pathogenesis of human BPH and PC

  16. "Short Take": Resistance of Skin Fibroblasts to Peroxide and UV Damage Predicts Hearing Loss in Aging Mice

    OpenAIRE

    Richard A. Miller; DOLAN, DAVID; Han, Melissa; Kohler, William; Schacht, Jochen

    2011-01-01

    Those mice whose skin-derived primary fibroblast cell lines resist lethal injury induced by hydrogen peroxide or UV light show lower age-related decline in hearing. Skin cell lines may provide an easily accessible surrogate index of intrinsic stress resistance that varies among individuals and influences the pace of neurosensory decline in aging mice.

  17. Extracellular depolymerization of hyaluronic acid in cultured human skin fibroblasts

    International Nuclear Information System (INIS)

    The chain length of [3H]hyaluronic acid synthesized by cultivating human skin fibroblasts in the presence of [3H]glucosamine was investigated. [3H]Hyaluronic acid obtained from the matrix fraction was excluded from a Sepharose CL-2B column irrespective of the incubation period, whereas that from the medium was depolymerized into a constant chain length (Mr = 40,000). The reducing and non-reducing terminals of the depolymerized hyaluronic acid were N-acetylglucosamine and glucuronic acid, respectively. Prolonged incubation produced no oligosaccharides as shown by examination of hyaluronidase digests, suggesting the presence of a novel endo-beta-N-acetylglucosaminidase in cultured human skin fibroblasts

  18. Proliferation index of camel skin fibroblast cells as nuclear donor

    International Nuclear Information System (INIS)

    Jaiselmeri is an excellent breed of riding camel, found in Jaiselmer and other adjoining districts of Western Rajasthan in India. Jaiselmeri camel like other pack animals are declining in India over the years due to increased mechanization and control of desert agriculture to some extent. The deep freezing technology on camel semen is poorly developed in India. The somatic cell technology has been developed at this Institute as an alternative tool of long-term conservation on endangered livestock breeds. For this study, samples of (0.25 cm2) skin tissue were collected from ear biopsy from elite male germplasm from National Research Centre on Camel, Bikaner. Skin tissues were cultured at 37 deg. C in Medium (DMEM+ Ham's F-12 nutritive mixture) supplemented with 10% fetal bovine serum, L-Glutamine and antibiotics in an incubator under 98% humidified and 5% Co2 atmosphere. The cell explants were visible from 12-16 days of culture. The cells were allowed to confluent in the TC flasks for additional 3-5 days till nearly 80% surface area is covered by the cells. The primary cells were harvested by usual trypsin-EDTA protocol. The cells were counted using Neubar's haemocytometer and cells were passaged subsequently. Since no reference values were available for camel skin fibroblasts, the present experiments were conducted to study the cell proliferation index, population doubling time, standard growth curve and cell viability using standard growth and MTT assays. It is shown that growth curves showed true sigmoid shape but a marked variation between the cell lines was observed. Moreover, cells, which grew faster attained plateau on day 6 while in slow growing cultures, the curve showed elevation even on day 8. This is probably due to non-availability of growing space for cells having faster growth rate. It was concluded that all animals do not produce karyoplast donors at equal rate or efficiency. Therefore, the growing cultures need to be compared with standard growth curve each time the cells are used as nuclear donor cells for cloning. Cell Proliferation Index: Cell multiplication rates vary considerably under different culture condition and slight change in environment or composition of medium may affect the proliferation of cells significantly. For camel skin fibroblast cells, the standard multiplication rate and the population doubling time was not known earlier. In order to study the proliferative indices of the growing cells using objective parameters, MTT assay was conducted. In this assay, the dividing and viable cells take up MTT [3- (4,5- dimethylthiozol- 2yl) 2,5 diphenyltetrazolium bromide] and a colour is developed. The intensity of colour is measured by ELISA reader at 540-570 nm. For this, 4000 cells per well were seeded in 96 well ELISA plate (flat bottom, Nunc) and cultured at 37 deg. C. First two rows of eight wells each were kept as negative and positive controls respectively. Rest of the 10 rows were kept as treatments. One row was harvested at an interval of 24 hours and adjoining row was treated with MTT solution for 4 hours. The MTT treated cells were fixed in 10% DMSO. Figures 2 and 3 show that the cell proliferation index both in terms of cell count and absorbance values in ELISA reader at appropriate wavelength was similar. From this study it is clear that MTT assay can give fairly accurate figures of cell proliferation rate of skin fibroblasts. Ploidy level: During long-term culture, the cells are likely to develop one or other type of chromosomal abnormalities. It must be ensured that the cells in different passages are checked for normal ploidy so that the viable clones can be developed from them. In order to see the utility of cells from Jaiselmeri camel as nuclear donor, the chromosomal profile was studied following the protocol described elsewhere. The 2N chromosomes up to passage No 4 (15th population doubling) was found to be normal (74XY) in 97% of the cells. From these preliminary studies it appears that camel skin fibroblast cells behave normally in culture and can serve as nuclear donors. These results will further help in camel cloning

  19. Extracellular depolymerization of hyaluronic acid in cultured human skin fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, T.; Takagaki, K.; Kubo, K.; Morikawa, A.; Tamura, S.; Endo, M. (Hirosaki Univ. School of Medicine (Japan))

    1990-10-15

    The chain length of ({sup 3}H)hyaluronic acid synthesized by cultivating human skin fibroblasts in the presence of ({sup 3}H)glucosamine was investigated. ({sup 3}H)Hyaluronic acid obtained from the matrix fraction was excluded from a Sepharose CL-2B column irrespective of the incubation period, whereas that from the medium was depolymerized into a constant chain length (Mr = 40,000). The reducing and non-reducing terminals of the depolymerized hyaluronic acid were N-acetylglucosamine and glucuronic acid, respectively. Prolonged incubation produced no oligosaccharides as shown by examination of hyaluronidase digests, suggesting the presence of a novel endo-beta-N-acetylglucosaminidase in cultured human skin fibroblasts.

  20. Effect of hormones and radiation on cultured human skin fibroblasts

    International Nuclear Information System (INIS)

    In order to elucidate the effects of hormones and radiation on skin fibroblasts in vitro, thyroxine (T4) and hydrocortisone were added either separately or in combination to cultures of human skin fibroblasts. In addition, other human skin fibroblast cultures were irradiated with 60Co-γ rays. The resulting effects of each treatment on the proliferation and morphology of the cells were investigated. There was a tendency for cell proliferation to be inhibited dose-dependently with an increase in the T4 concentration in the T4 group. Some of the cells of the T4 group were slightly flattened and there was an observable decrease in the number of fine villi and thread-type pseudopodia. Furthermore, morphologic changes in the cells such as a decrease and enlargement of the mitochondria and a decrease in the number of cristae were observed in the T4 group. A tendency for inhibition of the proliferation rate of the cells was observed in the radiation group. Immediately after irradiation of the cells, there was an increase in the number of heterogeneous residual bodies containing membrane components and amorphous materials and cells containing morphologically altered mitochondria were observed sporadically. (author)

  1. Treatment of Skin Avulsion Injuries with Basic Fibroblast Growth Factor

    Directory of Open Access Journals (Sweden)

    Hajime Matsumine, MD, PhD

    2015-04-01

    Full Text Available Summary: This report describes favorable outcomes in 9 patients with skin avulsion injuries of the extremities who underwent full-thickness skin grafting and basic fibroblast growth factor (bFGF application. Following removal of contaminated subcutaneous fat tissue on the inside of skin, the avulsed skin was processed into a full-thickness skin graft, with as much of the skin used as possible irrespective of damage. Several drainage holes (5–10 mm in diameter were made on the graft for drainage from the graft bed and to prevent seroma and hematoma formation. Genetically recombinant human bFGF was sprayed at a dose of 1 μg/cm2 onto the graft bed, which was then covered with the graft and sutured. Pressure immobilization with ointment gauzes and elastic bandages was administered for 1 week postoperatively, and the surface of the skin grafts that did not take was scraped away, preserving the revascularized dermal component on the debrided raw surface as much as possible. bFGF was sprayed again onto the debrided surface to promote epithelialization. Wound closure was achieved in all cases with conservative therapy. The surgical procedure was effective in preventing postoperative ulcer formation and scar contracture and resulted in wound healing with the formation of good-quality, flexible scars.

  2. Monomethylarsonous acid inhibited endogenous cholesterol biosynthesis in human skin fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Lei [Environmental Toxicology Graduate Program, University of California, Riverside, CA 92521-0403 (United States); Xiao, Yongsheng [Department of Chemistry, University of California, Riverside, CA 92521-0403 (United States); Wang, Yinsheng, E-mail: yinsheng.wang@ucr.edu [Environmental Toxicology Graduate Program, University of California, Riverside, CA 92521-0403 (United States); Department of Chemistry, University of California, Riverside, CA 92521-0403 (United States)

    2014-05-15

    Human exposure to arsenic in drinking water is a widespread public health concern, and such exposure is known to be associated with many human diseases. The detailed molecular mechanisms about how arsenic species contribute to the adverse human health effects, however, remain incompletely understood. Monomethylarsonous acid [MMA(III)] is a highly toxic and stable metabolite of inorganic arsenic. To exploit the mechanisms through which MMA(III) exerts its cytotoxic effect, we adopted a quantitative proteomic approach, by coupling stable isotope labeling by amino acids in cell culture (SILAC) with LC-MS/MS analysis, to examine the variation in the entire proteome of GM00637 human skin fibroblasts following acute MMA(III) exposure. Among the ∼ 6500 unique proteins quantified, ∼ 300 displayed significant changes in expression after exposure with 2 μM MMA(III) for 24 h. Subsequent analysis revealed the perturbation of de novo cholesterol biosynthesis, selenoprotein synthesis and Nrf2 pathways evoked by MMA(III) exposure. Particularly, MMA(III) treatment resulted in considerable down-regulation of several enzymes involved in cholesterol biosynthesis. In addition, real-time PCR analysis showed reduced mRNA levels of select genes in this pathway. Furthermore, MMA(III) exposure contributed to a distinct decline in cellular cholesterol content and significant growth inhibition of multiple cell lines, both of which could be restored by supplementation of cholesterol to the culture media. Collectively, the present study demonstrated that the cytotoxicity of MMA(III) may arise, at least in part, from the down-regulation of cholesterol biosynthesis enzymes and the resultant decrease of cellular cholesterol content. - Highlights: • MMA(III)-induced perturbation of the entire proteome of GM00637 cells is studied. • Quantitative proteomic approach revealed alterations of multiple cellular pathways. • MMA(III) inhibits de novo cholesterol biosynthesis. • MMA(III) perturbs Nrf2 pathway and selenoprotein synthesis.

  3. Monomethylarsonous acid inhibited endogenous cholesterol biosynthesis in human skin fibroblasts

    International Nuclear Information System (INIS)

    Human exposure to arsenic in drinking water is a widespread public health concern, and such exposure is known to be associated with many human diseases. The detailed molecular mechanisms about how arsenic species contribute to the adverse human health effects, however, remain incompletely understood. Monomethylarsonous acid [MMA(III)] is a highly toxic and stable metabolite of inorganic arsenic. To exploit the mechanisms through which MMA(III) exerts its cytotoxic effect, we adopted a quantitative proteomic approach, by coupling stable isotope labeling by amino acids in cell culture (SILAC) with LC-MS/MS analysis, to examine the variation in the entire proteome of GM00637 human skin fibroblasts following acute MMA(III) exposure. Among the ∼ 6500 unique proteins quantified, ∼ 300 displayed significant changes in expression after exposure with 2 μM MMA(III) for 24 h. Subsequent analysis revealed the perturbation of de novo cholesterol biosynthesis, selenoprotein synthesis and Nrf2 pathways evoked by MMA(III) exposure. Particularly, MMA(III) treatment resulted in considerable down-regulation of several enzymes involved in cholesterol biosynthesis. In addition, real-time PCR analysis showed reduced mRNA levels of select genes in this pathway. Furthermore, MMA(III) exposure contributed to a distinct decline in cellular cholesterol content and significant growth inhibition of multiple cell lines, both of which could be restored by supplementation of cholesterol to the culture media. Collectively, the present study demonstrated that the cytotoxicity of MMA(III) may arise, at least in part, from the down-regulation of cholesterol biosynthesis enzymes and the resultant decrease of cellular cholesterol content. - Highlights: • MMA(III)-induced perturbation of the entire proteome of GM00637 cells is studied. • Quantitative proteomic approach revealed alterations of multiple cellular pathways. • MMA(III) inhibits de novo cholesterol biosynthesis. • MMA(III) perturbs Nrf2 pathway and selenoprotein synthesis

  4. Low dose rate ionizing radiation induces increased growth capacities of d-deletion retinoblastoma skin fibroblasts

    International Nuclear Information System (INIS)

    Skin fibroblasts from normal children and three children with a 13q deletion retinoblastoma (Rb) were exposed to cumulative low doses of gamma rays. The typical response of normal donors was a reduction in the lifespan of irradiated fibroblasts, the precocity of the decline being inversely related to the dose received. In constrast, the lifespan of one Rb cell line (Rb1) was prolonged; irradiated cells with an increased growth potential showed a higher number of cells at confluency and more cells were entering DNA synthesis phase than in non-irradiated cells. Another Rb cell line (Rb2) demonstrated a normal lifespan following irradiation but foci were observed in irradiated cultures. Cytogenetic analysis revealed no selection of abnormal clones in these cell populations. The third Tb line examined (Rb3) responded like a normal cell line. It is suggested that irradiated skin fibroblasts derived from some patients with Rb are in certain cases able to express abnormal growth capacities which may be one of the manifestations of the high susceptibility of the individual's stromal cells to carcinogenic agents. (author)

  5. Tattoo ink nanoparticles in skin tissue and fibroblasts

    Science.gov (United States)

    Twigg, Peter C; Baker, Richard; Tobin, Desmond J

    2015-01-01

    Summary Tattooing has long been practised in various societies all around the world and is becoming increasingly common and widespread in the West. Tattoo ink suspensions unquestionably contain pigments composed of nanoparticles, i.e., particles of sub-100 nm dimensions. It is widely acknowledged that nanoparticles have higher levels of chemical activity than their larger particle equivalents. However, assessment of the toxicity of tattoo inks has been the subject of little research and ink manufacturers are not obliged to disclose the exact composition of their products. This study examines tattoo ink particles in two fundamental skin components at the nanometre level. We use atomic force microscopy and light microscopy to examine cryosections of tattooed skin, exploring the collagen fibril networks in the dermis that contain ink nanoparticles. Further, we culture fibroblasts in diluted tattoo ink to explore both the immediate impact of ink pigment on cell viability and also to observe the interaction between particles and the cells. PMID:26171294

  6. Tattoo ink nanoparticles in skin tissue and fibroblasts

    Directory of Open Access Journals (Sweden)

    Colin A. Grant

    2015-05-01

    Full Text Available Tattooing has long been practised in various societies all around the world and is becoming increasingly common and widespread in the West. Tattoo ink suspensions unquestionably contain pigments composed of nanoparticles, i.e., particles of sub-100 nm dimensions. It is widely acknowledged that nanoparticles have higher levels of chemical activity than their larger particle equivalents. However, assessment of the toxicity of tattoo inks has been the subject of little research and ink manufacturers are not obliged to disclose the exact composition of their products. This study examines tattoo ink particles in two fundamental skin components at the nanometre level. We use atomic force microscopy and light microscopy to examine cryosections of tattooed skin, exploring the collagen fibril networks in the dermis that contain ink nanoparticles. Further, we culture fibroblasts in diluted tattoo ink to explore both the immediate impact of ink pigment on cell viability and also to observe the interaction between particles and the cells.

  7. Fibroblast-mediated contraction in actinically exposed and actinically protected aging skin

    International Nuclear Information System (INIS)

    The changes in skin morphology over time are a consequence of both chronologic aging and the accumulation of environmental exposure. Through observation, we know that actinic radiation intensifies the apparent aging of skin. We have investigated the effects of aging and actinic radiation on the ability of fibroblasts to contract collagen-fibroblast lattices. Preauricular and postauricular skin samples were obtained from eight patients aged 49 to 74 undergoing rhytidectomy. The samples were kept separate, and the fibroblasts were grown in culture. Lattices constructed with preauricular fibroblasts consistently contracted more than lattices containing postauricular fibroblasts. The difference in amount of contraction in 7 days between sites was greatest for the younger patients and decreased linearly as donor age increased (r = -0.96). This difference may be due to preauricular fibroblasts losing their ability to contract a lattice as aging skin is exposed to more actinic radiation

  8. Exposure to Varying Strain Magnitudes Influences the Conversion of Normal Skin Fibroblasts Into Hypertrophic Scar Cells.

    Science.gov (United States)

    Kuang, Ruixia; Wang, Zhiguo; Xu, Quanchen; Cai, Xia; Liu, Tao

    2016-04-01

    Mechanical strain is a key contributor in the pathogenesis of hypertrophic scarring, whose optimal stretch magnitudes to initiate the differentiation of normal skin fibroblasts into aberrant fibroblasts phenotype remains largely unresolved. Influence of varying cyclic strain magnitudes on cultured human normal skin fibroblasts and its transformation into hypertrophic scar fibroblast-like phenotype is investigated in this study. Cultured fibroblasts isolated from hypertrophic scar and normal skin tissue were subjected to cyclic mechanical stretching under individual 10%, 15%, and 20% strain magnitudes at a frequency of 0.1 Hz for 24 hours. Stretched normal skin fibroblasts demonstrated significantly increased rates of cell proliferation, and also apparently oriented away nearly perpendicular to the applied stretching direction. Interestingly, the applied 10% strains magnitude resulted in a markedly enhanced cell proliferative ability compared with that of 20% strain magnitude. Parameters involving the mechanotransduction signaling, such as integrin β1 and P130Cas, were significantly improved at both mRNA and protein levels in the stretched normal skin fibroblasts, which was demonstrated in a negative magnitude-dependent manner. In addition, 10% strains magnitude triggered the highest expression levels of growth factor TGF-β1 and collagen matrix in stretched normal skin fibroblasts. Collectively, these results indicate that the 10% stretching magnitude, of the 3 strain magnitudes studied, is most effective for triggering the optimal mechanotransduction effects and biological responses inside cultured skin fibroblasts. The demonstrable conversion of normal skin fibroblasts into hypertrophic scar fibroblasts was also observed when 10% stretching magnitude was applied to cultured fibroblasts in vitro. PMID:26545222

  9. Lysosomal Storage Causes Cellular Dysfunction in Mucolipidosis II Skin Fibroblasts*

    Science.gov (United States)

    Otomo, Takanobu; Higaki, Katsumi; Nanba, Eiji; Ozono, Keiichi; Sakai, Norio

    2011-01-01

    Mucolipidosis II (ML-II) is a fatal inherited metabolic disease caused by deficiency of GlcNAc-phosphotransferase, which plays a role in generating the mannose 6-phosphate recognition marker on lysosomal enzymes. In ML-II, many lysosomal acid hydrolases are mistargeted out of cells, and lysosomes become filled with undigested substrates, which explains inclusion cell disease as an alternative name for this disease. In this study, we revealed various cellular phenotypes in ML-II skin fibroblasts. We quantitated phospholipid and cholesterol within cells and showed ∼2-fold accumulation in ML-II as compared with normal cells. Lysosomal pH of ML-II cells was higher than that of normal cells (5.29 ± 0.08 versus 4.79 ± 0.10, p < 0.001). The proliferated lysosomes in ML-II cells were accumulated ∼3-fold in amount as compared with normal cells. Intracellular logistics including endocytosis and mannose 6-phosphate receptor recycling were impaired in ML-II cells. To confirm whether these ML-II cellular phenotypes derive from deficient lysosomal acid hydrolases within lysosomes, we performed supplementation of lysosomal enzymes using a partially purified total enzyme mixture, which was derived from the conditioned culture medium of normal skin fibroblasts after NH4Cl treatment. This supplementation corrected all of the previously described ML-II phenotypes. In addition, the autophagic and mitochondrial impairment that we have previously reported improved, and inclusion bodies disappeared on electron micrography following total lysosomal enzyme supplementation. Our results indicate that various cellular phenotypes in ML-II are caused by the deficiency of many lysosomal enzymes and massive accumulation of undigested substrates. PMID:21846724

  10. Normal skin and hypertrophic scar fibroblasts differentially regulate collagen and fibronectin expression as well as mitochondrial membrane potential in response to basic fibroblast growth factor

    OpenAIRE

    Rui Song; Hui-Ning Bian; Wen Lai; Hua-De Chen; Ke-Seng Zhao

    2011-01-01

    Basic fibroblast growth factor (bFGF) regulates skin wound healing; however, the underlying mechanism remains to be defined. In the present study, we determined the effects of bFGF on the regulation of cell growth as well as collagen and fibronectin expression in fibroblasts from normal human skin and from hypertrophic scars. We then explored the involvement of mitochondria in mediating bFGF-inducedeffects on the fibroblasts. We isolated and cultivated normal and hypertrophic scar fibroblasts...

  11. Abnormal Collagen Metabolism in Cultured Skin Fibroblasts from Patients with Duchenne Muscular Dystrophy

    Science.gov (United States)

    Rodemann, H. Peter; Bayreuther, Klaus

    1984-08-01

    Total collagen synthesis is decreased by about 29% (P Duchenne muscular dystrophy (DMD) as compared with that in normal male skin fibroblasts in vitro. The reduction in collagen synthesis is associated with an approximately 2-fold increase in collagen degradation in DMD fibroblasts. Correlated to these alterations in the metabolism of collagen, DMD fibroblasts express a significantly higher hydroxyproline/proline ratio (DMD: 1.36-1.45; P < 0.01) than do normal fibroblasts (controls: 0.86-0.89). The increased hydroxylation of proline residues of collagen (composed of type I and type III) could be the cause for the enhanced degradation of collagen in DMD fibroblasts.

  12. Aging Alters Functionally Human Dermal Papillary Fibroblasts but Not Reticular Fibroblasts: A New View of Skin Morphogenesis and Aging

    OpenAIRE

    Mine, Solène; Fortunel, Nicolas O.; Pageon, Hervé; Asselineau, Daniel

    2008-01-01

    Understanding the contribution of the dermis in skin aging is a key question, since this tissue is particularly important for skin integrity, and because its properties can affect the epidermis. Characteristics of matched pairs of dermal papillary and reticular fibroblasts (Fp and Fr) were investigated throughout aging, comparing morphology, secretion of cytokines, MMPs/TIMPs, growth potential, and interaction with epidermal keratinocytes. We observed that Fp populations were characterized by...

  13. Mercury-induced micronuclei in skin fibroblasts of beluga whales

    Energy Technology Data Exchange (ETDEWEB)

    Gauthier, J.M.; Dubeau, H.; Rassart, E. [Univ. du Quebec, Montreal, Quebec (Canada). Dept. des Sciences Biologiques

    1998-12-01

    Beluga whales (Delphinapterus leucas) inhabiting the St. Lawrence estuary are highly contaminated with environmental pollutants and have a high incidence of cancer. Environmental contaminants may be partly responsible for the high incidence of cancer observed in this population. DNA damage plays an important role in the development of cancer. The micronuclei assay was used to test the genotoxic potential of mercury compounds in skin fibroblasts of an Arctic beluga whale. Both mercuric chloride (Hg) and methylmercury (MeHg) induced a highly significant dose-response increase of micronucleated cells. Statistically significant increases in micronucleated cells were observed for 0.5, 5, and 20 {micro}g/ml Hg and 0.05, 0.5, and 2 {micro}g/ml MeHg when compared to control cultures. Concentrations of 0.5, 5, and 20 {micro}g/ml Hg induced a two-, three- and fourfold increase of micronucleated cells, respectively. Treatment with MeHg was one order of magnitude more potent in inducing micronuclei and in inhibiting cell proliferation than Hg. Although results of this in vitro study do not imply that mercury compounds are involved in the etiology of cancer in St. Lawrence beluga whales, significant increases in micronuclei frequency were found at low concentrations of MeHg that are believed to be comparable to concentrations present in certain whales of this population.

  14. Phenotypic and functional changes in dermal primary fibroblasts isolated from intrinsically aged human skin.

    Science.gov (United States)

    Brun, Ccilia; Jean-Louis, Francette; Oddos, Thierry; Bagot, Martine; Bensussan, Armand; Michel, Laurence

    2016-02-01

    Dermal fibroblasts play a key role in maintaining skin homoeostasis by synthesizing and degrading extracellular matrix components. During ageing, they are subjected to changes, such as the loss of type I collagen expression and an increased synthesis of metalloproteinase I, leading to fragmentation of collagen fibrils with consequent reduction of the mechanical tension and defects of skin wound healing. Most information about fibroblast ageing was obtained from experiments performed on replicative-senescent dermal fibroblasts in vitro. However, the senescence status of fibroblasts isolated from intrinsically aged skins and its consequences on functionality need to be deeper investigated. Herein, we studied age-related phenotypic and functional alteration of fibroblasts from 'young' (50 years) donors. Our results brought evidence of the senescent status of 'old' fibroblasts by senescence associated ?-galactosidase (SA-?gal) positive staining and p16 expression. A PCR array focusing on senescence highlighted a subset of downregulated genes including cell cycle progression and ECM genes in 'old' fibroblasts as well as a subset of upregulated genes involved in senescence features. In 'old' fibroblasts, we measured a downregulation of proliferative and contractile capacities of migratory potential under PDGF stimulation and activation into myofibroblasts under TGF?. Old fibroblasts were also more sensitive to oxidative stress than 'young' ones. Of interest, downregulation of p16 expression partially reversed the senescent phenotype of 'old' fibroblasts but failed to restore their functional properties. In conclusion, our data brought evidence of phenotypic and functional differences between fibroblasts from young and intrinsically aged skin that may contribute to the alterations observed with ageing. PMID:26441147

  15. In vitro culture of skin fibroblast cells for potential cloning by nuclear transfer

    International Nuclear Information System (INIS)

    Donor cell lines were developed from skin tissue for the conservation of the endangered Jaiselmeri camel breed of India. Average cell proliferation rates varied from 0.82 to 0.69 in different passages, and population doubling time from 29.3 h to 34.8 h. Around 15 population doublings were accomplished during this culturing. Cell viability was 97 to 99% in different passages. Growth curves of cells from the JC-5 cell line reached a plateau on day 7, while the slower-growing cultures of JC-3 showed elevation even on day 10, possibly due to donor age differences. Cell proliferation rates by both cell count and MTT absorbance showed similar patterns, with a correlation coefficient of 0.79. MTT assay, a colorimetric method, can handle large samples in somatic cell cultures. Diploid chromosomal counts in passages 1, 3 and 5 were normal (2N=74, XY) in 97% of the cells. Occasional metaphase plates showed polyploidy. The present baseline data on standard growth curve, linear relationship in colorimetric assay for estimation of cell proliferation rate, and normal ploidy and karyological levels in camel skin fibroblast cells in multiplication could be useful in developing competent donor somatic cell lines for conservation now and revival of this camel breed by cloning in the future. (author)

  16. Fibroblast heterogeneity and its implications for engineering organotypic skin models in vitro.

    Science.gov (United States)

    Sriram, Gopu; Bigliardi, Paul Lorenz; Bigliardi-Qi, Mei

    2015-11-01

    Advances in cell culture methods, multidisciplinary research, clinical need to replace lost skin tissues and regulatory need to replace animal models with alternative test methods has led to development of three dimensional models of human skin. In general, these in vitro models of skin consist of keratinocytes cultured over fibroblast-populated dermal matrices. Accumulating evidences indicate that mesenchyme-derived signals are essential for epidermal morphogenesis, homeostasis and differentiation. Various studies show that fibroblasts isolated from different tissues in the body are dynamic in nature and are morphologically and functionally heterogeneous subpopulations. Further, these differences seem to be dictated by the local biological and physical microenvironment the fibroblasts reside resulting in "positional identity or memory". Furthermore, the heterogeneity among the fibroblasts play a critical role in scarless wound healing and complete restoration of native tissue architecture in fetus and oral mucosa; and excessive scar formation in diseased states like keloids and hypertrophic scars. In this review, we summarize current concepts about the heterogeneity among fibroblasts and their role in various wound healing environments. Further, we contemplate how the insights on fibroblast heterogeneity could be applied for the development of next generation organotypic skin models. PMID:26344860

  17. DNA repair in Bloom's syndrome skin fibroblasts after ultraviolet light irradiation

    International Nuclear Information System (INIS)

    Skin fibroblasts from a patient with Bloom's syndrome (86NoKi) were assayed for various DNA repair activities after ultraviolet light (UV) irradiation. Cultured fibroblasts as well as lymphocytes obtained from this patient showed a high frequency of spontaneous sister chromatid exchanges (SCEs). There was no significant difference between 86NoKi fibroblasts and skin fibroblasts from normal donors in the sensitivity to UV as measured by inactivation of colony forming activity, the capacity of host-cell reactivation (HCR) of UV-irradiated virus, and the amount of unscheduled DNA synthesis (UDS) after UV irradiation. However, the yield of UV-induced SCEs in 86NoKi cells was significantly higher than that in normal cells. (author)

  18. Deficiency of gamma-ray excision repair in skin fibroblasts from patients with Fanconi's anemia

    International Nuclear Information System (INIS)

    The capacity of preparations of skin fibroblasts from normal individuals and patients with Fanconi's anemia to excise gamma-ray products of the 5,6-dihydroxydihydrothymine type from exogenous DNA was investigated. The excision capacity of whole-cell homogenates of fibroblasts from two of four patients with Fanconi's anemia was substantially below normal. This repair deficiency was further pronounced in nuclear preparations from cells of the same two patients

  19. Distinct fibroblast lineages determine dermal architecture in skin development and repair

    Science.gov (United States)

    Driskell, Ryan R.; Simons, Ben D.; Charalambous, Marika; Ferron, Sacri R.; Herault, Yann; Pavlovic, Guillaume; Ferguson-Smith, Anne C.; Watt, Fiona M.

    2013-01-01

    Fibroblasts are the major mesenchymal cell type in connective tissue and deposit the collagen and elastic fibers of the extracellular matrix (ECM)1. Even within a single tissue fibroblasts exhibit remarkable functional diversity, but it is not known whether this reflects the existence of a differentiation hierarchy or is a response to different environmental factors. Here we show, using transplantation assays and lineage tracing, that the fibroblasts of skin connective tissue arise from two distinct lineages. One forms the upper dermis, including the dermal papilla that regulates hair growth and the arrector pili muscle (APM), which controls piloerection. The other forms the lower dermis, including the reticular fibroblasts that synthesise the bulk of the fibrillar ECM, and the pre-adipocytes and adipocytes of the hypodermis. The upper lineage is required for hair follicle formation. In wounded adult skin, the initial wave of dermal repair is mediated by the lower lineage and upper dermal fibroblasts are recruited only during re-epithelialisation. Epidermal beta-catenin activation stimulates expansion of the upper dermal lineage, rendering wounds permissive for hair follicle formation. Our findings explain why wounding is linked to formation of ECM-rich scar tissue that lacks hair follicles2-4. They also form a platform for discovering fibroblast lineages in other tissues and for examining fibroblast changes in ageing and disease. PMID:24336287

  20. Influence of caffeine and hyaluronicacid on collagen biosynthesis in human skin fibroblasts

    Directory of Open Access Journals (Sweden)

    Donejko M

    2014-10-01

    Full Text Available Magdalena Donejko,1 Andrzej Przylipiak,1 Edyta Rysiak,2 Katarzyna G?uszuk,2 Arkadiusz Sura?y?ski2 1Department of Esthetic Medicine, 2Department of Medicinal Chemistry, Faculty of Pharmacy, Medical University of Bia?ystok,Bia?ystok, Poland Aim: The aim of this study was to evaluate the effect of caffeine on collagen biosynthesis in human skin fibroblasts and the influence of hyaluronic acid (HA on this process. Materials and methods: Collagen, [3H]-thymidine incorporation, and prolidase activity were measured in confluent human skin fibroblast cultures that had been treated with 1, 2, and 5 mM caffeine and with caffeine and 500 g/mL HA. Western immunoblot analysis was performed to evaluate expression of 1-integrin receptor, insulin-like growth factor receptor phospho-Akt protein and mitogen-activated protein kinase (phospho-extracellular signal-regulated kinase. Results: Caffeine inhibited collagen biosynthesis in a dose-dependent manner. The mechanism of this process was found at the level of prolidase activity. Caffeine significantly inhibited the enzyme activity. The addition of HA had no effect on collagen biosynthesis or prolidase activity in fibroblasts incubated with caffeine. Caffeine also had an inhibitory effect on DNA biosynthesis. HA, however, did not have any significant effect on this process. The inhibition of the expression of 1-integrin and insulin-like growth factor receptor in fibroblasts incubated with the caffeine indicates a possible mechanism of inhibition of collagen biosynthesis. Conclusion: Caffeine reduces collagen synthesis in human cultured skin fibroblasts. HA did not have any significant protective effect on this process. This is the first study to our knowledge that reports caffeine-induced inhibition of collagen synthesis in human skin fibroblasts. Keywords: collagen, caffeine, hyaluronic acid, fibroblast

  1. Normal skin and hypertrophic scar fibroblasts differentially regulate collagen and fibronectin expression as well as mitochondrial membrane potential in response to basic fibroblast growth factor

    Directory of Open Access Journals (Sweden)

    Rui Song

    2011-05-01

    Full Text Available Basic fibroblast growth factor (bFGF regulates skin wound healing; however, the underlying mechanism remains to be defined. In the present study, we determined the effects of bFGF on the regulation of cell growth as well as collagen and fibronectin expression in fibroblasts from normal human skin and from hypertrophic scars. We then explored the involvement of mitochondria in mediating bFGF-inducedeffects on the fibroblasts. We isolated and cultivated normal and hypertrophic scar fibroblasts from tissue biopsies of patients who underwent plastic surgery for repairing hypertrophic scars. The fibroblasts were then treated with different concentrations of bFGF (ranging from 0.1 to 1000 ng/mL. The growth of hypertrophic scar fibroblasts became slower with selective inhibition of type I collagen production after exposure to bFGF. However, type III collagen expression was affected in both normal and hypertrophic scar fibroblasts. Moreover, fibronectin expression in the normal fibroblasts was up-regulated after bFGF treatment. bFGF (1000 ng/mL also induced mitochondrial depolarization in hypertrophic scar fibroblasts (P < 0.01. The cellular ATP level decreased in hypertrophic scar fibroblasts (P < 0.05, while it increased in the normal fibroblasts following treatment with bFGF (P < 0.01. These data suggest that bFGF has differential effects and mechanisms on fibroblasts of the normal skin and hypertrophic scars, indicating that bFGF may play a role in the early phase of skin wound healing and post-burn scar formation.

  2. DNA-repair after UV-irradiation in skin fibroblasts from patients with actinic keratosis

    International Nuclear Information System (INIS)

    Autoradiographic counting technique was utilized to measure the ultraviolet-induced unscheduled DNA synthesis of skin fibroblasts from 12 patients with chronic actinic keratosis and from 12 healthy donors of about the same age. In order to reveal a possible regional difference of DNA repair between the parts of the body ordinarily exposed and those parts unexposed to sunlight, two cell strains were used for each examined subject; one developed from the forehead skin and the other from the abdominal or axillary skin. Unscheduled DNA synthesis appeared depressed in actinic keratosis patients, as compared with controls. In all examined subjects, however, cell strains from exposed skin showed a DNA repair more active than cell strains from unexposed skin. These findings show that skin cancer may be promoted in actinic keratosis patients by a defect of DNA repair. The exalted DNA repair of chronically sun exposed skin is probably the consequence of a defensive process caused by enzymatic induction. (orig.)

  3. Possible role for metallothionein in the cellular defense mechanism against UVB irradiation in neonatal human skin fibroblasts

    International Nuclear Information System (INIS)

    The role of metallothionein (MT) in protecting skin cells against UVB irradiation was investigated. Fibroblast strains from normal adult (HS-K) and neonatal (NB1RGB) human skins as well as keratinocyte strains from human skin (SV40-HSK) and newborn Balb/c mouse skin (Pam 212) were exposed to UVB irradiation. (Author)

  4. Establishment of the first humpback whale fibroblast cell lines and their application in chemical risk assessment.

    Science.gov (United States)

    Burkard, Michael; Whitworth, Deanne; Schirmer, Kristin; Nash, Susan Bengtson

    2015-10-01

    This paper reports the first successful derivation and characterization of humpback whale fibroblast cell lines. Primary fibroblasts were isolated from the dermal connective tissue of skin biopsies, cultured at 37 C and 5% CO2 in the standard mammalian medium DMEM/F12 supplemented with 10% fetal bovine serum (FBS). Of nine initial biopsies, two cell lines were established from two different animals and designated HuWa1 and HuWa2. The cells have a stable karyotype with 2n=44, which has commonly been observed in other baleen whale species. Cells were verified as being fibroblasts based on their spindle-shaped morphology, adherence to plastic and positive immunoreaction to vimentin. Population doubling time was determined to be ?41 h and cells were successfully cryopreserved and thawed. To date, HuWa1 cells have been propagated 30 times. Cells proliferate at the tested temperatures, 30, 33.5 and 37 C, but show the highest rate of proliferation at 37 C. Short-term exposure to para,para'-dichlorodiphenyldichloroethylene (p,p'-DDE), a priority compound accumulating in southern hemisphere humpback whales, resulted in a concentration-dependent loss of cell viability. The effective concentration which caused a 50% reduction in HuWa1 cell viability (EC50 value) was approximately six times greater than the EC50 value for the same chemical measured with human dermal fibroblasts. HuWa1 exposed to a natural, p,p'-DDE-containing, chemical mixture extracted from whale blubber showed distinctively higher sensitivity than to p,p'-DDE alone. Thus, we provide the first cytotoxicological data for humpback whales and with establishment of the HuWa cell lines, a unique in vitro model for the study of the whales' sensitivity and cellular response to chemicals and other environmental stressors. PMID:26363275

  5. Sperm functional changes and fertilization in vitro in co-culture with human skin fibroblasts.

    Science.gov (United States)

    Wetzels, A M; Van der Auwera, I; Bastiaans, B A; Goverde, H J; Hollanders, H M; Hamilton, C J

    1995-01-01

    This study was undertaken to evaluate the effects of human skin fibroblast monolayers on human sperm function and fertilization in vitro. Sperm function was evaluated using the hamster oocyte penetration assay (HOPA) and zona binding assay (ZBA) in medium alone and in co-culture with human skin fibroblast monolayers and suspensions. The ZBA was also studied in fibroblast conditioned medium and in bovine oviduct cell monolayers and suspensions. Fertilization was measured both in in-vitro fertilization (IVF) couples with a normal semen analysis (first study; randomized) and in IVF couples with subnormal semen analysis (second study; each patient served as its own control). The HOPA results were not significantly different with or without fibroblasts. In all co-culture situations and in conditioned medium the ZBA scored significantly lower than medium alone. No significant differences with respect to IVF were observed between the co-culture and the control group in either study. The mean fertilization rate per patient was approximately 60% in the group with normal semen analysis and approximately 25% in the group with abnormal semen analysis. From this study we conclude that although co-culture with human skin fibroblasts and epithelial cells influences the results of some sperm function tests, it does not influence fertilization in vitro. PMID:7745043

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

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Hyun Sook; Kang, Ki Mun; Na, Jae Boem; Chai, Gyu Young [Gyeongsang National University College of Medicine, Jinju (Korea, Republic of); Lee, Sang Wook [University of Ulsan College of Medicine, Asan Medical Center, Seoul (Korea, Republic of)

    2006-09-15

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

  7. Enhanced biosynthesis of human skin collagenase in fibroblast cultures from recessive dystrophic epidermolysis bullosa.

    OpenAIRE

    Valle, K J; Bauer, E A

    1980-01-01

    Using a sensitive, specific immunoprecipitation method, the biosynthesis of human skin collagenase was studied in fibroblast cultures from patients with recessive dystrophic epidermolysis bullosa. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of solubilized immunoprecipitates showed two 3H-labeled procollagenase species that comigrated with those harvested from control cultures. Recessive dystrophic epidermolysis bullosa cultures accumulated increased amounts of collagenase. Both ...

  8. Complementation studies of isovaleric acidemia and glutaric aciduria type II using cultured skin fibroblasts.

    Science.gov (United States)

    Dubiel, B; Dabrowski, C; Wetts, R; Tanaka, K

    1983-01-01

    Using cultured skin fibroblasts, we studied the heterogeneity of inborn errors of leucine metabolism such as isovaleric acidemia (IVA), glutaric aciduria type II (GA II), and multiple carboxylase deficiency (MC). We first developed a simple macromolecular-labeling test to measure the ability of cells to oxidize [1-14C]isovaleric acid in situ in culture. Cells from two different lines were fused using polyethylene glycol, and the ability of the heterokaryons to oxidize [1-14C]isovaleric acid was tested by the macromolecular-labeling test. The MC line complemented with all 10 IVA lines tested; heterokaryons showed 99 +/- 68% more activity than the unfused mixture of component cells. GA II/IVA heterokaryons exhibited poor growth, but when the culture remained confluent, the GA II cells complemented with all six IVA lines tested, showing a 71 +/- 41% increase in activity. The relatively large standard deviations are due to a few experiments in which significant enhancement of macromolecular-labeling test activity was not observed upon fusion, but significant complementation was clearly observed in repeats of the same combinations. These results are consistent with our previous findings, which indicated that the decreased ability of GA II cells to oxidize isovaleryl-CoA involves a defective electron-transporting system rather than a defective isovaleryl-CoA dehydrogenase. IVA/IVA heterokaryons showed no complementation in any combination tested, indicating no detectable heterogeneity in isovaleric acidemia. This finding indicates that the same gene is mutated in all IVA lines. Previous results indicated that this gene codes for isovaleryl-CoA dehydrogenase. PMID:6630517

  9. Characterization of the camel skin cell line Dubca.

    Science.gov (United States)

    Klopries, M; Wernery, U; Kaaden, O R

    1995-01-01

    A skin fibroblast cell culture was established from a 2-month-old dromedary foetus. The cells were transformed by infection with SV40 and cloned in soft agar. The established cell line is now designated Dubca cells (Dubai camel) and has been in permanent culture for 95 passages. The cell culture was examined morphologically, chromosome preparations made and DNA fingerprinting performed by hybridization with the oligonucleotide probe (GTG)5. SV40 large T antigen was detected by western blotting. The viral host range was determined by infection with viruses of different families. Camelpox virus (CaPV) bovine herpesvirus-1 (BHV-1), vesicular stomatitis virus (VSV) and border disease virus (BDV) could be propagated in these cells. PMID:8556315

  10. Assessment of the potential skin irritation of lysine-derivative anionic surfactants using mouse fibroblast and human keratinocytes as an alternative to animal testing

    OpenAIRE

    Snchez Molina, Lourdes; Mitjans Arnal, Montserrat; Infante Martnez-Pardo, Ma. Rosa; Vinardell Martnez-Hidalgo, Ma. Pilar

    2004-01-01

    Purpose. The aim of this study was to identify new surfactants with low skin irritant properties for use in pharmaceutical and cosmetic formulations, employing cell culture as an alternative method to in vivo testing. In addition, we sought to establish whether potential cytotoxic properties were related to the size of the counterions bound to the surfactants. Methods. Cytotoxicity was assessed in the mouse fibroblast cell line 3T6, and the human keratinocyte cell line NCTC 2544, using the MT...

  11. Multi-layered culture of human skin fibroblasts and keratinocytes through three-dimensional freeform fabrication.

    Science.gov (United States)

    Lee, Wonhye; Debasitis, Jason Cushing; Lee, Vivian Kim; Lee, Jong-Hwan; Fischer, Krisztina; Edminster, Karl; Park, Je-Kyun; Yoo, Seung-Schik

    2009-03-01

    We present a method to create multi-layered engineered tissue composites consisting of human skin fibroblasts and keratinocytes which mimic skin layers. Three-dimensional (3D) freeform fabrication (FF) technique, based on direct cell dispensing, was implemented using a robotic platform that prints collagen hydrogel precursor, fibroblasts and keratinocytes. A printed layer of cell-containing collagen was crosslinked by coating the layer with nebulized aqueous sodium bicarbonate. The process was repeated in layer-by-layer fashion on a planar tissue culture dish, resulting in two distinct cell layers of inner fibroblasts and outer keratinocytes. In order to demonstrate the ability to print and culture multi-layered cell-hydrogel composites on a non-planar surface for potential applications including skin wound repair, the technique was tested on a poly(dimethylsiloxane) (PDMS) mold with 3D surface contours as a target substrate. Highly viable proliferation of each cell layer was observed on both planar and non-planar surfaces. Our results suggest that organotypic skin tissue culture is feasible using on-demand cell printing technique with future potential application in creating skin grafts tailored for wound shape or artificial tissue assay for disease modeling and drug testing. PMID:19108884

  12. Basal Level of Autophagy is Increased in Aging Human Skin Fibroblasts In Vitro, But Not in Old Skin

    OpenAIRE

    Demirovic, Dino; Nizard, Carine; Rattan, Suresh

    2015-01-01

    Intracellular autophagy (AP) is a stress response that is enhanced under conditions of limitation of amino acids, growth factors and other nutrients, and also when macromolecules become damaged, aggregated and fibrillated. Aging is generally accompanied by an increase in intracellular stress due to all the above factors. Therefore, we have compared the basal levels of AP in serially passaged human facial skin fibroblasts undergoing aging and replicative senescence in vitro, and ex vivo in the...

  13. Genotoxicity of alpha particles in human embryonic skin fibroblasts

    International Nuclear Information System (INIS)

    Cell inactivation and induced mutation frequencies at the hypoxanthine-guanine phosphoribosyl transferase (HGPRT) locus have been measured in cultured human fibroblasts (GM10) exposed to α particles from 238 Pu and 250 kVp X rays. The survival curves resulting from exposure to α particles are exponential. The mean lethal dose, D0, is approximately 1.3 Gy for X rays and 0.25 Gy for α particles. As a function of radiation dose, mutation induction at the HGPRT locus was linear for α particles whereas the X-ray-induced mutation data were better fitted by a quadratic function. When mutation frequencies were plotted against the log of survival, mutation frequency at a given survival level was greater in cells exposed to α particles than to X rays

  14. Differential effects of estradiol, raloxifene and tamoxifen on estrogen receptor expression in cultured human skin fibroblasts.

    Science.gov (United States)

    Haczynski, Jozef; Tarkowski, Rafal; Jarzabek, Katarzyna; Wolczynski, Slawomir; Magoffin, Denis A; Czarnocki, Krzysztof J; Ziegert, Maria; Jakowicki, Jerzy; Jakimiuk, Artur J

    2004-06-01

    The balance between ER-alpha and ER-beta in fibroblasts may be crucial in the physiological response to ligands. Up- or down-regulation of the ERs in response to different compounds could mediate the reversal of certain age-related changes in skin and connective tissue. The time-dependent effects of 17-beta estradiol, raloxifene and tamoxifen on ER-alpha and ER-beta mRNA expression in the skin fibroblast cultures were performed. Experiments were carried out in primary cultures of human skin fibroblasts obtained from postmenopausal women. The cells were cultured in medium containing: 2 micromol/l estradiol (E2), 4 micromol/l tamoxifen (Tx) or 4 micromol/l raloxifene (Rx) for 7, 24 and 32 h. ER-alpha and ER-beta mRNAs were measured by quantitative assays based on reverse transcription (RT) of the mRNA and polymerase chain reaction (PCR) amplification of the cDNA. We suggest that ER-alpha and ER-beta are co-expressed in human postmenopausal skin fibroblast and documented that the level of mRNA expression of ERs in this tissue is estradiol, raloxifene or tamoxifen regulated as a mechanism to control the action of those ligands on the cell. On the basis of ER mRNA expression levels, fibroblast response to estradiol appears to be modulated by up-regulation of ER-beta rather than ER-alpha. Two of the examined SERMs appear to have different response to modulation of ERs: response of raloxifen is modulated by up-regulation of ER-beta, and no changes in expression of ER-alpha and tamoxifen response seem to be modulated by ER down-regulation in short-term or up-regulation during longer treatment. PMID:15138633

  15. Lysosomal Storage Causes Cellular Dysfunction in Mucolipidosis II Skin Fibroblasts*

    OpenAIRE

    Otomo, Takanobu; Higaki, Katsumi; Nanba, Eiji; Ozono, Keiichi; Sakai, Norio

    2011-01-01

    Mucolipidosis II (ML-II) is a fatal inherited metabolic disease caused by deficiency of GlcNAc-phosphotransferase, which plays a role in generating the mannose 6-phosphate recognition marker on lysosomal enzymes. In ML-II, many lysosomal acid hydrolases are mistargeted out of cells, and lysosomes become filled with undigested substrates, which explains inclusion cell disease as an alternative name for this disease. In this study, we revealed various cellular phenotypes in ML-II skin fibroblas...

  16. DNA-protein crosslinking in normal human skin fibroblasts exposed to solar ultraviolet wavelengths

    International Nuclear Information System (INIS)

    Three normal human skin fibroblast cell lines were exposed to the simulated solar UV radiation produced by a fluorescent sunlamp (wavelength components shorter than either 295, 305 or 315 nm were excluded). The level of DNA-protein crosslinks (DPC) was then measured in those cells either immediately after irradiation or following a 24 h incubation. Cells were exposed to fluences that induce similar levels of DPC. For cells exposed to 10 kJ m-2 of sunlamp UV>295 nm, the level of DPC exhibited a 2-5-fold increase following incubation. In contrast, 40-100% of the DPC were removed upon incubation of cells irradiated with either 10 kJ m-2 of sunlamp UV>305 nm or 150 kJ m-2 of sunlamp UV>315 nm. A major difference between the effects induced by these wavelength regions is that, in addition to DPC, a very high level of pyrimidine dimers is also produced by sunlamp UV>295 nm, whereas much lower dimer yields result from treatment with either sunlamp UV>305 nm or sunlamp UV>315 nm. A potential role for type II DNA topoisomerase in the formation of these DPC resulting from either the change in conformational structure caused by the presence of a high level of dimers or an involvement of this enzyme in dimer excision repair is discussed. (author)

  17. Morphofunctional characteristics of fibroblasts (McCoy cell line) cultured with magnesium preparations

    OpenAIRE

    L. V. Didenko; T. G. Borovaya; E. A. Kost; E A Ulubieva; A. V. Timofeeva; A. G. Avtandilov

    2015-01-01

    Aim. To study the effect of magnesium orotate, magnesium/pyridoxine combination and magnesium sulfate on fibroblast morphofunctional characteristics in cell culture of fibroblasts (McCoy line).Material and methods. The study of fibroblasts (McCoy line) with the addition of magnesium-containing preparations (magnesium orotate, magnesium/pyridoxine combination, magnesium sulphate) to the culture medium was performed using scanning and transmission electron microscopy.Results. When adding into t...

  18. Protective effect of maghemite nanoparticles on ultraviolet-induced photo-damage in human skin fibroblasts

    International Nuclear Information System (INIS)

    This study examined the optical properties of an oxidized form of maghemite (γ-Fe2O3) nanoparticles and their protective effects against the photoaging of human skin fibroblasts irradiated with ultraviolet (UV) light. Nanoparticles with diameters ranging from 8.7 to 12 nm were prepared using a chemical co-precipitation method. The nanoparticles were coated with two surfactants to obtain a water-based product. The onset of the absorption of the γ-Fe2O3 nanoparticles in the UV-visible absorption spectra increased with increasing particle size. The γ-Fe2O3 nanoparticles significantly inhibited the production of matrix metalloproteinase-1 in human skin fibroblast HS 68 cells by 60% compared with the UV-irradiated control. These results suggest that γ-Fe2O3 nanoparticles have photoprotective properties, and have potential use as an agent against photoaging

  19. Oxidant stress leads to transcriptional activation of the human heme oxygenase gene in cultured skin fibroblasts

    International Nuclear Information System (INIS)

    Treatment of cultured human skin fibroblasts with near-UV radiation, hydrogen peroxide, and sodium arsenite induces accumulation of heme oxygenase mRNA and protein. In this study, these treatments led to a dramatic increase in the rate of RNA transcription from the heme oxygenase gene but had no effect on mRNA stability. Transcriptional activation, therefore, appears to be the major mechanism of stimulation of expression of this gene by either oxidative stress or sulfydryl reagents

  20. Protective effect of selenium and zinc on UV-A damage in human skin fibroblasts

    International Nuclear Information System (INIS)

    Ultraviolet A radiation participates in cytotoxicity and carcinogensis of the skin by a mechanism involving the generation of reactive oxygen species. Endogenous antiradical defense systems utilize metalloenzymes including Se-dependent glutathione peroxidase and Cu and Zn superoxide dismutase. The aim of the present work was to determine the protective effect of two trace elements, Se and Zn, on cultured human diploid fibroblasts exposed to UV-A radiation (broad-spectrum source with a maximum intensity at 375 nm). (Author)

  1. DNA damage in human skin fibroblasts exposed to UVA light used in clinical PUVA treatment

    International Nuclear Information System (INIS)

    Human skin fibroblasts were irradiated with a clinically used UVA light source. The doses (1.1 and 3 J/cm2) were similar to those reaching the dermis during clinical PUVA treatment of psoriasis. DNA strand breaks, as determined by alkaline elution, were formed in a dose-dependent way and disappeared within 1 hr of postincubation at 37 degrees C. These findings have clinical implications since UVA-induced DNA damage may be accompanied by mutagenic and tumor promoting effects

  2. Recovery from UV-induced potentially lethal damage in systemic lupus erythematosus skin fibroblasts

    International Nuclear Information System (INIS)

    The repair of ultraviolet light-induced potentially lethal damage was investigated in density-inhibited skin fibroblast cell strains derived from patients with systemic lupus erythematosus. The effect of exposure to polychromatic ultraviolet light composed of environmentally relevant wavelengths or to the more commonly studied, short wavelength (254 nm) ultraviolet light was studied. Systemic lupus erythematosus cells, which are hypersensitive to ultraviolet light under growth promoting conditions, were able to repair potentially lethal damage as well as normal cells. (author)

  3. L-Lactate Protects Skin Fibroblasts against Aging-Associated Mitochondrial Dysfunction via Mitohormesis

    OpenAIRE

    Jaroslav Zelenka; Aleš Dvořák; Lukáš Alán

    2015-01-01

    A moderate elevation of reactive oxygen species (ROS) production and a mild inhibition of mitochondrial respiratory chain have been associated with a health promotion and a lifespan extension in several animal models of aging. Here, we tested whether this phenomenon called mitohormesis could be mediated by L-lactate. The treatment with 5 mM L-lactate significantly increased H2O2 production and slightly inhibited the respiration in cultured skin fibroblasts and in isolated mitochondria. The L-...

  4. Sustained systemic delivery of monoclonal antibodies by genetically modified skin fibroblasts

    DEFF Research Database (Denmark)

    Nol, D; Pelegrin, M; Brockly, F; Lund, Anders Henrik; Piechaczyk, M

    2000-01-01

    In vivo production and systemic delivery of therapeutic antibodies by engineered cells might advantageously replace injection of purified antibodies for treating a variety of life-threatening diseases, including cancer, acquired immunodeficiency syndrome, and autoimmune diseases. We report here...... that skin fibroblasts retrovirally transduced to express immunoglobulin genes can be used for sustained long-term systemic delivery of cloned antibodies in immunocompetent mice. Importantly, no anti- idiotypic response against the ectopically expressed model antibody used in this study was observed...

  5. The effect of ursolic and oleanolic acids on human skin fibroblast cells

    OpenAIRE

    Helena Donica; Piotr Niedziela; Anna Matysik-Woźniak; Roman Paduch; Magdalena Wójciak-Kosior

    2012-01-01

    In this article, we look at how ursolic and oleanolic acids can be used for the purpose of quality control of natural products used in dermatocosmetology as well as of various other therapeutic preparations. Ursolic acid (UA) and oleanolic acid (OA) are pentacyclic triterpenes and they are constituents of many medicinal herbs. In this study, we analyzed the cytotoxic and anti-proliferative activity of OA and UA against normal human skin fibroblasts (HSF). Additionally, the scavenging activity...

  6. Isolation and characterization of SSEA3(+) stem cells derived from goat skin fibroblasts.

    Science.gov (United States)

    Yang, Zhongcai; Liu, Jun; Liu, Hongliang; Qiu, Mingning; Liu, Qingqing; Zheng, Liming; Pang, Meijun; Quan, Fusheng; Zhang, Yong

    2013-06-01

    Novel stem cells expressing stage-specific embryonic antigen 3 (SSEA-3) reside among human dermal fibroblasts and are known as multilineage-differentiating stress-enduring (Muse) cells. They enhance the generation efficiency of induced pluripotent stem cells. However, Muse cells have only been found in humans. We aimed to isolate SSEA3-positive cells from terminally differentiated skin fibroblasts of adult goat and determine their pluripotency. Cell clusters from SSEA3(+) populations possessed stem cell-like morphological features and normal karyotypes, were consistently positive for alkaline phosphatase, and expressed stem cell pluripotency markers. These SSEA3(+) cells remained undifferentiated over eight passages in suspension culture and were able to differentiate into cells of all three germ layers in vitro and in vivo. Our combined findings suggest that a subset of adult stem cells expressing SSEA3 also exist among adult goat skin fibroblasts. We are the first to report that multipotent adult goat cells exist among terminally differentiated goat skin in suspension culture. Our results also provide a promising platform for generation of a transgenic goat, because the undifferentiated state of stem cells was thought to be more efficient as donor cells for somatic cell nuclear transfer. PMID:23668861

  7. Radioprotective effect of c-ski on rat skin fibroblast in vitro

    International Nuclear Information System (INIS)

    Objective: To examine radioprotective effect of c-ski on rat skin fibroblast in vitro and explore its possible mechanism. Methods: The effect of soft X-ray irradiation at dose varied from 2 to 8 Gy on cell apoptosis in rat skin fibroblast were determined by flow cytometry with Annexin-V-FITC-PI labelling. The effect of c-ski gene transfection on cell apoptosis was evaluated after soft X-ray irradiation of 4 Gy. The protein expressions of Bax and Bcl-2 after c-ski gene transfection were measured with the Western blot method. Results: Soft X-ray irradiation increases cell apoptosis, and the increase is proportional to the irradiation dose. Apoptosis ratio increases with time since the irradiation, and reaches its peak at 36h after the irradiation, c-ski gene was observed to markedly decrease apoptosis index at 24 h after soft X-ray irradiation of 4 Gy compared to the control group, significant increase of the protein expression of Bcl-2 was observed. C-ski gene was found no significant effect on the protein expression of Bax. Conclusion: c-ski gene can decrease radiation sensitivity of skin fibroblast, promoting Bcl-2 protein expression is one of its possible mechanism for this radioprotective effects. (authors)

  8. Induced pluripotent stem cell lines derived from equine fibroblasts.

    Science.gov (United States)

    Nagy, Kristina; Sung, Hoon-Ki; Zhang, Puzheng; Laflamme, Simon; Vincent, Patrick; Agha-Mohammadi, Siamak; Woltjen, Knut; Monetti, Claudio; Michael, Iacovos Prodromos; Smith, Lawrence Charles; Nagy, Andras

    2011-09-01

    The domesticated horse represents substantial value for the related sports and recreational fields, and holds enormous potential as a model for a range of medical conditions commonly found in humans. Most notable of these are injuries to muscles, tendons, ligaments and joints. Induced pluripotent stem (iPS) cells have sparked tremendous hopes for future regenerative therapies of conditions that today are not possible to cure. Equine iPS (EiPS) cells, in addition to bringing promises to the veterinary field, open up the opportunity to utilize horses for the validation of stem cell based therapies before moving into the human clinical setting. In this study, we report the generation of iPS cells from equine fibroblasts using a piggyBac (PB) transposon-based method to deliver transgenes containing the reprogramming factors Oct4, Sox2, Klf4 and c-Myc, expressed in a temporally regulated fashion. The established iPS cell lines express hallmark pluripotency markers, display a stable karyotype even during long-term culture, and readily form complex teratomas containing all three embryonic germ layer derived tissues upon in vivo grafting into immunocompromised mice. Our EiPS cell lines hold the promise to enable the development of a whole new range of stem cell-based regenerative therapies in veterinary medicine, as well as aid the development of preclinical models for human applications. EiPS cell could also potentially be used to revive recently extinct or currently threatened equine species. PMID:21347602

  9. Heterogeneous response to X-ray and ultraviolet light irradiations of cultured skin fibroblasts in two families with Gardner's Syndrome

    International Nuclear Information System (INIS)

    A heterogeneous response to X-ray and far UV (254 nm) light irradiations was found in cultured skin fibroblast lines from 2 separate families with Gardner's syndrome. When compared to 2 normal control cultures and cultures from 2 patients with nonfamilial colon cancer, cultures from 4 clinically affected members of family 1 showed increased sensitivity to the lethal effects of both X-ray and UV light irradiations. These cells also showed a delayed pattern of X-ray potentially lethal damage repair (PLDR) and absent UV PLDR. In contrast, cultures from 3 members of family 2 (2 of whom were clinically affected) showed a normal response of survival and PLDR to both X-ray and UV light irradiations. Thus increased sensitivity of cultured skin fibroblasts to X-ray and UV light irradiations was not a consistent in vitro finding in patients with Gardner's syndrome. However, in families with Gardner's syndrome who demonstrate in vitro radiosensitivity, additional studies are needed to assess the usefulness of these techniques in detecting affected individuals prior to the development of colon carcinoma and other manifestations

  10. Radiosensitivity of fibroblasts obtained from a caf-au-lait spot and normal-appearing skin of a patient with neurofibromatosis (NF-6).

    Science.gov (United States)

    Hannan, M A; Smith, B P; Sigut, D; Sackey, K

    1990-07-15

    Fibroblast cells derived from a caf-au-lait spot and normal-appearing skin of a neurofibromatosis (NF-6) patient were studied for radiosensitivity in comparison with two normal cell lines used as "controls." No difference in radiosensitivity was observed between the patient's cell lines and the controls using acute gamma-irradiation. However, a markedly increased radiosensitivity of the fibroblasts obtained from the patient's skin of normal appearance was demonstrated after chronic gamma-irradiation. The cells from the caf-au-lait spot showed intermediate sensitivity to chronic irradiation as compared with the control cell lines and the fibroblasts derived from the normal skin of the patient. These results showed the usefulness of chronic irradiation in detecting increased cellular radiosensitivity which may result from a unique DNA repair defect in an NF patient. We suggest that enhanced genetic changes in radiosensitive NF patients may lead to formation of caf-au-lait lesions and certain tumors. Such a transformation may be associated with production of radiotolerant cells. PMID:2113426

  11. Radiosensitivity of fibroblasts obtained from a cafe-au-lait spot and normal-appearing skin of a patient with neurofibromatosis (NF-6)

    International Nuclear Information System (INIS)

    Fibroblast cells derived from a cafe-au-lait spot and normal-appearing skin of a neurofibromatosis (NF-6) patient were studied for radiosensitivity in comparison with two normal cell lines used as controls. No difference in radiosensitivity was observed between the patient's cell lines and the controls using acute gamma-irradiation. However, a markedly increased radiosensitivity of the fibroblasts obtained from the patient's skin of normal appearance was demonstrated after chronic gamma-irradiation. The cells from the cafe-au-lait spot showed intermediate sensitivity to chronic irradiation as compared with the control cell lines and the fibroblasts derived from the normal skin of the patient. These results showed the usefulness of chronic irradiation in detecting increased cellular radiosensitivity which may result from a unique DNA repair defect in an NF patient. We suggest that enhanced genetic changes in radiosensitive NF patients may lead to formation of cafe-au-lait lesions and certain tumors. Such a transformation may be associated with production of radiotolerant cells

  12. Radiosensitivity of fibroblasts obtained from a cafe-au-lait spot and normal-appearing skin of a patient with neurofibromatosis (NF-6)

    Energy Technology Data Exchange (ETDEWEB)

    Hannan, M.A.; Smith, B.P.; Sigut, D.; Sackey, K. (King Faisal Specialist Hospital and Research Centre, Riyadh (Saudi Arabia))

    1990-07-15

    Fibroblast cells derived from a cafe-au-lait spot and normal-appearing skin of a neurofibromatosis (NF-6) patient were studied for radiosensitivity in comparison with two normal cell lines used as controls. No difference in radiosensitivity was observed between the patient's cell lines and the controls using acute gamma-irradiation. However, a markedly increased radiosensitivity of the fibroblasts obtained from the patient's skin of normal appearance was demonstrated after chronic gamma-irradiation. The cells from the cafe-au-lait spot showed intermediate sensitivity to chronic irradiation as compared with the control cell lines and the fibroblasts derived from the normal skin of the patient. These results showed the usefulness of chronic irradiation in detecting increased cellular radiosensitivity which may result from a unique DNA repair defect in an NF patient. We suggest that enhanced genetic changes in radiosensitive NF patients may lead to formation of cafe-au-lait lesions and certain tumors. Such a transformation may be associated with production of radiotolerant cells.

  13. Loss of a putative tumor suppressor locus after gamma-ray-induced neoplastic transformation of HeLa x Skin fibroblast human cell hybrids

    International Nuclear Information System (INIS)

    The nontumorigenic HeLa x skin fibroblast hybrid cell line, CGL1, can be induced to re-express HeLa tumor-associated cell surface antigen, p75-IAP (intestinal alkaline phosphatase), with resulting neoplastic transformation, by exposure to γ radiation. This has allowed the human hybrid system to be developed into a quantitative in vitro model for radiation-induced neoplastic transformation of human cells. Recently, several γ-ray-induced IAP-expression mutants (GIMs) of the nontumorigenic HeLa x skin fibroblast hybrid CGL1 were isolated and all were tumorigenic when injected subcutaneously into nude mice. Control cell lines which were negative for p75-IAP (CONs) were also isolated from irradiated populations, and none were found to be tumorigenic. We have now begun to investigate the molecular basis of radiation-induced neoplastic transformation in this system by studying the potential genetic linkage between p75/IAP expression, tumorigenicity and damage to a putative tumor suppressor locus on fibroblast chromosome 11. Previous analysis of rare spontaneous segregants has indicated that this locus is involved in the regulation of tumorigenicity and in the expression of the HeLa tumor-associated cell surface marker intestinal alkaline phosphatase (p75-IAP) in this system. Therefore, analysis by restriction fragment length polymorphism and chromosome painting have been performed for chromosome 11, and for chromosome 13 as a control, for the p75/IAP-positive GIM and p75/IAP-negative CON cell lines. We report that in five of eight of the GIMs large-scale damage to the fibroblast chromosome 11's is evident (four GIMs have lost one complete copy of a fibroblast chromosome 11 heavily damaged). None of the CONs, however (0/5), have lost a complete copy of either fibroblast chromosome 11. No large-scale damage to the control chromosome 13's was detected in the GIMs or CONs. 49 refs., 3 figs., 2 tabs

  14. Resistance to 1,25-dihydroxyvitamin D. Association with heterogeneous defects in cultured skin fibroblasts

    International Nuclear Information System (INIS)

    We evaluated the interaction of [3H]1,25(OH)2D3 with skin fibroblasts cultured from normal subjects or from affected members of six kindreds with rickets and resistance to 1-alpha, 25(OH)2D [1,25(OH)2D]. We analyzed two aspects of the radioligand interaction; nuclear uptake with dispersed, intact cells at 37 degrees C and binding at 0 degrees C with soluble extract (cytosol) prepared from cells disrupted in buffer containing 300 mM KCl and 10 mM sodium molybdate. With normal fibroblasts the affinity and capacity of nuclear uptake of [3H]1,25(OH)2D3 were 0.5 nM and 10,300 sites per cell, respectively; for binding with cytosol these were 0.13 nM and 8,900 sites per cell, respectively. In all cases where the radioligand bound with high affinity in nucleus or cytosol, the nucleus- or cytosol-associated radioligand exhibited normal sedimentation velocity on sucrose density gradients. When two kindreds exhibited similar patterns (i.e. pattern a or c) with the analyses of cultured fibroblasts, clinical features in affected members suggested that the underlying genetic defects were not identical. In conclusion: (a) Fibroblasts cultured from human skin manifest nuclear uptake and cytosol binding of [3H]1,25(OH)2D3 that is an expression of the genes determining these processes in target tissues. (b) Based upon data from clinical evaluations and from analyses of cultured fibroblasts, severe resistance to 1,25(OH)2D resulted from five or six distinct genetic mutations in six kindreds

  15. Resistance to 1,25-dihydroxyvitamin D. Association with heterogeneous defects in cultured skin fibroblasts

    International Nuclear Information System (INIS)

    The authors evaluated the interaction of [3H]1,25(OH)2D3 with skin fibroblasts cultured from normal subjects or from affected members of six kindreds with rickets and resistance to 1-alpha, 25(OH)2D [1,25(OH)2D]. They analyzed two aspects of the radioligand interaction; nuclear uptake with dispersed, intact cells at 37 degrees C and binding at 0 degrees C with soluble extract (cytosol) prepared from cells disrupted in buffer. With normal fibroblasts the affinity and capacity of nuclear uptake of [3H]1,25(OH)2D3 were 0.5 nM and 10,300 sites per cell, respectively; for binding with cytosol these were 0.13 nM and 8,900 sites per cell, respectively. The following four patterns of interaction with [3H]1,25(OH)2D3 were observed with cells cultured from affected patients. In all cases where the radioligand bound with high affinity in nucleus or cytosol, the nucleus- or cytosol-associated radioligand exhibited normal sedimentation velocity on sucrose density gradients. When two kindreds exhibited similar patterns (i.e. pattern a or c) with the analyses of cultured fibroblasts, clinical features in affected members suggested that the underlying genetic defects were not identical. In conclusion: (a) Fibroblasts cultured from human skin manifest nuclear uptake and cytosol binding of [3H]1,25(OH)2D3 that is an expression of the genes determining these processes in target tissues. (b) Based upon data from clinical evaluations and from analyses of cultured fibroblasts, severe resistance to 1,25(OH)2D resulted from five or six distinct genetic mutations in six kindreds

  16. Abnormal phenotype of cultured fibroblasts in human skin with chronic radiotherapy damage

    International Nuclear Information System (INIS)

    Purpose: The pathophysiological aspects of radiation-induced fibrosis (RIF) have not been well characterized. We therefore cultured human fibroblasts from samples of skin with RIF to investigate the long-term effects of therapeutic irradiation. Materials and methods: Biopsies of normal and RIF skin were obtained from patients previously irradiated for cancer, without recurrence. Cells were extracted from dermis samples by the outgrowth technique, seeded as monolayers and cultured at confluence. Enzyme activities and proteins were assayed, RNA was isolated and Northern blot analysis was performed on surviving cells between passages 2 and 5. Results: RIF cell cultures displayed heterogeneous fibroblasts populations. The initial outgrowth consisted of one-third small cells that floated rapidly, one-third spindle-shaped cells migrating far from the explant to form islets and one-third large pleiomorphic cells. In subsequent subcultures, surviving cells exhibited either myofibroblastic characteristics with a normal proliferative capacity or senescent morphology with a reduced proliferative capacity. These RIF cells had a brief finite lifespan, with dramatically reduced growth rate during their initial outgrowth and the following passages. Study of the antioxidant metabolism showed that Mn superoxide dismutase and catalase activities were significantly weaker in surviving RIF cells than healthy fibroblasts. These exhausted RIF cells exhibited no overexpression of transforming growth factor β or tissue inhibitor of metalloproteinase. Conclusion: Irradiation may lead to apparently contradictory effects such as fibrosis and necrosis in clinical practice. In cell culture, we observed two main cellular phenotypes which may be related to both processes, i.e. myofibroblast-like cells and fibrocyte-like cells. These two phenotypes may represent two steps in the differentiation induced as a long-term effect of therapeutic irradiation of the skin. Cell culture probably accelerates the induction of the terminal differentiation in RIF fibroblasts. (Copyright (c) 1998 Elsevier Science B.V., Amsterdam. All rights reserved.)

  17. Preventive effects of tamarind seed coat extract on UVA-induced alterations in human skin fibroblasts.

    Science.gov (United States)

    Phetdee, Khemjira; Rakchai, Racharat; Rattanamanee, Kwanchai; Teaktong, Thanasak; Viyoch, Jarupa

    2014-01-01

    One of the most damaging actions on skin is from solar radiation, particularly from its ultraviolet (UV) component, through the formation of oxidative species. Thus, an antioxidant strategy that prevents the formation of these oxidants could form the basis of an efficacious cutaneous protectant. Many herbal materials contain antioxidant polyphenols, and this study assessed the possibility that tamarind seed coat extract could fulfill this role. An alcoholic extract of the tamarind (Tamarindus indica L.) seed coat showed stronger antioxidant activity (2,2-diphenyl-1-picrylhydrazyl inhibition, EC(50) = 12.9 μg/ml) than L-ascorbic acid (EC(50) = 22.9 μg/ml) and α-tocopherol (EC(50) = 29.3 μg/ml). In cultured fibroblasts taken from human skin, hydrogen peroxide (100-1000 μM) damaged 62-92% of the cells compared to only 35-47% when the cells were preincubated in extract (200 μg/ml) for 24 h. UVA (40 J/cm2) irradiation of human fibroblasts damaged 25% of the cells but the death rate was reduced to 10% with extract. UV irradiation increased the proportion of cells arrest in G(0)/G(1) phase (from 59% to 78%) but this was largely prevented by the extract (64%), according to flow cytometry. Intracellular total glutathione of UVA-irradiated cells pretreated with the extract increased to 10-25% compared to the non-pretreated group at 24-72 h after irradiation. Fibroblasts typically increased matrix metalloproteinase-1 secretion after photodamage, and this is prevented by the extract. This is the first report showing that tamarind seed coat extract is an antioxidant and can protect human skin fibroblasts from cellular damage produced by UVA and thus may form the foundation for an antiaging cosmetic. PMID:24602819

  18. In vitro study of RRS HA injectable mesotherapy/biorevitalization product on human skin fibroblasts and its clinical utilization.

    Science.gov (United States)

    Deglesne, Pierre-Antoine; Arroyo, Rodrigo; Ranneva, Evgeniya; Deprez, Philippe

    2016-01-01

    Mesotherapy/biorevitalization with hyaluronic acid (HA) is a treatment approach currently used for skin rejuvenation. Various products with a wide range of polycomponent formulations are available on the market. Most of these formulations contain noncross-linked HA in combination with a biorevitalization cocktail, formed by various amounts of vitamins, minerals, amino acids, nucleotides, coenzymes, and antioxidants. Although ingredients are very similar among the different products, in vitro and clinical effects may vary substantially. There is a real need for better characterization of these products in terms of their action on human skin or in vitro skin models. In this study, we analyzed the effect of the RRS(®) (Repairs, Refills, Stimulates) HA injectable medical device on human skin fibroblasts in vitro. Skin fibroblast viability and its capacity to induce the production of key extracellular matrix were evaluated in the presence of different concentrations of RRS HA injectable. Viability was evaluated through colorimetric MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay, and key extracellular matrix genes, type I collagen and elastin, were quantified by quantitative polymerase chain reaction. Results demonstrated that RRS HA injectable could promote human skin fibroblast viability (+15%) and increase fibroblast gene expression of type I collagen and elastin by 9.7-fold and 14-fold in vitro, respectively. These results demonstrate that mesotherapy/biorevitalization products can, at least in vitro, effectively modulate human skin fibroblasts. PMID:26966384

  19. In vitro study of RRS HA injectable mesotherapy/biorevitalization product on human skin fibroblasts and its clinical utilization

    Science.gov (United States)

    Deglesne, Pierre-Antoine; Arroyo, Rodrigo; Ranneva, Evgeniya; Deprez, Philippe

    2016-01-01

    Mesotherapy/biorevitalization with hyaluronic acid (HA) is a treatment approach currently used for skin rejuvenation. Various products with a wide range of polycomponent formulations are available on the market. Most of these formulations contain noncross-linked HA in combination with a biorevitalization cocktail, formed by various amounts of vitamins, minerals, amino acids, nucleotides, coenzymes, and antioxidants. Although ingredients are very similar among the different products, in vitro and clinical effects may vary substantially. There is a real need for better characterization of these products in terms of their action on human skin or in vitro skin models. In this study, we analyzed the effect of the RRS (Repairs, Refills, Stimulates) HA injectable medical device on human skin fibroblasts in vitro. Skin fibroblast viability and its capacity to induce the production of key extracellular matrix were evaluated in the presence of different concentrations of RRS HA injectable. Viability was evaluated through colorimetric MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay, and key extracellular matrix genes, type I collagen and elastin, were quantified by quantitative polymerase chain reaction. Results demonstrated that RRS HA injectable could promote human skin fibroblast viability (+15%) and increase fibroblast gene expression of type I collagen and elastin by 9.7-fold and 14-fold in vitro, respectively. These results demonstrate that mesotherapy/biorevitalization products can, at least in vitro, effectively modulate human skin fibroblasts.

  20. Redox Imbalance and Morphological Changes in Skin Fibroblasts in Typical Rett Syndrome

    Science.gov (United States)

    Amabile, Sonia; Belmonte, Giuseppe; Valacchi, Giuseppe; Galano, Jean-Marie; Ciccoli, Lucia; Renieri, Alessandra; Hayek, Joussef

    2014-01-01

    Evidence of oxidative stress has been reported in the blood of patients with Rett syndrome (RTT), a neurodevelopmental disorder mainly caused by mutations in the gene encoding the Methyl-CpG-binding protein 2. Little is known regarding the redox status in RTT cellular systems and its relationship with the morphological phenotype. In RTT patients (n = 16) we investigated four different oxidative stress markers, F2-Isoprostanes (F2-IsoPs), F4-Neuroprostanes (F4-NeuroPs), nonprotein bound iron (NPBI), and (4-HNE PAs), and glutathione in one of the most accessible cells, that is, skin fibroblasts, and searched for possible changes in cellular/intracellular structure and qualitative modifications of synthesized collagen. Significantly increased F4-NeuroPs (12-folds), F2-IsoPs (7.5-folds) NPBI (2.3-folds), 4-HNE PAs (1.48-folds), and GSSG (1.44-folds) were detected, with significantly decreased GSH (−43.6%) and GSH/GSSG ratio (−3.05 folds). A marked dilation of the rough endoplasmic reticulum cisternae, associated with several cytoplasmic multilamellar bodies, was detectable in RTT fibroblasts. Colocalization of collagen I and collagen III, as well as the percentage of type I collagen as derived by semiquantitative immunofluorescence staining analyses, appears to be significantly reduced in RTT cells. Our findings indicate the presence of a redox imbalance and previously unrecognized morphological skin fibroblast abnormalities in RTT patients. PMID:24987493

  1. Skin fibroblasts from a D-deletion type retinoblastoma patient are abnormally X-ray sensitive

    International Nuclear Information System (INIS)

    Retinoblastoma is a rare malignant eye tumour that appears either spontaneously or in genetically predisposed persons. The latter group is composed of persons who inherit the tumour with a dominant mode of transmission (the familial type) and those who have a deletion in the long arm of chromosome 13 referred to as the D-deletion type. When this deletion is present it is observed in many somatic cells and is often associated with structural defects. Survivors of the genetic forms of retinoblastoma have an increased risk of the development of cancers at other sites. A single genetic locus is unlikely to predispose many somatic cells to tumour formation unless a fundamental molecular defect, possibly related to DNA repair, is present. In order to investigate this hypothesis a study was made of the in vitro X-ray sensitivity of skin fibroblasts derived from three retinoblastoma patients, comprising a pair of twins with the familial type accompanied by no gross chromosome abnormalities, and a patient with the D-deletion type. It was found that fibroblasts derived from the D-deletion patient were significantly more radiosensitive than those from the other two patients. X-ray survival curves are shown. It is concluded that skin fibroblasts derived from a patient with the D-deletion variant of retinoblastoma are abnormally radiosensitive. Future investigations may indicate a specific defect in molecular repair of DNA that will explain the predisposition of these patients to the development of other tumours. (U.K.)

  2. Mitochondrial ribosomal protein S18-2 evokes chromosomal instability and transforms primary rat skin fibroblasts

    KAUST Repository

    Kashuba, Elena

    2015-05-12

    We have shown earlier that overexpression of the human mitochondrial ribosomal protein MRPS18-2 (S18-2) led to immortalization of primary rat embryonic fibroblasts. The derived cells expressed the embryonic stem cell markers, and cellular pathways that control cell proliferation, oxidative phosphorylation, cellular respiration, and other redox reactions were activated in the immortalized cells. Here we report that, upon overexpression of S18-2 protein, primary rat skin fibroblasts underwent cell transformation. Cells passed more than 300 population doublings, and two out of three tested clones gave rise to tumors in experimental animals. Transformed cells showed anchorage-independent growth and loss of contact inhibition; they expressed epithelial markers, such as E-cadherin and β-catenin. Transformed cells showed increased telomerase activity, disturbance of the cell cycle, and chromosomal instability. Taken together, our data suggest that S18-2 is a newly identified oncoprotein that may be involved in cancerogenesis.

  3. Differentiation state of skin fibroblast cultures versus risk of subcutaneous fibrosis after radiotherapy

    International Nuclear Information System (INIS)

    Background and purpose: There is increasing evidence for patient-to-patient variation in the response of normal tissue to radiotherapy. Recently, it has been suggested that accumulation of functional fibrocytes may be a key step in the development of radiation-induced fibrosis. Therefore, we have examined a possible relationship between the differentiation state of untreated fibroblasts and the risk of radiation-induced subcutaneous fibrosis in individual patients. Materials and methods: We used skin fibroblast cultures isolated from eight postmastectomy radiotherapy patients whose individual clinical radiosensitivity was assessed by the mean excess risk of fibrosis. Different types of potentially mitotic progenitor fibroblasts (MF) and postmitotic functional fibrocytes (PMF) in the terminal differentiation lineage (MFI approaches MFII approaches MFIII approaches PMF) were scored morphologically in clonal culture. Progression of differentiation was quantified by the ratio L/E of colony-forming late (MFIII and late MFII) and early (MFI and early MFII) progenitors. Results: We observed a correlation between the ratio L/E and the mean risk of fibrosis (rS=0.743, P=0.03), indicating an approximately 10-fold increase in L/E with an increasing risk of fibrosis. This was paralleled by a decreasing trend in the absolute numbers of early progenitor types. By contrast, there was no significant correlation between the plating efficiency and the risk of fibrosis. Conclusions: The data suggest that the risk of fibrosis increases with the progression of the differentiation of untreated progenitor fibroblasts, indicating that the progression of fibroblast differentiation may be a co-factor in the development of radiation-induced fibrosis. If this hypothesis is validated, it provides a rationale for a novel predictive test to identify patients with an increased risk of subcutaneous fibrosis. (Copyright (c) 1998 Elsevier Science B.V., Amsterdam. All rights reserved.)

  4. Characterisation of the kynurenine pathway in skin-derived fibroblasts and keratinocytes.

    Science.gov (United States)

    Sheipouri, Diba; Grant, Ross; Bustamante, Sonia; Lovejoy, David; Guillemin, Gilles J; Braidy, Nady

    2015-06-01

    Acute UVB exposure triggers inflammation leading to the induction of indoleamine 2,3 dioxygenase (IDO1), one of the first enzymes in the kynurenine pathway (KP) for tryptophan degradation. However, limited studies have been undertaken to determine the catabolism of tryptophan within the skin. The aim of this study was two fold: (1) to establish if the administration of the proinflammatory cytokine interferon-gamma (IFN-γ) and/or UVB radiation elicits differential KP expression patterns in human fibroblast and keratinocytes; and (2) to evaluate the effect of KP metabolites on intracellular nicotinamide adenine dinucleotide (NAD(+) ) levels, and cell viability. Primary cultures of human fibroblasts and keratinocytes were used to examine expression of the KP at the mRNA level using qPCR, and at the protein level using immunocytochemistry. Cellular responses to KP metabolites were assessed by examining extracellular lactate dehydrogenase (LDH) activity and intracellular NAD(+) levels. Major downstream KP metabolites were analyzed using GC/MS and HPLC. Our data shows that the KP is fully expressed both in human fibroblasts and keratinocytes. Exposure to UVB radiation and/or IFN-γ causes significant changes in the expression pattern of downstream KP metabolites and enzymes. Exposure to various concentrations of KP metabolites showed marked differences in cell viability and intracellular NAD(+) production, providing support for involvement of the KP in the de novo synthesis of NAD(+) in the skin. This new information will have a significant impact on our understanding of the pathogenesis of UV related skin damage and the diagnosis of KP related disease states. PMID:25639585

  5. Aloin Protects Skin Fibroblasts from Heat Stress-Induced Oxidative Stress Damage by Regulating the Oxidative Defense System

    OpenAIRE

    Liu, Fu-Wei; Liu, Fu-Chao; Wang, Yu-Ren; Tsai, Hsin-I; Yu, Huang-Ping

    2015-01-01

    Oxidative stress is commonly involved in the pathogenesis of skin damage induced by environmental factors, such as heat stress. Skin fibroblasts are responsible for the connective tissue regeneration and the skin recovery from injury. Aloin, a bioactive compound in Aloe vera, has been reported to have various pharmacological activities, such as anti-inflammatory effects. The aim of this study was to investigate the protective effect of aloin against heat stress-mediated oxidative stress in hu...

  6. Membrane damage induced in cultured human skin fibroblasts by UVA irradiation

    International Nuclear Information System (INIS)

    Irradiation of cultured human skin fibroblasts with ultraviolet light from 320 to 400 nm (UVA) leads to a decrease in the membrane fluidity exemplified by an enhanced fluorescence anisotropy of the lipophilic fluorescent probe 1-[4-trimethylamino)-phenyl]-6-phenylhexa-1,3,5-triene. This UVA-induced decrease in fluidity is associated with lactate dehydrogenase leakage in the supernatant. Vitamin E, an inhibitor of lipid peroxidation, exerts a protective effect on both phenomena. Therefore, this UVA-induced damage in membrane properties may be related to lipid peroxidation processes. Moreover, exponentially growing cells are more sensitive to these UVA-induced alterations than confluent cells. (Author)

  7. Fibroblast radiosensitivity versus acute and late normal skin responses in patients treated for breast cancer

    International Nuclear Information System (INIS)

    Purpose/Objective: To determine if the radiosensitivity of normal human skin fibroblasts, measured in early passage cultures, is significantly correlated with the degree of acute or late normal skin damage in patients treated for breast cancer with radiotherapy. Methods and Materials: In the 1970s, a series of breast cancer patients was treated at the Department of Oncology in Gothenburg, Sweden with postoperative irradiation to the parasternal region. Patients were treated bilaterally using different fractionation schedules and doses to the right and left fields. Peak acute reactions were scored on a six-point scale, and skin erythema was measured by reflectance spectrophotometry. Telangiectasia was graded over time on a six-point scale. In April 1992, two small skin biopsies were obtained from 22 patients in two treatment groups (i.e., four dose-fractionation schedules) and, using either delayed or immediate plating, fibroblast radiosensitivity was measured in early passage cultures by clonogenic survival, after high and low dose-rate irradiations. Survival at 2.0 Gy (SF2) was calculated from complete survival curves. Results: To test assay reproducibility, SF2 values derived from paired biopsies of the same patient (12 cases) were compared. A reasonably good correlation (p = 0.075) was obtained for SF2s determined by high dose-rate irradiations with immediate plating, but not for delayed plating or low dose-rate treatments. The median coefficient of variation in the replicate SF2s after high dose-rate treatment and immediate plating was 13%, suggesting that the poor correlation in paired SF2 values is due to the magnitude of the uncertainty in SF2 relative to the overall spread in SF2 values between patients (CV = 28%). Individual SF2 values and averaged values from patients with data from two biopsies were compared with the acute and late clinical reactions. A significant negative correlation was found between SF2 and relative clinical response, but only when averaged high dose-rate SF2 values and telangiectasia scores were compared. There was no significant correlation between average SF2 values and acute responses or between individual SF2 measurements and either the acute or late clinical response. Conclusion: The results of this study suggest that the degree of late telangiectasia is at least partially dependent upon the intrinsic cellular radiosensitivity of normal fibroblasts, but the relationship is not clear cut. Multiple replicate assays are necessary to obtain reliable estimates of fibroblast SF2 values using current techniques

  8. Near-ultraviolet radiation-induced lipid peroxidation and membrane effects in Escherichia coli and human skin fibroblasts

    International Nuclear Information System (INIS)

    The first part of this thesis examines the response of an unsaturated fatty acid auxotroph, Escherichia coli K1060 to broad-band near-UV radiation. Sensitivity, lipid peroxidation and leakage of rubidium from irradiated cells were found to increase with increasing unsaturation of membrane fatty acids. The involvement of singlet oxygen was implicated by an increase in sensitivity, lipid peroxidation and leakage of rubidium following irradiation in deuterium oxide. Some factors influencing survival following irradiation were investigated, where lower growth rates were shown to enhance survival. In the second part, the study was extended to human fibroblasts where a normal human skin fibroblast strain, GM730 and a strain derived from an actinic reticuloid patient, AR6LO, are compared. Lipid peroxidation was measured in both cell lines following broad-band near-UV irradiation. Membrane activity, as assessed by the pinocytic uptake of 14C-sucrose and its subsequent release from the cell, was measured. Near-UV irradiation was found to increase such activity in both strains. Vitamin E and Trolox-C were found to decrease this response in AR6LO but not GM730 cells. The final part consists of preliminary investigations into the near-UV induced peroxidation of fatty acids and liposomes, and the subsequent increase in the level of hydroperoxides in the hours following irradiation. (author)

  9. Basic fibroblast growth factor is beneficial for postoperative color uniformity in split-thickness skin grafting.

    Science.gov (United States)

    Akita, Sadanori; Akino, Kozo; Yakabe, Aya; Tanaka, Katsumi; Anraku, Kuniaki; Yano, Hiroki; Hirano, Akiyoshi

    2010-01-01

    Color changes of visible and exposed body surfaces, such as the face and extremities, after burn injury or surgery, such as skin grafting, flap, or sclerotherapy for vascular malformations, are sometimes a concern. The consequences reduce the satisfaction of both patients and physicians. An easy and reproducible method has not yet been established for an objective analysis of color changes; therefore, we tested a hand-held color analyzer (NF-333; Nippon Denshoku Co. Ltd) with data transport to a computer database and analysis software for posttreatment skin color change. The parameters included L, a, and b, which measure clarity, red, and yellow, respectively. Two groups were prospectively divided with 20 (11 females and nine males) patients per group. One group received skin grafting plus basic fibroblast growth factor (bFGF) spray daily and the other group received only skin grafting. The patients were randomized by the date of their first visit to our hospital. Patients were treated with bFGF on odd days, while patients who came on even days were included in the non-bFGF-treated group. The donor site for skin grafting was the lateral thighs and the thickness was similar in both groups. The results were compared at 1-year posttreatment follow-up. Clinical and objective assessments of the scars were performed 1 to years after complete healing. Color change differentials in comparison with the surrounding skin were lower with bFGF treatment in all parameters (p<0.01), along with clinical assessment with the Vancouver Scar Scale; therefore, the treatment contribute to a better color match with skin grafting postoperatively. PMID:20868384

  10. Human skin fibroblast stromelysin: structure, glycosylation, substrate specificity, and differential expression in normal and tumorigenic cells

    International Nuclear Information System (INIS)

    The authors have purified and determined the complete primary structure of human stromelysin, a secreted metalloprotease with a wide range of substrate specificities. Human stromelysin is synthesized in a preproenzyme form with a calculated size of 53,977 Da and a 17-amino acid long signal peptide. Prostromelysin is secreted in two forms, with apparent molecular masses on NaDodSO4/PAGE of 60 and 57 kDa. Human stromelysin is capable of degrading proteoglycan, fibronectin, laminin, and type IV collagen but not interstitial type I collagen. The enzyme is not capable of activating purified human fibroblast procollagenase. Analysis of its primary structure shows that stromelysin is in all likelihood the human analog of rat transin, which is an oncogene transformation-induced protease. The pattern of enzyme expression in normal and tumorigenic cells revealed that human skin fibroblasts in vitro secrete stromelysin constitutively. Human fetal lung fibroblasts transformed with simian virus 40, human bronchial epithelial cells transformed with the ras oncogene, fibrosarcoma cells (HT-1080), and a melanoma cell strain (A 2058), do not express this protease nor can the enzyme be induced in these cells by treatment with phorbol 12-myristate 13-acetate. The data indicate that the expression and the possible involvement of secreted metalloproteases in tumorigenesis result from a specific interaction between the transforming factor and the target cell, which may vary in different species

  11. In vitro study of RRS HA injectable mesotherapy/biorevitalization product on human skin fibroblasts and its clinical utilization

    Directory of Open Access Journals (Sweden)

    Deglesne PA

    2016-02-01

    Full Text Available Pierre-Antoine Deglesne,* Rodrigo Arroyo,* Evgeniya Ranneva, Philippe Deprez Research and Development, SKIN TECH PHARMA GROUP, Castelló d'Empúries, Spain  *These authors contributed equally to this work Abstract: Mesotherapy/biorevitalization with hyaluronic acid (HA is a treatment approach currently used for skin rejuvenation. Various products with a wide range of polycomponent formulations are available on the market. Most of these formulations contain noncross-linked HA in combination with a biorevitalization cocktail, formed by various amounts of vitamins, minerals, amino acids, nucleotides, coenzymes, and antioxidants. Although ingredients are very similar among the different products, in vitro and clinical effects may vary substantially. There is a real need for better characterization of these products in terms of their action on human skin or in vitro skin models. In this study, we analyzed the effect of the RRS® (Repairs, Refills, Stimulates HA injectable medical device on human skin fibroblasts in vitro. Skin fibroblast viability and its capacity to induce the production of key extracellular matrix were evaluated in the presence of different concentrations of RRS HA injectable. Viability was evaluated through colorimetric MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay, and key extracellular matrix genes, type I collagen and elastin, were quantified by quantitative polymerase chain reaction. Results demonstrated that RRS HA injectable could promote human skin fibroblast viability (+15% and increase fibroblast gene expression of type I collagen and elastin by 9.7-fold and 14-fold in vitro, respectively. These results demonstrate that mesotherapy/biorevitalization products can, at least in vitro, effectively modulate human skin fibroblasts.Keywords: mesotherapy, medical device, RRS, collagen, elastin, extracellular matrix

  12. Oxidative stress in skin fibroblasts cultures from patients with Parkinson's disease

    Directory of Open Access Journals (Sweden)

    Arenas Joaquín

    2010-10-01

    Full Text Available Abstract Background In the substantia nigra of Parkinson's disease (PD patients, increased lipid peroxidation, decreased activities of the mitochondrial complex I of the respiratory chain, catalase and glutathione-peroxidase, and decreased levels of reduced glutathione have been reported. These observations suggest that oxidative stress and mitochondrial dysfunction play a role in the neurodegeneration in PD. We assessed enzymatic activities of respiratory chain and other enzymes involved in oxidative processes in skin fibroblasts cultures of patients with PD. Methods We studied respiratory chain enzyme activities, activities of total, Cu/Zn- and Mn-superoxide-dismutase, gluthatione-peroxidase and catalase, and coenzyme Q10 levels in skin fibroblasts cultures from 20 Parkinson's disease (PD patients and 19 age- and sex- matched healthy controls. Results When compared with controls, PD patients showed significantly lower specific activities for complex V (both corrected by citrate synthase activity and protein concentrations. Oxidized, reduced and total coenzyme Q10 levels (both corrected by citrate synthase and protein concentrations, and activities of total, Cu/Zn- and Mn-superoxide-dismutase, gluthatione-peroxidase and catalase, did not differ significantly between PD-patients and control groups. Values for enzyme activities in the PD group did not correlate with age at onset, duration, scores of the Unified Parkinson's Disease Rating scales and Hoehn-Yahr staging. Conclusions The main result of this study was the decreased activity of complex V in PD patients. This complex synthesizes ATP from ADP using an electrochemical gradient generated by complexes I-IV. These results suggest decreased energetic metabolism in fibroblasts of patients with PD.

  13. Sensitivity of cultured skin fibroblasts from patients with neurofibromatosis to DNA-damaging agents

    International Nuclear Information System (INIS)

    Neurofibromatosis (NF) is an autosomal dominant disorder associated with various constitutional abnormalities as well as a striking predisposition for malignant and nonmalignant neoplasms, both in cells originating in and not originating in the neural crest. We have examined the sensitivity of cultured skin fibroblasts from patients with neurofibromatosis to several types of DNA damage. Fibroblasts in Dulbecco's modified Eagle's medium were plated at 10(2) to 2 X 10(4) cells per 75 cm2 tissue culture plates, and exposed to various doses of gamma radiation (leads to DNA scission), actinomycin D, or mitomycin C. Cells were reincubated for 15 to 40 days until surviving colonies exhibited greater than 30-50 cells. Plates were then stained with 1% methylene blue and the colonies counted, with surviving fraction determined relative to plating efficiency. Nine skin fibroblast cell strains from normal individuals were studied as controls. One neurofibromatosis (NF) cell strain, SB23, exhibited normal sensitivity to all three DNA-damaging agents studied in early (7-8) and middle (12-13) in vitro passage. Strain GM0622, on the other hand, exhibited normal sensitivity to the three DNA-damaging agents studied at early passage, but showed a significant decrease in survival after exposure to both gamma radiation (D0 = 106 rad) and actinomycin D (D0 = 0.024 mcg/ml) with increasing passage. Strain GM1639 exhibited decreased survival after actinomycin D exposure at early passage (D0 = 0.017 mcg/ml), with normal survival after exposure to gamma radiation and mitomycin C at the same passage

  14. Relationship between DNA double-strand breaks, cell killing, and fibrosis studied in confluent skin fibroblasts derived from breast cancer patients

    DEFF Research Database (Denmark)

    Dikomey, E; Brammer, I; Johansen, Jørgen; Bentzen, Søren M; Overgaard, J

    2000-01-01

    To investigate the relationship between DNA double-strand breaks (dsbs), cell killing, and fibrosis using skin fibroblasts derived from breast cancer patients who received postmastectomy radiotherapy.......To investigate the relationship between DNA double-strand breaks (dsbs), cell killing, and fibrosis using skin fibroblasts derived from breast cancer patients who received postmastectomy radiotherapy....

  15. Mutagenicity of 8-methoxypsoralen and long-wave ultraviolet irradiation in diploid human skin fibroblasts

    International Nuclear Information System (INIS)

    Cell killing and the induction of mutation were studied in dividing and non-dividing human skin fibroblasts as a result of treatment by 8-methoxypsoralen (8-MOP) and long-wave UV irradiation (UVA). The cytotoxic effect was highly dependent upon the duration of the UVA exposure. The frequency of mutations increased linearly with the UVA dose at concentrations of 10 and 0.25 μl 8-MOP/ml, the latter representing the concentration in the skin during PUVA treatment. The number of mutations induced per unit dose (= per μg 8-MOP/ml per joule UVA/m2) was calculated: for dividing cells this value was 3.3 x 10-8 per cell and for non-dividing cells 0.6 x 10.8-8 per cell. On the basis of these values the expected number of induced mutants in the human skin per session of photochemotherapy is 1.2 x 10-5, and per 30 years of maintenance therapy 1.3 x 10-2 per cell. A comparison was made between this frequency and the frequency to be expected from spontaneous mutation. In addition the significance of absence in patients of SCE induction by photochemotherapy is discussed. (orig.)

  16. Effects of bosentan on collagen type I synthesis on in vitro culture of scleroderma skin fibroblasts

    Directory of Open Access Journals (Sweden)

    S. Soldano

    2011-01-01

    Full Text Available The present study evaluated the effects of a non-selective endothelin (ETA/B receptors antagonist, on collagen type I (COLI synthesis on in vitro culture of scleroderma (SSc skin fibroblasts (Fb. Fb were obtained from skin biopsies of 6 female SSc patients (mean age 64. 1±6 years, after informed consent and Ethical Committee Approval. Cells were treated with endothelin-I [ET-I, 100nM] for 24 and 48 hrs, pre-treated for I hr with ETA/B receptors antagonist [10nM] alone or followed by ET-I for 24 and 48 hrs. Untreated Fb were used as controls. Immunocytochemistry and western blot analysis were performed to evaluate COLI synthesis. ET-I increased COLI synthesis both at 24 and 48 hrs when compared to controls. ETA/B receptor antagonost blocks the increased COLI synthesis ET-I-mediated both at 24 and 48 hrs vs. ET-I. Results showed that ET-I receptors blockage by ETA/B receptors antagonist might prevent the excessive synthesis of COLI, supporting its positive action in the management of skin fibrosis.

  17. Immortalized human skin fibroblast feeder cells support growth and maintenance of both human embryonic and induced pluripotent stem cells

    OpenAIRE

    Unger, C; S. Gao; Cohen, M; Jaconi, M; Bergstrom, R; Holm, F.; A. Galan; Sanchez, E.; Irion, O; Dubuisson, J B; Giry-Laterriere, M; Salmon, P; Simon, C.; O Hovatta; Feki, A

    2009-01-01

    BACKGROUND: Feeder cells are frequently used for the early-stage of derivation and culture of human embryonic stem cell (hESC) lines. METHODS: We established a conditionally immortalized human foreskin fibroblast line that secreted basic fibroblast growth factor (bFGF). These cells were used as feeder cells for hESC culture and induced pluripotent stem (iPS) cell derivation and expansion. This conditional immortalization was performed using lentiviral vector (LV) mediated transduction of Bmi-...

  18. Mesenchymal stem cell-conditioned medium accelerates skin wound healing: An in vitro study of fibroblast and keratinocyte scratch assays

    International Nuclear Information System (INIS)

    We have used in vitro scratch assays to examine the relative contribution of dermal fibroblasts and keratinocytes in the wound repair process and to test the influence of mesenchymal stem cell (MSC) secreted factors on both skin cell types. Scratch assays were established using single cell and co-cultures of L929 fibroblasts and HaCaT keratinocytes, with wound closure monitored via time-lapse microscopy. Both in serum supplemented and serum free conditions, wound closure was faster in L929 fibroblast than HaCaT keratinocyte scratch assays, and in co-culture the L929 fibroblasts lead the way in closing the scratches. MSC-CM generated under serum free conditions significantly enhanced the wound closure rate of both skin cell types separately and in co-culture, whereas conditioned medium from L929 or HaCaT cultures had no significant effect. This enhancement of wound closure in the presence of MSC-CM was due to accelerated cell migration rather than increased cell proliferation. A number of wound healing mediators were identified in MSC-CM, including TGF-β1, the chemokines IL-6, IL-8, MCP-1 and RANTES, and collagen type I, fibronectin, SPARC and IGFBP-7. This study suggests that the trophic activity of MSC may play a role in skin wound closure by affecting both dermal fibroblast and keratinocyte migration, along with a contribution to the formation of extracellular matrix.

  19. Cerebrohepatorenal Syndrome (CHRS) of Zellweger: lysosomal enzyme activities, sulfation of glycosaminoglycans, and pipecolic acid levels in cultured skin fibroblasts

    International Nuclear Information System (INIS)

    The defect in the cerebrohepatorenal syndrome (CHRS), a fatal hereditary disorder primarily affecting neurological development, is unknown. Three areas were studied for specific biochemical abnormalities which might aid in diagnosis and understanding of the disorder: (1) Clinico-pathological similarities to inherited degenerative neurologic disorders suggested decreased activity of certain lysosomal enzymes. Assays of ?-galactosidase, ?-hexosaminidase, ?-mannosidase, and arylsulfatase A activities in fibroblasts from four infants with CHRS indicated no deficiency of enzyme activities. (2) Undersulfation of glycosaminoglycans (GAGs) has been reported in patients with the clinically similar Lowe's syndrome. The rate and amount of incorporation of 35SO4 = into intracellular 35S-GAGs up to 48 hours was comparable in fibroblasts from six CHRS infants and controls. Loss of 35-GAGs also followed a normal pattern. (3) Because pipecolic acid (PA) has been reported to be elevated in body fluids of patients with CHRS, cultured skin fibroblasts were examined for such an abnormality. Lysosomal enzyme activities and metabolism of sulfated glycosaminoglycans appear to be normal in cultured skin fibroblasts from infants with CHRS. Despite the sensitivity of the method, examination of pipecolic acid in cultured skin fibroblasts does not seem to be useful for diagnosis of CHRS

  20. Epigenetic conversion of adult dog skin fibroblasts into insulin-secreting cells.

    Science.gov (United States)

    Brevini, T A L; Pennarossa, G; Acocella, F; Brizzola, S; Zenobi, A; Gandolfi, F

    2016-05-01

    Diabetes is among the most frequently diagnosed endocrine disorder in dogs and its prevalence continues to increase. Medical management of this pathology is lifelong and challenging because of the numerous serious complications. A therapy based on the use of autologous viable insulin-producing cells to replace the lost β cell mass would be very advantageous. A protocol to enable the epigenetic conversion of canine dermal fibroblasts, obtained from a skin biopsy, into insulin-producing cells (EpiCC) is described in the present manuscript. Cells were briefly exposed to the DNA methyltransferase inhibitor 5-azacytidine (5-aza-CR) in order to increase their plasticity. This was followed by a three-step differentiation protocol that directed the cells towards the pancreatic lineage. After 36 days, 38 ± 6.1% of the treated fibroblasts were converted into EpiCC that expressed insulin mRNA and protein. Furthermore, EpiCC were able to release insulin into the medium in response to an increased glucose concentration. This is the first evidence that generating a renewable autologous, functional source of insulin-secreting cells is possible in the dog. This procedure represents a novel and promising potential therapy for diabetes in dogs. PMID:27033591

  1. Establishment, characterization, and toxicological application of loggerhead sea turtle (Caretta caretta) primary skin fibroblast cell cultures.

    Science.gov (United States)

    Webb, Sarah J; Zychowski, Gregory V; Bauman, Sandy W; Higgins, Benjamin M; Raudsepp, Terje; Gollahon, Lauren S; Wooten, Kimberly J; Cole, Jennifer M; Godard-Codding, Cline

    2014-12-16

    Pollution is a well-known threat to sea turtles but its impact is poorly understood. In vitro toxicity testing presents a promising avenue to assess and monitor the effects of environmental pollutants in these animals within the legal constraints of their endangered status. Reptilian cell cultures are rare and, in sea turtles, largely derived from animals affected by tumors. Here we describe the full characterization of primary skin fibroblast cell cultures derived from biopsies of multiple healthy loggerhead sea turtles (Caretta caretta), and the subsequent optimization of traditional in vitro toxicity assays to reptilian cells. Characterization included validating fibroblast cells by morphology and immunocytochemistry, and optimizing culture conditions by use of growth curve assays with a fractional factorial experimental design. Two cell viability assays, MTT and lactate dehydrogenase (LDH), and an assay measuring cytochrome P4501A (CYP1A) expression by quantitative PCR were optimized in the characterized cells. MTT and LDH assays confirmed cytotoxicity of perfluorooctanoic acid at 500 ?M following 72 and 96 h exposures while CYP1A5 induction was detected after 72 h exposure to 0.1-10 ?M benzo[a]pyrene. This research demonstrates the validity of in vitro toxicity testing in sea turtles and highlights the need to optimize mammalian assays to reptilian cells. PMID:25384208

  2. Effect of Fe3O4 Nanoparticles on Skin Tumor Cells and Dermal Fibroblasts

    OpenAIRE

    Lirija Alili; Swetlana Chapiro; Gernot U. Marten; Schmidt, Annette M.; Klaus Zanger; Peter Brenneisen

    2015-01-01

    Iron oxide (Fe3O4) nanoparticles have been used in many biomedical approaches. The toxicity of Fe3O4 nanoparticles on mammalian cells was published recently. Though, little is known about the viability of human cells after treatment with Fe3O4 nanoparticles. Herein, we examined the toxicity, production of reactive oxygen species, and invasive capacity after treatment of human dermal fibroblasts (HDF) and cells of the squamous tumor cell line (SCL-1) with Fe3O4 nanoparticles. These nanoparticl...

  3. Cytotoxic and Oxidative Stress Caused by Cadmium and Lead on Human Skin Fibroblast Cells

    Directory of Open Access Journals (Sweden)

    Ali Beman Zaree Mahmodabady

    2006-01-01

    Full Text Available Introduction: Heavy metals are important occupational andenvironmental pollutants that cause damage to various organs.Although there is no effective therapy for such a poisoning,metallothionein has been shown to play a key role in thedetoxification of cadmium (Cd. Evidence in the literature suggeststhat superoxide dismutase, glutathione peroxidase, and catalaseconstitute important defense mechanisms against oxygen toxicity inthe cells. The aim of this study was to investigate the effect ofcadmium chloride and Pb-acetate on antioxidant enzymes in thehuman skin fibroblast cells (HF2FF.Material and Methods: The human skin fibroblast (HF2FF cellswere incubated in serum-free medium containing 20 μM CdCl2 for18 hr three times a week. The same exposure to an equimolar doseof Pb-acetate was performed. After each exposure and after threetimes exposure the cells were collected and cell viability, thecontents of superoxide dismutase (SOD, catalase, glutathioneperoxidase (GSH-Px, GSH and malondialdehyde (MDA weremeasured.Results: Cd caused cytotoxicity and inhibition of glutathioneperoxidase (GSH-Px and SOD activity, as well as depletion of thereduced form of glutathione (GSH in the cell. The level of lipidperoxidation (LP was increased, but catalase activity was notsignificantly altered. These defects were increased with repeatedexposures. The same exposure to an equimolar dose of Pb-acetateevoked only inhibition of GSH-Px and SOD. The values of GSH,catalase and LP activity remained unchanged.Conclusion: The inhibition of GSH-Px and SOD may be consideredas an important biomarker of the toxic effect of metals.

  4. Dramatic increase in oxidative stress in carbon-irradiated normal human skin fibroblasts

    International Nuclear Information System (INIS)

    Skin complications were recently reported after carbon-ion (C-ion) radiation therapy. Oxidative stress is considered an important pathway in the appearance of late skin reactions. We evaluated oxidative stress in normal human skin fibroblasts after carbon-ion vs. X-ray irradiation. Survival curves and radiobiological parameters were calculated. DNA damage was quantified, as were lipid peroxidation (LPO), protein carbonylation and antioxidant enzyme activities. Reduced and oxidized glutathione ratios (GSH/GSSG) were determined. Pro-inflammatory cytokine secretion in culture supernatants was evaluated. The relative biological effectiveness (RBE) of C-ions vs. X-rays was 4.8 at D0 (irradiation dose corresponding to a surviving fraction of 37%). Surviving fraction at 2 Gy (SF2) was 71.8% and 7.6% for X-rays and C-ions, respectively. Compared with X-rays, immediate DNA damage was increased less after C-ions, but a late increase was observed at D10% (irradiation dose corresponding to a surviving fraction of 10%). LPO products and protein carbonyls were only increased 24 hours after C-ions. After X-rays, superoxide dismutase (SOD) activity was strongly increased immediately and on day 14 at D0% (irradiation dose corresponding to a surviving fraction of around 0%), catalase activity was unchanged and glutathione peroxidase (GPx) activity was increased only on day 14. These activities were decreased after C-ions compared with X-rays. GSH/GSSG was unchanged after X-rays but was decreased immediately after C-ion irradiation before an increase from day 7. Secretion of IL-6 was increased at late times after X-ray irradiation. After C-ion irradiation, IL-6 concentration was increased on day 7 but was lower compared with X-rays at later times. C-ion effects on normal human skin fibroblasts seemed to be harmful in comparison with X-rays as they produce late DNA damage, LPO products and protein carbonyls, and as they decrease antioxidant defences. Mechanisms leading to this discrepancy between the two types of radiation should be investigated. (authors)

  5. Age-associated reduction of cell spreading induces mitochondrial DNA common deletion by oxidative stress in human skin dermal fibroblasts: implication for human skin connective tissue aging

    OpenAIRE

    Quan, Chunji; Cho, Moon Kyun; Perry, Daniel; Quan, Taihao

    2015-01-01

    Background Reduced cell spreading is a prominent feature of aged dermal fibroblasts in human skin in vivo. Mitochondrial DNA (mtDNA) common deletion has been reported to play a role in the human aging process, however the relationship between age-related reduced cell spreading and mtDNA common deletion has not yet been reported. Results To examine mtDNA common deletion in the dermis of aged human skin, the epidermis was removed from full-thickness human skin samples using cryostat. mtDNA comm...

  6. A quantitative assessment of changes in the dermal fibroblast population of pig skin after single doses of X-rays

    International Nuclear Information System (INIS)

    Changes in the density of fibroblast nuclei in reticular dermis of pigs was studied from 6 to 104 weeks after a single dose of 15.4 Gy of X-rays. The largest decrease in fibroblasts occurred between 12 and 26 weeks after irradiation; after this there was only a slight fall in fibroblast number until 104 weeks when observations ceased. At 26 weeks and later times after irradiation reduction in the density of fibroblast nuclei in the reticular dermis was dose-dependent for single doses in the range 8.0-20.7 Gy. The dose-response curve had an initial shoulder, after which the fall in the fibroblast nuclear density was linearly related to dose. Data obtained between 26 weeks and 104 weeks after irradiation, could be fitted by the same dose-response curve. The fall in the counts of fibroblast nuclei was compared with earlier studies. The loss of fibroblasts occurred after an initial reduction in blood flow in the pig skin but was concomitant with general reduction in dermal thickness. (author)

  7. Radiation-induced comet-formation in human skin fibroblasts from radiotherapy patients with different normal tissue reactions

    International Nuclear Information System (INIS)

    Background: In clinical radiotherapy most patients tolerate the applied dosage with no or moderate side effects. However, 5 to 10% of all individuals show increased acute and/or late reactions. In-vitro test systems are investigated for their suitability for predictive purposes. This paper attempts a correlation between the induction and repair of DNA damage measured in the comet assay and the clinical observed reaction in order to evaluate the suitability of the comet assay for prediction of radiation sensitivity. Patients and methods: Skin fibroblasts of 30 patients with average tissue reactions or acute and/or late increased side effects and cell lines of 4 individuals carrying the heritable disease ataxia telangiectasia (AT) were irradiated in vitro. The induction and repair of DNA damage was measured at different time points after irradiation in the comet assay (single cell gel electrophoresis). These results were compared to the acute and late clinical reactions classified according to the RTOG grading system. Results: The radiation induced DNA damage decreased over time reflecting DNA repair. Cells of the AT individuals showed and elevated damage induction and a reduced repair capacity compared to patients with average tissue reactions. Fibroblast of patients with increased acute and late side effects exhibited slower DNA repair. In addition to the known lack of cell cycle control, our results indicate that AT cells show reduced DNA repair capacity. Conclusions: The comet assay seems to be able to detect some types of increased individual radiation sensitivity. In contrast to other predictive in-vitro tests, the comet assay needs less time and fewer cells, which would be useful in a clinical setting. (orig.)

  8. Aloin Protects Skin Fibroblasts from Heat Stress-Induced Oxidative Stress Damage by Regulating the Oxidative Defense System

    Science.gov (United States)

    Wang, Yu-Ren; Tsai, Hsin-I; Yu, Huang-Ping

    2015-01-01

    Oxidative stress is commonly involved in the pathogenesis of skin damage induced by environmental factors, such as heat stress. Skin fibroblasts are responsible for the connective tissue regeneration and the skin recovery from injury. Aloin, a bioactive compound in Aloe vera, has been reported to have various pharmacological activities, such as anti-inflammatory effects. The aim of this study was to investigate the protective effect of aloin against heat stress-mediated oxidative stress in human skin fibroblast Hs68 cells. Hs68 cells were first incubated at 43°C for 30 min to mimic heat stress. The study was further examined if aloin has any effect on heat stress-induced oxidative stress. We found that aloin protected Hs68 cells against heat stress-induced damage, as assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and lactate dehydrogenase assay. Aloin protected Hs68 cells by regulating reactive oxygen species production and increasing the levels of glutathione, cytosolic and mitochondrial superoxide dismutase. Aloin also prevented the elevation of thiobarbituric acid reactive substances and the reduction of 8-OH-dG induced by heat stress. These results indicated that aloin protected human skin fibroblasts from heat stress-induced oxidative stress damage by regulating the oxidative defense system. PMID:26637174

  9. Electrically Activated Primary Human Fibroblasts Improve In Vitro and In Vivo Skin Regeneration.

    Science.gov (United States)

    Rouabhia, Mahmoud; Park, Hyun Jin; Zhang, Ze

    2016-08-01

    Electrical stimulation (ES) changes cellular behaviors and thus constitutes a potential strategy to promote wound healing. However, well-controlled in vitro findings have yet to be translated to in vivo trials. This study was to demonstrate the feasibility and advantages of transplanting electrically activated cells (E-Cells) to help wound healing. Primary human skin fibroblasts were activated through well defined ES and cultured with keratinocytes to generate engineered human skin (EHS), which were transplanted to nu/nu mice. The electrically activated EHS grafts were analyzed at 20 and 30 days post-grafting, showing faster wound closure, thick epidermis, vasculature, and functional basement membrane containing laminin and type IV collagen that were totally produced by the implanted human cells. Because a variety of cells can be electrically activated, E-Cells may become a new cell source and the transplantation of E-Cells may represent a new strategy in wound healing and tissue engineering. J. Cell. Physiol. 231: 1814-1821, 2016. © 2015 Wiley Periodicals, Inc. PMID:26661681

  10. The protective effects of fucosterol against skin damage in UVB-irradiated human dermal fibroblasts.

    Science.gov (United States)

    Hwang, Eunson; Park, Sang-Yong; Sun, Zheng-wang; Shin, Heon-Sub; Lee, Don-Gil; Yi, Tae Hoo

    2014-06-01

    Exposure to ultraviolet (UV) light causes matrix metalloproteinase (MMP) overexpression and extracellular matrix depletion, leading to skin photoaging. The activation of MMP is related to increased interlukin-6 (IL-6) and type I procollagen production, which is regulated by transforming growth factor-β1 (TGF-β1). Activator protein-1 (AP-1) activation induces MMP-1 production and reduces type I procollagen secretion. Fucosterol, which is extracted and purified from the brown algae Hizikia fusiformis, is a phytosterol. We assessed the effects of fucosterol on photodamage and investigated its molecular mechanism of action in UVB-irradiated normal human dermal fibroblasts by using enzyme-linked immunosorbent assay, Western blot analysis, and reverse transcription-polymerase chain reaction. Our results showed that fucosterol significantly decreased the UVB-induced expression of MMP-1, IL-6, p-c-Jun, and p-c-Fos. Additionally, fucosterol markedly increased the UVB-induced production of type I procollagen and TGF-β1. Our results indicate that fucosterol regulates MMP-1 and type I procollagen expression by modulating AP-1 and TGF-β1 signaling and that MMP-1 activation is correlated with IL-6. These data suggest that fucosterol is a promising botanical agent to protect against skin photodamage. PMID:24142195

  11. 3,3'-Dihydroxyisorenieratene and isorenieratene prevent UV-induced DNA damage in human skin fibroblasts.

    Science.gov (United States)

    Wagener, Sarah; Völker, Tanja; De Spirt, Silke; Ernst, Hansgeorg; Stahl, Wilhelm

    2012-08-01

    Skin cancer is among the most frequent neoplastic malignancies and exposure to UV irradiation is a major risk factor. In addition to topical sunscreens, photoprotection by dietary antioxidants such as carotenoids or polyphenols has been suggested as a means of prevention. Isorenieratene (IR) and dihydroxyisorenieratene (DHIR) are aromatic carotenoids with particular antioxidant properties produced by Brevibacterium linens. The aim of this study was to investigate the photoprotective and antioxidant activities of DHIR and IR in comparison to the nonaromatic carotenoid lutein in human dermal fibroblasts. Incubation of the cells with DHIR and IR significantly decreased the UV-induced formation of cyclobutane pyrimidine dimers and formation of DNA strand breaks. Lipid oxidation was lowered as determined by the formation of malondialdehyde as a biomarker. Both aromatic carotenoids also prevented oxidatively generated damage to DNA as demonstrated by a decrease in DNA strand breaks associated with the formation of oxidized DNA bases. These data highlight the multifunctional photoprotective properties of aromatic carotenoids, which may be suitable natural compounds for the prevention of skin cancer. PMID:22634149

  12. Biochemical mechanisms of skin radiation burns inhibition and healing by the volumetric autotransplantation of fibroblasts and of keratinocytes with fibroblasts composition

    Directory of Open Access Journals (Sweden)

    L. V. Altukhova

    2015-09-01

    Full Text Available Mechanisms of influence of volumetric autotransplantation of fibroblasts and of the mixture of fibroblasts and keratinocytes on the development of the local 3rd degree X-ray burn and the radiation skin ulcer in guinea pigs were investigated. We used deepadministration into the irradiation zone on its perimeter of 6 doses, which contained (150–160×103 fibroblasts and (130–140×103 keratinocytes in 100 µl. It is shown that this autotransplantation carried out 1 hour after the irradiation, and then every 24 hours, reduces the area of burn on the 35th day, compared to the control by 63%. Radiation ulcer appears on the 10th day after irradiation and is completely healed on the 25th day. With the same regimen of administration of only fibroblasts containing (200–210×103 cells in 100 µl, these parameters of treatment were equal to 31% on 4th and 35th day, respectively. It is shown that as a result of radiation in the area of burn the level of gene expression of collagen types I and III, elastin, fibronectin, vinculin, decorin, hyaluronansynthases 1, 2, 3, matrix metalloproteinases 1, 2, 3, 7, 9 and hyaluronidase is reduced. Besides, in the burn area the level of gene expression of transforming growth factor α, fibroblast growth factors 1, 2, 8 and anti-inflammatory cytokines – interleukin 10 and transforming growth factor-β1 – is reduced, while the level of gene expression of proinflammatory cytokine (interleykin1β increases. Both types of autotransplantation cause the growth of the expression level of all the structural genes and regulatory proteins of biopolymers and decrease in the expression level of interleukin 1β, which leads to activation of tissue regeneration and healing of the burn wound. Reasonsfor the higher efficiency of autotransplantation using the mixture of fibroblasts and keratinocytes compared to autotransplantation by fibroblasts only are both the larger total number of live cells regularly replacing dead cells in the burn area, and mutual stimulation of auto-fibroblasts and auto-keratinocytes to proliferate and to synthesize biologically active substances, i.e. cytokines and growth factors.

  13. Influence of hyperthermia on the phosphorylation of ribosomal protein S6 from human skin fibroblasts and meningioma cells

    DEFF Research Database (Denmark)

    Richter, W W; Zang, K D; Issinger, O G

    1983-01-01

    Skin fibroblasts and meningioma cells, derived from primary cultures of the same patients have been used to study the influence of hyperthermia on (i) cell morphology and (ii) phosphorylation pattern of ribosomal and ribosome-associated proteins. Incubation of tumour cells and fibroblasts up to 7 h...... at 42 degrees C did not significantly change the cell morphology as compared to control cells kept at 37 degrees C. At 42 degrees C ribosomal protein S6 is shifted cathodically indicating a loss of negative charge, however no quantitative dephosphorylation of S6 was observed. Meningioma cells and...

  14. Relationship between the in vitro radiosensitivity of skin fibroblasts and the expression of subcutaneous fibrosis, telangiectasia, and skin erythema after radiotherapy

    International Nuclear Information System (INIS)

    Objective: To investigate if the occurrence of subcutaneous fibrosis after radiotheraphy in an unselected group of breast cancer patients is related to cellular radiosensitivity of skin fibroblasts as measured in a clonogenic assay. Materials and methods: An in vitro colony-forming assay of normal fibroblast radiosensitivity was applied to primary skin biopsies from 31 breast cancer patients who received post-mastectomy radiotherapy with large doses per fraction (2.7-3.9 Gy) more than 10 years earlier. Three clinical normal-tissue endpoints were assessed. Two late endpoints, subcutaneous fibrosis and telangiectasia, were evaluated in three treatments fields by a single experienced clinician. In addition, skin erythema had been assessed at the end of treatment by members of the staff and junior staff. >From previous analyses of normal tissue response, individual clinical radiosensitivity could be assessed as 'excess risk' of each of the three reactions. This was defined as the difference between the actual observed response in the patient and the expected response estimated from individual treatment characteristics in a linear quadratic (LQ) mixture model and, for the two late endpoints, with correction for the follow-up time. This clinical radioresponsiveness was compared with the in vitro radiosensitivity of the skin fibroblasts. To this end, the fractions of colony-forming cells after graded single doses were fitted by an LQ survival curve using non-linear and linear regression from which the surviving fraction at 3.5 Gy (SF3.5) was estimated. Assessment at 3.5 Gy was chosen to reflect the fraction size during clinical radiotherapy. Results: A statistically significant variability of in vitro radiosensitivity between patients could be detected for both SF2 (P = 0.0095) and SF3.5 (P = 0.0008). A significant correlation was observed between SF3.5 and excess risk of fibrosis (rs -0.46, P = 0.009) while no association was found between fibroblast radiosensitivity and either the occurrence of severe skin telangiectasia or the acute endpoint skin erythema. Conclusion: These results suggest that variability in the occurrence of subcutaneous fibrosis, but not telangiectasia or erythema, after radiotherapy is partly accounted for by differences in cellular radiosensitivity of normal skin fibroblasts

  15. Generation of Transgenic Porcine Fibroblast Cell Lines Using Nanomagnetic Gene Delivery Vectors.

    Science.gov (United States)

    Grześkowiak, Bartosz F; Hryhorowicz, Magdalena; Tuśnio, Karol; Grzeszkowiak, Mikołaj; Załęski, Karol; Lipiński, Daniel; Zeyland, Joanna; Mykhaylyk, Olga; Słomski, Ryszard; Jurga, Stefan; Woźniak, Anna

    2016-05-01

    The transgenic process allows for obtaining genetically modified animals for divers biomedical applications. A number of transgenic animals for xenotransplantation have been generated with the somatic cell nuclear transfer (SCNT) method. Thereby, efficient nucleic acid delivery to donor cells such as fibroblasts is of particular importance. The objective of this study was to establish stable transgene expressing porcine fetal fibroblast cell lines using magnetic nanoparticle-based gene delivery vectors under a gradient magnetic field. Magnetic transfection complexes prepared by self-assembly of suitable magnetic nanoparticles, plasmid DNA, and an enhancer under an inhomogeneous magnetic field enabled the rapid and efficient delivery of a gene construct (pCD59-GFPBsd) into porcine fetal fibroblasts. The applied vector dose was magnetically sedimented on the cell surface within 30 min as visualized by fluorescence microscopy. The PCR and RT-PCR analysis confirmed not only the presence but also the expression of transgene in all magnetofected transgenic fibroblast cell lines which survived antibiotic selection. The cells were characterized by high survival rates and proliferative activities as well as correct chromosome number. The developed nanomagnetic gene delivery formulation proved to be an effective tool for the production of genetically engineered fibroblasts and may be used in future in SCNT techniques for breeding new transgenic animals for the purpose of xenotransplantation. PMID:27048425

  16. Double trisomy mosaic (47,XXX/48,XXX,+13) confirmed by FISH and skin fibroblast culture

    Energy Technology Data Exchange (ETDEWEB)

    Lieber, E.; Grady, V.; Dosik, H. [Interfaith Medical Center, Brooklyn, NY (United States)] [and others

    1994-09-01

    A 4 lb 8 oz female was born to a 49-year-old woman (P1200G12) at 40 weeks. The baby had tetralogy of Fallot, polydactyly, microcephaly, low set simple ears, posterior cleft of the soft palate and overlapping flexion deformities of both hands. The eyes were deep set. The clinical impression was trisomy 13. The baby is not doing well and needs a gastrotomy tube for feeding. Sucking is allright but swallowing is impeded. An MRI showed an anomaly of the corpus callosum. The ophthalmological examination showed no abnormalities. A chromosome study on a 2-day peripheral blood sample resulted in poor growth and poor morphology; however, 20 Giemsa-banded cells revealed a 47,XXX karyotype. A second specimen was obtained to search for mosaicism and a blood smear revealed nuclear projections on the neutrophils. FISH analysis using whole chromosome painting probe (Life Technologies) first identified the extra chromosome number 13, the final results showing five of sixty metaphase cells (8.3%) with trisomy 13. Cytogenetic analysis using Giemsa-banding technique revealed four cells in fifty examined (8.0%) with a 48,XXX,+13 karyotype. In order to further evaluate the mosaicism, cytogenetic analysis of a skin fibroblast culture was performed. Twenty one of twenty three cells examined (91.3%) showed the 48,XXX,+13 karyotype. FISH analysis of the skin biopsy revealed eighteen of twenty cells (90.9%) with the trisomy 13. The FISH technique is an important enhancement to routine cytogenetic studies when they do not immediately correlate with clinical impressions.

  17. Thyroid hormone excess stimulates the synthesis of proteoglycan in human skin fibroblasts in culture

    International Nuclear Information System (INIS)

    We previously demonstrated that proteoglycan accumulated in the affected skin of circumscribed pretibial myxedema of Graves' disease. As an underlying mechanism responsible for the accumulation, we sought to determine whether excess thyroid hormone was partially responsible for the increase in proteoglycan synthesis. Human skin fibroblasts were cultured in Ham's F-10 medium containing 1% Nutridoma with graded doses of T3(0.184 x 10-9 to 46 x 10-9 mol/l) and were labelled with [35S]sulphate and [3H]glucosamine. Proteoglycans were purified by Sephadex G-50, Q-Sepharose chromatography with NaCl-gradient and Sepharose CL-6B chromatography. 35S and 3H incorporated into dermatan sulphate proteoglycan and heparan sulphate proteoglycan and 3H incorporated into hyaluronan were measured. 35S and 3H incorporation into dermatan sulphate proteoglycan was minimum at a T3 concentration of 0.184 x 10-9 mol/l, and increased with increasing doses of T3 up to 46 x 10-9 mol/l. 35S and 3H incorporation into heparan sulphate proteoglycan also increased with increasing-doses of T3. 3H incorporation into hyaluranan was not influenced at all by T3. The increased incorporation of 35S into proteoglycan in high-T3 culture reflects the increased synthesis of proteoglycan because 1. the extent of sulphation of disaccharides examined by thin-layer chromatography was not altered by T3; 2. the specific activity of [35S]sulphate was not influenced by T3, and 3. T3 did not decrease the degradation rate of cell-associated proteoglycan. (author)

  18. Extracellular Matrix Modulates Morphology, Growth, Oxidative Stress Response and Functionality of Human Skin Fibroblasts during Aging In Vitro

    DEFF Research Database (Denmark)

    Jørgensen, Peter; Rattan, Suresh

    2014-01-01

    The Hayflick system of cellular aging and replicative senescence in vitro has been used widely in both basic and applied research in biogerontology. The state of replicative senescence is generally considered to be irreversible, but is modifiable by genetic and environmental manipulations. Some...... recent observations indicate that replicative lifespan, senescence and functionality of cells in vitro can be significantly affected by the quality of the extra cellular matrix (ECM). Following up on those reports, here we show that using the ECM prepared from early passage young cells, partial...... rejuvenation of serially passaged human facial skin fibroblasts was possible in pre-senescent middle-aged cells, but not in fully senescent late passage cells. ECM from young cells improved the appearance, viability, stress tolerance and wound healing ability of skin fibroblasts. Furthermore, young ECM...

  19. Growth and motility of human skin fibroblasts on multilayer strong polyelectrolyte films.

    Science.gov (United States)

    Wytrwal, Magdalena; Koczurkiewicz, Paulina; Zrubek, Karol; Niemiec, Wiktor; Michalik, Marta; Kozik, Bartłomiej; Szneler, Edward; Bernasik, Andrzej; Madeja, Zbigniew; Nowakowska, Maria; Kepczynski, Mariusz

    2016-01-01

    Polyelectrolyte multilayers (PEMs) have found application in modifying material surfaces to make them adhesive or non-adhesive for animal cells. However, PEMs made of strong polyelectrolytes are not fully recognized in the literature. This study focuses on the interplay between the properties of PEM assembled from strong polyelectrolytes and cell adhesion and motility. Strong polycations (with quaternary ammonium groups) and a polyanion (with sulfonate groups) were obtained by modification of poly(allylamine hydrochloride) (PAH). Two types of multilayer films were assembled from these PAH derivatives and used to investigate the behavior of human skin fibroblasts (HSFs). The effect of surface charge, hydrophobicity, and film thickness on adhesion of HSFs in a serum-containing medium was studied with immunofluorescence microscopy. The results showed that adhesion of HSFs was strongly depended on the chemical functions of the terminal layer, whereas the wettability was not important. The surface of PEM can be strongly cytophobic (the quaternary ammonium terminal groups) or strongly cytophilic (the sulfonate terminal groups). Finally, the motile activity of HSFs seeded on glass coated with a varying number of polymer layers was investigated. It was demonstrated using an in vitro model that coating the substrate with only two polymer layers can considerably increase the average speed of HSFs movement and stimulate cell migration into the wound. PMID:26407058

  20. Radiosensitivity of skin fibroblasts from atomic bomb survivors with and without breast cancer

    International Nuclear Information System (INIS)

    Fibroblasts were established in vitro from skin biopsies obtained from 55 women and one man with or without breast cancer and with or without exposure to radiation from the atomic bomb explosion in Hiroshima. The radiosensitivity of these cells was evaluated by clonogenic assays after exposure to X rays or to fission neutrons from a 252Cf source. Data were fitted to a multitarget model, S/S0 = A[1-(1-ekD)N], for both X-ray and neutron dose-survival curves. A single-hit model, S/S0 = AekD, fits the neutron dose-survival responses as well. These was no difference in the means or variances of radiosensitivity between exposed and nonexposed groups, or between patients with or without breast cancer. Hence, although the sample is not large, it provides no support for the hypothesis that A-bomb radiation preferentially induces breast cancer in women whose cells in vitro are sensitive to cell killing by radiation. (author)

  1. DNA-protein crosslinking in normal human skin fibroblasts exposed to ultraviolet radiation

    International Nuclear Information System (INIS)

    Cultured normal human skin fibroblasts were exposed to different fluences of 254 nm UV and the levels of DNA-protein crosslinks (DPC) measured with alkaline elution immediately after irradiation or following a 24-hour incubation (370C). For cells exposed to 10J/m/sup 2/ and then incubated, the level of DPC decreased to that of unexposed cells. When the fluences increased, the levels of DPC measured following a 24-hour incubation increased as compared with non-incubated cells. At fluences higher than 100J/m/sup 2/, the DPC levels of incubated cells exceeded the DPC levels of non-incubated cells. When the single strand breaks (SSB) and double strand breaks (DSB) were measured under a deproteinized condition with alkaline elution and neutral elution, respectively, the levels of SSB and DSB were higher for cells with than for cells without post-irradiation incubation. The simultaneous increase of DPC and proteinase-sensitive SSB and DSB for cells given post-irradiation incubation suggests that a significant part of the DPC observed during post-UV-irradiation incubation were the DNA strand breaks that were tightly associated with proteins. A potential role for type II DNA topoisomerase in the formation of these DPC resulting from either the change in conformational structure caused by the presence of a high level of dimers or an involvement of this enzyme in dimer excision repair will be discussed

  2. Radiosensitivity of skin fibroblasts from atomic bomb survivors with and without breast cancer

    International Nuclear Information System (INIS)

    Fibroblasts were established in vitro from skin biopsies obtained from 55 women and 1 man with or without breast cancer and with or without exposure to radiation from the atomic bomb explosion in Hiroshima. The radiosensitivity of these cells was evaluated by clonogenic assays after exposure to X-rays or to fission neutrons from a 252Cf source. Data were fitted to a multitarget model, S/S0 = A [1 - (1 - ekD)N], for both X-ray and neutron dose-survival curves. A single hit model, S/S0 = AekD, fits the neutron dose-survival responses as well. There were no differences in the means or variances of radiosensitivity between exposed and nonexposed groups or between patients with or without breast cancer. Hence, although the sample is not large, it provides no support for the hypothesis that atomic bomb radiation preferentially induces breast cancer in women whose cells in vitro are sensitive to cell killing by radiation

  3. Skin substitutes based on allogenic fibroblasts or keratinocytes for chronic wounds not responding to conventional therapy: a retrospective observational study.

    Science.gov (United States)

    Pajardi, Giorgio; Rapisarda, Vicenzo; Somalvico, Francesco; Scotti, Andrea; Russo, Giulia Lo; Ciancio, Francesco; Sgr, Arturo; Nebuloni, Manuela; Allevi, Raffaele; Torre, Maria L; Trabucchi, Emilio; Marazzi, Mario

    2016-02-01

    Chronic wounds are an expression of underlying complex pathologies and have a high incidence. Skin substitutes may represent an alternative approach to treat chronic ulcers. The aim of this retrospective observational study was to evaluate the wound reduction using skin substitutes based on allogenic fibroblasts or keratinocytes in 30 patients not responding to conventional therapy. Wound bed was prepared, then keratinocytes on Laserskin() to treat superficial wounds or fibroblasts on Hyalograft 3D(R) to treat deep leg ulcers were applied, and finally wounds were treated with a secondary dressing composed of nanocrystalline silver. Once a week constructs were removed and new bioengineered products were applied, as well as nanocrystalline silver medication. In none of the cases under examination did any complications arise relating to the treatment. We also achieved a reduction in wound dimension and exudates, and an increase in wound bed score. Postoperative assessment shows a degree of healing that is statistically higher in the group treated with keratinocytes as compared with the fibroblast group. This retrospective study improves our understanding and defines the clinical indications for the various uses of the two types of skin substitutes. PMID:24517418

  4. Studies of the UVC-sensitivity of non-tumorigenic and tumorigenic human cell hybrids (HeLa x skin fibroblasts)

    International Nuclear Information System (INIS)

    The UVC-sensitivities of a non-tumorigenic and a tumorigenic human cell hybrid (HeLa x skin fibroblasts) are compared and contrasted. The tumorigenic cells differ from the non-tumorigenic cells in that they have lost one copy each of chromosomes 11 and 14. For exponentially growing cultures, the tumorigenic cells are considerably more resistant than the non-tumorigenic cells. For confluent cultures, the differential in photosensitivity is much less. Flow cytometric studies of cell cycle distributions of both exponentially growing and confluent cultures of these cells indicate that the differences in photosensitivity cannot be explained by differences in cell cycle distribution. Studies of the kinetics of potentially lethal damage repair (PLDR) in confluent cultures of both cell lines indicate little or no recovery over the first 6h followed by an increase in survival over the next 12-24h. These data are consistent with previously published observations in human skin fibroblasts where the kinetics of PLDR reflected the kinetics of thymine dimer loss. The data are not consistent with 6-4 photoproducts being a potentially lethal lesion since such damage is rapidly repaired in human cells. (author)

  5. Skin Equivalent Tissue-Engineered Construct: Co-Cultured Fibroblasts/ Keratinocytes on 3D Matrices of Sericin Hope Cocoons

    Science.gov (United States)

    Nayak, Sunita; Dey, Sancharika; Kundu, Subhas C.

    2013-01-01

    The development of effective and alternative tissue-engineered skin replacements to autografts, allografts and xenografts has became a clinical requirement due to the problems related to source of donor tissue and the perceived risk of disease transmission. In the present study 3D tissue engineered construct of sericin is developed using co-culture of keratinocytes on the upper surface of the fabricated matrices and with fibroblasts on lower surface. Sericin is obtained from “Sericin Hope” silkworm of Bombyx mori mutant and is extracted from cocoons by autoclave. Porous sericin matrices are prepared by freeze dried method using genipin as crosslinker. The matrices are characterized biochemically and biophysically. The cell proliferation and viability of co-cultured fibroblasts and keratinocytes on matrices for at least 28 days are observed by live/dead assay, Alamar blue assay, and by dual fluorescent staining. The growth of the fibroblasts and keratinocytes in co-culture is correlated with the expression level of TGF-β, b-FGF and IL-8 in the cultured supernatants by enzyme-linked immunosorbent assay. The histological analysis further demonstrates a multi-layered stratified epidermal layer of uninhibited keratinocytes in co-cultured constructs. Presence of involucrin, collagen IV and the fibroblast surface protein in immuno-histochemical stained sections of co-cultured matrices indicates the significance of paracrine signaling between keratinocytes and fibroblasts in the expression of extracellular matrix protein for dermal repair. No significant amount of pro inflammatory cytokines (TNF-α, IL-1β and nitric oxide) production are evidenced when macrophages grown on the sericin matrices. The results all together depict the potentiality of sericin 3D matrices as skin equivalent tissue engineered construct in wound repair. PMID:24058626

  6. Molecular alterations of tropoelastin and proteoglycans induced by tobacco smoke extracts and ultraviolet A in cultured skin fibroblasts

    International Nuclear Information System (INIS)

    Functional integrity of normal skin is dependent on the balance between the biosynthesis and degradation of extracellular matrix, primarily composed of collagen, elastin and proteoglycans. In our previous studies, we found that tobacco smoke extracts decreased expressions of type I and III procollagen and induced matrix metalloproteinase-1 (MMP-1) and MMP-3 in the cultured skin fibroblasts. We here further investigated the effects of tobacco smoke extracts or ultraviolet A (UVA) treatments on the expression of tropoelastin (soluble elastin protein), and versican and decorin (proteoglycans) in cultured skin fibroblasts. The mRNA of tropoelastin increased by tobacco smoke extracts or UVA irradiation. Versican was markedly shown to decrease after these treatments by using western blotting and the mRNA of versican V0 also significantly decreased. UVA treatment did not show remarkable change in decorin protein, but resulted in marked decrease of decorin D1 mRNA. In contrast to UVA irradiation, the treatments of tobacco smoke extracts resulted in significant increase in decorin, while mRNA of decorin D1 decreased as compared to the control. MMP-7 increased after the treatment of tobacco smoke extracts or UVA. These results indicated that common molecular features might underlie the skin premature aging induced by tobacco smoke extracts and UVA, including abnormal regulation of extracellular matrix deposition through elevated MMPs, reduced collagen production, abnormal tropoelastin accumulation, and altered proteoglycans. (author)

  7. Hypersensitivity of skin fibroblasts from basal cell nevus syndrome patients to killing by ultraviolet B but not by ultraviolet C radiation

    International Nuclear Information System (INIS)

    Basal cell nevus syndrome (BCNS) is an autosomal dominant genetic disorder in which the afflicted individuals are extremely susceptible to sunlight-induced skin cancers, particularly basal cell carcinomas. However, the cellular and molecular basis for BCNS is unknown. To ascertain whether there is any relationship between genetic predisposition to skin cancer and increased sensitivity of somatic cells from BCNS patients to killing by UV radiation, we exposed skin fibroblasts established from unexposed skin biopsies of several BCNS and age- and sex-matched normal individuals to either UV-B (280-320 nm) or UV-C (254 nm) radiation and determined their survival. The results indicated that skin fibroblasts from BCNS patients were hypersensitive to killing by UV-B but not UV-C radiation as compared to skin fibroblasts from normal individuals. DNA repair studies indicated that the increased sensitivity of BCNS skin fibroblasts to killing by UV-B radiation was not due to a defect in the excision repair of pyrimidine dimers. These results indicate that there is an association between hypersensitivity of somatic cells to killing by UV-B radiation and the genetic predisposition to skin cancer in BCNS patients. In addition, these results suggest that DNA lesions (and repair processes) other than the pyrimidine dimer are also involved in the pathogenesis of sunlight-induced skin cancers in BCNS patients. More important, the UV-B sensitivity assay described here may be used as a diagnostic tool to identify presymptomatic individuals with BCNS

  8. Repair of α- or X-radiation-induced DNA damage in cultured human skin fibroblasts

    International Nuclear Information System (INIS)

    The authors previously reported the RBE for cell killing and mutagenicity at the hgprt locus in cultured human skin fibroblasts (GM10) was 7 and 13-18, respectively, for high-LET /sup 238/Pu-emitted α-particles compared to low-LET 250 kVp X-rays. Dose fractionation studies indicated repair of sublethal and mutational damages in X-irradiated cells, whereas, this repair was not observed in cells exposed to α-radiation. In this report, the agents present data on radiation-induced DNA single-strand and double-strand breaks (ssb and dsb) and their repair in both proliferating and confluent monolayers of GM10 cells. Employing sensitive alkaline and neutral DNA filter elution analyses, rates of ssb induced by X-rays were 6-fold that observed for α-particles, whereas rates of dsb induced by α-particles were 2.4-fold greater than those induced by X-rays. Rejoining the ssb induced by either radiation was rapid and >90% were repaired within 60 min post-irradiation incubation at 370. The time necessary for 50% repair (t/sub 1/2/) of X-ray-induced dsb was --40 min, however, for ---irradiated cells (20-110 Gy) the t/sub 1/2/ was 4-8-fold greater. Even after prolonged post-irradiation incubation (48 hr) using confluent GM10, --30% dsb remained unrepaired. This class of unrepaired DNA lesions probably contributes to the elevated RBE for high-LET radiation

  9. Elongation of 20-carbon polyunsaturated fatty acids by human skin fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Banerjee, N.; Rosenthal, M.D.

    1986-05-01

    Human skin fibroblasts readily incorporate exogenous arachidonate (20:4(n-6)) and eicosapentaenoate (20:5(n-3)) into cellular phospholipids and triacylglycerol. The extent of incorporation of 1.25 ..mu..M (/sup 14/C)20:4(n-6) and (/sup 14/C)20:5(n-3) from culture medium with delipidized serum protein is similar, 20% in 1 hr increasing to 60-70% within 8 hr. Elongation of incorporated (/sup 14/C)20:5(n-3) to (/sup 14/C)22:5(n-3) is extensive, 40% by 8 hr and 85% by 48 hr. Elongation of (/sup 14/C)20:4(n-6) to (/sup 14/C)22:4(n-6) is <1/2 that of (/sup 1/$C)20:5(n-3) and plateaus at approx.20% of incorporated /sup 14/C-fatty acid. Although incorporated, exogenous 22:4(n-6) is not an effective inhibitor of the elongation of (/sup 14/C)arachidonate; however, exogenous 20:5(n-3) is inhibitory, with an ID/sub 50/ of 5 ..mu..M. With exogenous concentrations of (/sup 14/C)arachidonate from 0.4 - 10 ..mu..M, the percentage incorporated in 24 hr remains relatively constant. By contrast, the extent of elongation of incorporated (/sup 14/C)arachidonate increases from 12% at 0.4 ..mu..M to 43% at 10 ..mu..M. Under these conditions, elongation of incorporated (/sup 14/C)20:5(n-3) is approx.75%. Thus, in these cells, selectivity of the elongation system results in differential metabolism of 20-carbon n-6 and n-3 fatty acids. Furthermore, arachidonate appears to act as a positive modulator of its own elongation.

  10. Cytotoxic and mutagenic effects of high let charged particles on human skin fibroblasts

    International Nuclear Information System (INIS)

    Cytotoxic and mutagenic effects of high LET charged particles were quantitatively measured using primary cultures of human skin fibroblasts. The span of LETs selected were from 25 keV/μm(330 MeV/u) to 920 keV/μm (600 MeV/u). Mutations were scored at the hypoxanthine-guanine phosphoribosyl transferase (HPRT) locus using 6-thioguanine (6-TG) for selection. Exposure to these high LET charged particles resulted in exponential survival curves whereas mutant induction was fitted by a linear model. The Relative Biological Effect (RBE) for cell-killing ranged from 3.73 to 1.25, while that for mutant induction ranged from 5.74 to 0.48. Maximum RBE values were obtained at the LET of 150 keV/μm. The inactivation cross-section (σi) and the action cross-section for mutant induction (σm) ranged from 2.2 to 92.0 μm2 and 0.09 to 5.56 x 10-3 μm2 respectively, the maximum values were obtained by 56Fe with an LET of 200 keV/μm. The mutagenicity (σm/σi) ranged from 2.05 to 7.99 x 10-5 with the maximum value at 150 keV/μm. Furthermore, the results of multiplex polymerase chain reaction (PCR) of some of the mutants induced by charged particles indicate that higher LET beams are more likely to cause larger deletions in the hprt locus. (author)

  11. Establishment and biological research of the Jining Grey goat fibroblast line

    OpenAIRE

    BAI, Chunyu; WANG, Dianjun; SU, Xiaohua; Zhang, Minghai; Guan, Weijun; MA, YUEHUI

    2012-01-01

    Using Jining Grey goat ear marginal tissues as experimental materials, we succeeded in establishing a fibroblast cell bank, including 32 samples, by means of a primary explant and cryopreservation technique. The results from the biological analysis suggested that the population doubling time of the cell line was approximately 48 h. The diploid accounted for 98.35%, the isozyme analysis of the lactic dehydrogenase and malic dehydrogenase disproved cross contamination, and the results of the ba...

  12. Endogenous glutathione protects human skin fibroblasts against the cytotoxic action of UVB, UVA and near-visible radiations

    International Nuclear Information System (INIS)

    Both the UVB (290-320 nm) and UVA (320-380 nm) regions of sunlight damage human skin cells but, particularly at the longer wavelengths, information is scant concerning the mechanism(s) of damage induction and the roles of cellular defense mechanisms. Following extensive glutathione depletion of cultured human skin fibroblasts, the cells become strongly sensitized to the cytotoxic action of near-visible (405 nm), UVA (334 nm, 365 nm) and UVB (313 nm) but not UVC (254 nm) radiations. In the critical UVB region, the magnitude of the protection afforded by endogenous glutathione approaches that of the protection provided by excision repair. The results suggest that a significant fraction of even UVB damage can be mediated by free radical attack and that a major role of glutathione in human skin cells is to protect them from the cytotoxic action of sunlight. (author)

  13. Effect of low intensity laser interaction with human skin fibroblast cells using fiber-optic nano-probes.

    Science.gov (United States)

    Pal, Gopalendu; Dutta, Ashim; Mitra, Kunal; Grace, Michael S; Amat, Albert; Romanczyk, Tara B; Wu, Xingjia; Chakrabarti, Kristi; Anders, Juanita; Gorman, Erik; Waynant, Ronald W; Tata, Darrell B

    2007-03-01

    Over the past forty years, many efforts have been devoted to study low power laser light interactions with biological systems. Some of the investigations were performed in-vitro, on bulk cell populations. Our present work was undertaken to apply specially engineered fiber-optic based nano-probes for the precise delivery of laser light on to a single cell and to observe production of low power laser light induced reactive oxygen species (ROS). A normal human skin fibroblast (NHF) cell line was utilized in this investigation and the cells were irradiated under two different schemes of exposure: (1) an entire NHF cell population within a Petri dish using a fan beam methodology, and (2) through the precise delivery of laser energy on to a single NHF cell using fiber-optic nano-probe. Photobiostimulative studies were conducted through variation of laser intensity, exposure time, and the energy dose of exposure. Laser irradiation induced enhancement in the rate of cell proliferation was observed to be dependent on laser exposure parameters and the method of laser delivery. The total energy dose (fluence) had a greater influence on the enhancement in the rate of cellular proliferation than compared to laser intensity. The enhancement in the growth rate was observed to have a finite life-time of several days after the initial laser exposure. Fluorescent life-time imaging of ROS was performed during the nano-based single cell exposure method. The kinetics of ROS generation was found to depend strongly on the laser fluence and not on the laser intensity. PMID:17224276

  14. Methylmalonic and propionic acidemias: lipid profiles of normal and affected human skin fibroblasts incubated with [1-14C]propionate

    International Nuclear Information System (INIS)

    Normal human skin fibroblasts and those from methylmalonic acidemia and propionic acidemia patients were grown in culture. Following incubation with [1-14C]propionate, the major lipid classes in the cells were separated by thin layer chromatography and isolated fractions analyzed by radio gas chromatography for the presence of odd-numbered long-chain fatty acids; the pattern of even-numbered long-chain fatty acids was obtained also. Normal fibroblasts incorporated a small percentage of propionate into odd-numbered fatty acids which were present in all lipids studied. The abnormal cells incorporated a larger amount while maintaining the characteristic ratios of odd-numbered fatty acids found in the normal line. Most of the radioactivity was associated with phospholipids which are the predominant constituents of cell membranes. A characteristic C15/C17 ratio was found for different phospholipids and the triglyceride fraction; pentadecanoic acid was the principal odd-numbered fatty acid utilized in the assembly of complex lipids. Compared to even-numbered long-chain fatty acids the absolute amount of odd-numbered fatty acids was low (1-2%), even in affected cells. An unusual polar lipid fraction was isolated in the course of the study. In the normal cell it contained several unlabeled eicosanoids which were missing from the same fraction of both affected cell lines

  15. Impaired colony-forming ability following ? irradiation of skin fibroblasts from tuberous scierosis patients

    International Nuclear Information System (INIS)

    The radiosensitivity of cultured dermal fibroblasts from human subjects afflicted with tuberous sclerosis (TS), a hereditary neurocutaneous syndrome, was assessed by assaying loss of colony-forming ability in response to acute ?-ray exposure. Related to control strains from clinically normal donors, three cell lines (GM1635, GM1643, GM2333) from two affected patients displayed enhanced sensitivity to inactivation by 60Co ?-ray treatment, whether administered oxically (air-saturated) or hypoxically (N2-gassed); a fourth strain (GM1644) from a third patient exhibited normal radiosensitivity under both treatment conditions. The post-?-irradiaton colony-forming ability of the three hypersensitive TS strains was intermediate between that of normal controls and that of strains from patients inheriting the radiotherapy-sensitive neurovascular disorder ataxia telangiectasia. The variability in the radioresponse of the TS stains (three sensitive and one normal) is not surprising, considering the widely recognized clinical heterogeneity in the disease. Our findings, aside from providing a laboratory marker for early (possible presymptomatic) detection of persons at high risk for TS, may lead to a better understanding of the origin and progressive development of this multifaceted syndrome

  16. The Apoptotic Effects of the P300 Activator on Breast Cancer and Lung Fibroblast Cell Lines

    OpenAIRE

    Mohammad Reza Salahshoor; Cyrus Jalili; Ahmad Shebanizadeh Darehdori; Mozafar Khazaei; Shiva Roshankhah; Rostam Ghorbani

    2013-01-01

    Background: P300 is an enzyme that acetylates histones during stress. It alsoacetylates several non-histone proteins, including P53 which is the most important tumorsuppressor gene. P53 plays an important role in the apoptosis of tumor cells. Hereby,this study describes the potency of cholera toxin B subunit as a P300 activator to induceapoptosis in a breast cancer cell line (MCF-7) and a lung fibroblast cell line (MRC-5)as a non-tumorigenic control sample. Methods: MCF-7 and MRC-5 were cultu...

  17. Acute leukemia after radiotherapy in a patient with Turcot's syndrome. Impaired colony formation in skin fibroblast cultures after irradiation

    International Nuclear Information System (INIS)

    Colonic polyposis and carcinoma developed in a woman with Turcot's syndrome at the age of 31 years; astrocytoma developed when she was 37. Her brother and sister had died of astrocytoma at the ages of 18 and 33 years, respectively. Progressive neutropenia developed in the patient three months after radiotherapy for her brain tumor and acute myelomonocytic leukemia 19 months after treatment. Three laboratories independently evaluated cultures of her skin fibroblasts for in vitro sensitivity to cell killing (loss of colony-forming ability) by x-rays. Survival assays consistently revealed slight but significant radiosensitivity in an early-passage (six to 10 doublings) fibroblast subculture. A later subculture (21 to 29 doublings) showed no abnormality, a possible effect of selective in vitro loss of radiosensitive cells

  18. Effect of wavelength and fluence on morphology, cellular and genetic integrity of diabetic wounded human skin fibroblasts

    Science.gov (United States)

    Abrahamse, H.; Hawkins, D.; Houreld, N.

    2006-02-01

    An alternative treatment modality for diabetic wound healing includes low level laser therapy (LLLT). Biostimulation of such wounds may be of benefit to patients by reducing healing time. Structural, cellular and genetic events in diabetic wounded human skin fibroblasts (WS1) were evaluated after exposing cells in culture to a Helium-Neon (632.8nm), a Diode laser (830nm) and a Nd:YAG (Neodynium:Yttrium-Allumina-Gallium) laser (1064nm) at either 5J/cm2 or 16J/cm2. Cells were exposed twice a week and left 24 hours post-irradiation prior to measuring effects. Structural changes were evaluated by assessing colony formation, haptotaxis and chemotaxis. Cellular changes were evaluated using cell viability, (adenosine-triphosphate, ATP production), and proliferation, (alkaline phosphatase, ALP and basic fibroblast growth factor, bFGF expression), while the Comet assay evaluated DNA damage and cytotoxicity was determined assessing membrane permeability for lactate dehydrogenase (LDH). Caspase 3/7 activity was used as an estimate of apoptosis as a result of irradiation. The irradiated diabetic wounded cells showed structural, cellular as well as molecular resilience comparable to that of unwounded normal skin fibroblast cells. With regards to fluence, 5J/cm2 elicit positive cellular and structural responses while 16J/cm2 increases cellular and genetic damage and cellular morphology is altered. Different wavelengths of LLLT influences the beneficial outcomes of diabetic wounded cells and although all three wavelengths elicit cellular effects, the penetration depth of 830nm plays a significant role in the healing of diabetic wounded human fibroblast cells. Results from this study validate the contribution of LLLT to wound healing and elucidate the biochemical effects at a cellular level while highlighting the role of different dosages and wavelengths in LLLT.

  19. Fabrication of a nanofibrous scaffold with improved bioactivity for culture of human dermal fibroblasts for skin regeneration

    International Nuclear Information System (INIS)

    Engineering dermal substitutes with electrospun nanofibres have lately been of prime importance for skin tissue regeneration. Simple electrospinning technology served to produce nanofibrous scaffolds morphologically and structurally similar to the extracellular matrix of native tissues. The nanofibrous scaffolds of poly(l-lactic acid)-co-poly(ε-caprolactone) (PLACL) and PLACL/gelatin complexes were fabricated by the electrospinning process. These nanofibres were characterized for fibre morphology, membrane porosity, wettability and chemical properties by FTIR analysis to culture human foreskin fibroblasts for skin tissue engineering. The nanofibre diameter was obtained between 282 and 761 nm for PLACL and PLACL/gelatin scaffolds; expressions of amino and carboxyl groups and porosity up to 87% were obtained for these fibres, while they also exhibited improved hydrophilic properties after plasma treatment. The results showed that fibroblasts proliferation, morphology, CMFDA dye expression and secretion of collagen were significantly increased in plasma-treated PLACL/gelatin scaffolds compared to PLACL nanofibrous scaffolds. The obtained results prove that the plasma-treated PLACL/gelatin nanofibrous scaffold is a potential biocomposite material for skin tissue regeneration.

  20. Fabrication of a nanofibrous scaffold with improved bioactivity for culture of human dermal fibroblasts for skin regeneration

    Energy Technology Data Exchange (ETDEWEB)

    Chandrasekaran, Arun Richard; Venugopal, J; Sundarrajan, S; Ramakrishna, S, E-mail: nnijrv@nus.edu.s [Healthcare and Energy Materials Laboratory, Nanoscience and Nanotechnology Initiative, Faculty of Engineering, National University of Singapore (Singapore)

    2011-02-15

    Engineering dermal substitutes with electrospun nanofibres have lately been of prime importance for skin tissue regeneration. Simple electrospinning technology served to produce nanofibrous scaffolds morphologically and structurally similar to the extracellular matrix of native tissues. The nanofibrous scaffolds of poly(l-lactic acid)-co-poly({epsilon}-caprolactone) (PLACL) and PLACL/gelatin complexes were fabricated by the electrospinning process. These nanofibres were characterized for fibre morphology, membrane porosity, wettability and chemical properties by FTIR analysis to culture human foreskin fibroblasts for skin tissue engineering. The nanofibre diameter was obtained between 282 and 761 nm for PLACL and PLACL/gelatin scaffolds; expressions of amino and carboxyl groups and porosity up to 87% were obtained for these fibres, while they also exhibited improved hydrophilic properties after plasma treatment. The results showed that fibroblasts proliferation, morphology, CMFDA dye expression and secretion of collagen were significantly increased in plasma-treated PLACL/gelatin scaffolds compared to PLACL nanofibrous scaffolds. The obtained results prove that the plasma-treated PLACL/gelatin nanofibrous scaffold is a potential biocomposite material for skin tissue regeneration.

  1. Effect of bradykinin on prostaglandin production by human skin fibroblasts in culture

    International Nuclear Information System (INIS)

    In studying the effect of bradykinin on prostaglandin formation in fibroblasts, two general types of assays have been employed. Radioimmunoassays were utilized to determine specific prostaglandins, PGI2 and 6-keto-PGF, using tritium as radiolabel. A second procedure involved initial incubation of the fibroblasts with [14C]arachidonate and identification and quantification of the metabolites by chromatographic methods. After radiolabelling of fibroblasts with [14C]arachidonate, bradykinin was found to release radiolabel from membrane lipids approximately 2-fold. In the presence of bradykinin, there was a close coupling of phospholipase S2 activity, arachidonate mobilization, and synthesis of a specific prostaglandin, PGI2

  2. Contact guidance of chick embryo neurons on single scratches in glass and on underlying aligned human skin fibroblasts.

    Science.gov (United States)

    S?epie?, E; Stanisz, J; Korohoda, W

    1999-01-01

    The influence of substratum topography on the morphology and orientation of neurites of chick embryo neurons was studied. Two series of experiments are reported. One concerned the behaviour of growth cones when the axons become contact-guided by the surface texture. The second studied contact guidance of neurites extending on a compact layer of fixed aligned human skin fibroblasts (HSF). It was observed that when the growth cones of sensory neurons isolated from dorsal root ganglions encountered a single scratch in a glass surface (0.1-2 microm in depth and diameter) they turned and continued movement following the axis of the scratch. These neurons became contact-guided as a result of the sequence of events. The growth cone filopodia recognized the irregularity in the substratum surface, whereas the growth cone lamella stabilized contact with the scratch and moved forward along the scratch axis. Scanning electron microscope revealed that the single scratches 150 nm in width and ca. 100 nm deep growth cone filopodia less than 200 nm in diameter could detect and react by turning into them. These filopodia extensions followed the edge of scratches. However, phase contrast and Nomarski's differential interference contrast appeared insufficient for analysis of primary contact guidance of fine growth cone filopodia which themselves are often less than 200 nm. In neuron cultures on fixed aligned HSF, the neuron aggregates assumed spindle-like shapes, and sparsely seeded individual neurons extended axons along the long axes of the fibroblasts. The axons extended significantly further on the fixed underlying fibroblasts than on collagen-covered glass. In crowded cultures of neurons, the cells extended neurites ignoring both the surface anisotropy (the scratches) and the orientation of the aligned fibroblasts. Immunofluorescence staining of neurons with antibodies against neurofilaments made it possible to analyse their shape and orientation on the fibroblasts. Computer-assisted image analysis permitted the observed alignment of the neurites to be characterized quantitatively. PMID:10561119

  3. DNA damage in wounded, hypoxic and acidotic human skin fibroblast cell cultures after low laser irradiation

    Science.gov (United States)

    Hawkins Evans, D.; Mbene, A.; Zungu, I.; Houreld, N.; Abrahamse, H.

    2009-02-01

    Phototherapy has become more popular and widely used in the treatment of a variety of medical conditions. To ensure sound results as evidence of its effectiveness, well designed experiments must be conducted when determining the effect of phototherapy. Cell culture models such as hypoxic, acidotic and wounded cell cultures simulating different disease conditions including ischemic heart disease, diabetes and wound healing were used to determine the effect of laser irradiation on the genetic integrity of the cell. Even though phototherapy has been found to be beneficial in a wide spectrum of conditions, it has been shown to induce DNA damage. However, this damage appears to be repairable. The risk lies in the fact that phototherapy may help the medical condition initially but damage DNA at the same time leaving undetected damage that may result in late onset, more severe, induced medical conditions including cancer. Human skin fibroblasts were cultured and used to induce a wound (by the central scratch model), hypoxic (by incubation in an anaerobic jar, 95% N2 and 5% O2) and acidotic (reducing the pH of the media to 6.7) conditions. Different models were irradiated using a Helium-Neon (632.8 nm) laser with a power density of 2.07 mW/cm2 and a fluence of 5 J/cm2 or 16 J/cm2. The effect of the irradiation was determined using the Comet assay 1 and 24 h after irradiation. In addition, the Comet assay was performed with the addition of formamidopyrimidine glycosylase (FPG) obviating strand brakes in oxidized bases at a high fluence of 16 J/cm2. A significant increase in DNA damage was seen in all three injured models at both 1 and 24 h post-irradiation when compared to the normal un-injured cells. However, when compared to non-irradiated controls the acidotic model showed a significant decrease in DNA damage 24 h after irradiation indicating the possible induction of cellular DNA repair mechanisms. When wounded cells were irradiated with higher fluences of 16 J/cm2, there was a significant increase in DNA damage in irradiated cells with and without the addition of FPG. These results are indicative of the importance of both cell injury model as well as fluence when assessing the effect of phototherapy on DNA integrity.

  4. Curcumin induces heme oxygenase-1 in normal human skin fibroblasts through redox signaling : relevance for anti-aging intervention

    OpenAIRE

    Lima, Cristo?va?o F.; Wilson, Cristina Pereira; Rattan, Suresh

    2011-01-01

    Curcumin, a component of the spice turmeric, was tested for its potential hormetic anti-aging effects as an inducer of mild stress. Methods and results: Early passage young human skin fibroblasts treated with low doses of curcumin (below 20 mM) showed a time- and concentration-dependent induction of heme oxygenase-1 (HO-1), followed by compensatory increase in glutathione-S-transferase activity, GSH levels and GSH/GSSG ratio. These effects were preceded by induction of oxidative stress (in...

  5. Mechanisms of sulfur mustard analog 2-chloroethyl ethyl sulfide-induced DNA damage in skin epidermal cells and fibroblasts

    OpenAIRE

    Inturi, Swetha; Tewari-Singh, Neera; Gu, Mallikarjuna; Shrotriya, Sangeeta; Gomez, Joe; Agarwal, Chapla; White, Carl W.; Agarwal, Rajesh

    2011-01-01

    Employing mouse skin epidermal JB6 cells and dermal fibroblasts, here we examined the mechanisms of DNA damage by 2-chloroethyl ethyl sulfide (CEES), a monofunctional analog of sulfur mustard (SM). CEES exposure caused H2A.X and p53 phosphorylation as well as p53 accumulation in both cell types, starting at 1 h, that was sustained for 24 h, indicating a DNA-damaging effect of CEES, which was also confirmed and quantified by alkaline comet assay. CEES exposure also induced oxidative stress and...

  6. Effects of sulfate deprivation on the production of chondroitin/dermatan sulfate by cultures of skin fibroblasts from normal and diabetic individuals

    International Nuclear Information System (INIS)

    Human skin fibroblast monolayer cultures from two normal men, three Type I diabetic men, and one Type I diabetic woman were incubated with [3H]glucosamine in the presence of diminished concentrations of sulfate. Although total synthesis of [3H]chondroitin/dermatan glycosaminoglycans varied somewhat between cell lines, glycosaminoglycan production was not affected within any line when sulfate levels were decreased from 0.3 mM to 0.06 mM to 0.01 mM to 0 added sulfate. Lowering of sulfate concentrations resulted in diminished sulfation of chondroitin/dermatan in a progressive manner, so that overall sulfation dropped to as low as 19% for one of the lines. Sulfation of chondroitin to form chondroitin 4-sulfate and chondroitin 6-sulfate was progressively and equally affected by decreasing the sulfate concentration in the culture medium. However, sulfation to form dermatan sulfate was preserved to a greater degree, so that the relative proportion of dermatan sulfate to chondroitin sulfate increased. Essentially all the nonsulfated residues were susceptible to chondroitin AC lyase, indicating that little epimerization of glucuronic acid residues to iduronic acid had occurred in the absence of sulfation. These results confirm the previously described dependency of glucuronic/iduronic epimerization on sulfation, and indicate that sulfation of the iduronic acid-containing disaccharide residues of dermatan can take place with sulfate concentrations lower than those needed for 6-sulfation and 4-sulfation of the glucuronic acid-containing disaccharide residues of chondroitin. There were considerable differences among the six fibroblast lines in susceptibility to low sulfate medium and in the proportion of chondroitin 6-sulfate, chondroitin 4-sulfate, and dermatan sulfate. However, there was no pattern of differences between normals and diabetics

  7. Effects of sulfate deprivation on the production of chondroitin/dermatan sulfate by cultures of skin fibroblasts from normal and diabetic individuals

    Energy Technology Data Exchange (ETDEWEB)

    Silbert, C.K.; Humphries, D.E.; Palmer, M.E.; Silbert, J.E. (Veterans Administration Outpatient Clinic, Boston, MA (USA))

    1991-02-15

    Human skin fibroblast monolayer cultures from two normal men, three Type I diabetic men, and one Type I diabetic woman were incubated with (3H)glucosamine in the presence of diminished concentrations of sulfate. Although total synthesis of (3H)chondroitin/dermatan glycosaminoglycans varied somewhat between cell lines, glycosaminoglycan production was not affected within any line when sulfate levels were decreased from 0.3 mM to 0.06 mM to 0.01 mM to 0 added sulfate. Lowering of sulfate concentrations resulted in diminished sulfation of chondroitin/dermatan in a progressive manner, so that overall sulfation dropped to as low as 19% for one of the lines. Sulfation of chondroitin to form chondroitin 4-sulfate and chondroitin 6-sulfate was progressively and equally affected by decreasing the sulfate concentration in the culture medium. However, sulfation to form dermatan sulfate was preserved to a greater degree, so that the relative proportion of dermatan sulfate to chondroitin sulfate increased. Essentially all the nonsulfated residues were susceptible to chondroitin AC lyase, indicating that little epimerization of glucuronic acid residues to iduronic acid had occurred in the absence of sulfation. These results confirm the previously described dependency of glucuronic/iduronic epimerization on sulfation, and indicate that sulfation of the iduronic acid-containing disaccharide residues of dermatan can take place with sulfate concentrations lower than those needed for 6-sulfation and 4-sulfation of the glucuronic acid-containing disaccharide residues of chondroitin. There were considerable differences among the six fibroblast lines in susceptibility to low sulfate medium and in the proportion of chondroitin 6-sulfate, chondroitin 4-sulfate, and dermatan sulfate. However, there was no pattern of differences between normals and diabetics.

  8. Inhibition of Oxidative Stress by Low-Molecular-Weight Polysaccharides with Various Functional Groups in Skin Fibroblasts

    Directory of Open Access Journals (Sweden)

    Gregor P. C. Drummen

    2013-09-01

    Full Text Available The aim of this study was to evaluate the in cellulo inhibition of hydrogen-peroxide-induced oxidative stress in skin fibroblasts using different low-molecular-weight polysaccharides (LMPS prepared from agar (LMAG, chitosan (LMCH and starch (LMST, which contain various different functional groups (i.e., sulfate, amine, and hydroxyl groups. The following parameters were evaluated: cell viability, intracellular oxidant production, lipid peroxidation, and DNA damage. Trolox was used as a positive control in order to allow comparison of the antioxidant efficacies of the various LMPS. The experimentally determined attenuation of oxidative stress by LMPS in skin fibroblasts was: LMCH > LMAG > LMST. The different protection levels of these LMPS may be due to the physic-chemical properties of the LMPS functional groups, including electron transfer ability, metal ion chelating capacities, radical stabilizing capacity, and the hydrophobicity of the constituent sugars. The results suggest that LMCH might constitute a novel and potential dermal therapeutic and sun-protective agent.

  9. Chromosomal radiosensitivity during the G2 cell-cycle period of skin fibroblasts from individuals with familial cancer

    International Nuclear Information System (INIS)

    The authors reported previously that human cells after neoplastic transformation in culture had acquired an increased susceptibility to chromatid damage induced by x-irradiation during the G2 phase of the cell cycle. Evidence suggested that this results from deficient DNA repair during G2 phase. Cells derived from human tumors also showed enhanced G2-phase chromosomal radiosensitivity. Furthermore, skin fibroblasts from individuals with genetic diseases predisposing to a high risk of cancer, including ataxia-telangiectasia, Bloom syndrome, Fanconi anemia, and xeroderma pigmentosum exhibited enhanced G2-phase chromosomal radiosensitivity. The present study shows that apparently normal skin fibroblasts from individuals with familial cancer--i.e., from families with a history of neoplastic disease--also exhibit enhanced G2-phase chromosomal radiosensitivity. This radiosensitivity appears, therefore, to be associated with both a genetic predisposition to cancer and a malignant neoplastic state. Furthermore, enhanced G2-phase chromosomal radiosensitivity may provide the basis for an assay to detect genetic susceptibility to cancer

  10. Analysis of carnitine esters by radio-high performance liquid chromatography in cultured skin fibroblasts from patients with mitochondrial fatty acid oxidation disorders.

    Science.gov (United States)

    Schmidt-Sommerfeld, E; Bobrowski, P J; Penn, D; Rhead, W J; Wanders, R J; Bennett, M J

    1998-08-01

    Acylcarnitines are important diagnostic markers for inborn errors of fatty acid oxidation, but their analysis in body fluids may not always be reliable. Recently, disease-specific acylcarnitine profiles generated by cultured skin fibroblasts were reported to facilitate the diagnosis by localizing a specific enzymatic defect in the mitochondrial beta-oxidation pathway. Using a novel methodologic approach, fibroblasts from 16 patients with inborn errors of fatty acid oxidation and 13 control subjects were preincubated with L-[3H]carnitine to label the intracellular carnitine pool. Cells were subsequently incubated with unlabeled palmitic acid and, after methanol extraction of cells and media, labeled free carnitine and acylcarnitines were analyzed by radio-HPLC. Quantitation was based on the integrated radioactivity of individual peaks relative to the total radioactivity recovered. In control cell lines, all saturated acylcarnitines were detected, and reference values were established. With the exception of one cell line deficient in electron transfer flavoprotein, all mutant cell lines showed abnormal and disease-specific relative concentrations of acylcarnitines. Advantages of the method include use of a small number of cells, no need for trypsinization and permeabilization of cells before incubation, simple extraction without purification of the specimen before HPLC, and relatively inexpensive equipment. The method allows a focused approach to the subsequent, more laborious confirmation of a particular disease by direct enzymatic and/or molecular analysis. It remains to be established whether the method can replace widely used global measurements of fatty acid oxidation rates in vitro that do not provide specific information about the enzyme deficiency involved. PMID:9702916

  11. Mutations of ATIC and ADSL affect purinosome assembly in cultured skin fibroblasts from patients with AICA-ribosiduria and ADSL deficiency.

    Science.gov (United States)

    Baresova, Veronika; Skopova, Vaclava; Sikora, Jakub; Patterson, David; Sovova, Jana; Zikanova, Marie; Kmoch, Stanislav

    2012-04-01

    The purinosome is a multienzyme complex composed by the enzymes active in de novo purine synthesis (DNPS) that cells transiently assemble in their cytosol upon depletion or increased demand of purines. The process of purinosome formation has thus far been demonstrated and studied only in human epithelial cervical cancer cells (HeLa) and human liver carcinoma cells (C3A) transiently expressing recombinant fluorescently labeled DNPS proteins. Using parallel immunolabeling of various DNPS enzymes and confocal fluorescent microscopy, we proved purinosome assembly in HeLa, human hepatocellular liver carcinoma cell line (HepG2), sarcoma osteogenic cells (Saos-2), human embryonic kidney cells (HEK293), human skin fibroblasts (SF) and primary human keratinocytes (KC) cultured in purine-depleted media. Using the identical approach, we proved in cultured skin fibroblasts from patients with AICA-ribosiduria and ADSL deficiency that various mutations of ATIC and ADSL destabilize to various degrees of purinosome assembly and found that the ability to form purinosomes correlates with clinical phenotypes of individual ADSL patients. Our results thus shown that the assembly of functional purinosomes is fully dependent on the presence of structurally unaffected ATIC and ADSL complexes and presumably also on the presence of all the other DNPS proteins. The results also corroborate the hypothesis that the phenotypic severity of ADSL deficiency is mainly determined by structural stability and residual catalytic capacity of the corresponding mutant ADSL protein complexes, as this is prerequisite for the formation and stability of the purinosome and at least partial channeling of succinylaminoimidazolecarboxamide riboside-ADSL enzyme substrates-through the DNPS pathway. PMID:22180458

  12. Effects of hyperthermia and ionizing radiation in normal and ataxia telangiectasia human fibroblast lines

    International Nuclear Information System (INIS)

    The effects of 450C hyperthermia and ? radiation have been studied in three normal human fibroblast lines (GM38, GM730, WI38) and compared to the effects in two lines derived from patients with the hereditary disease ataxia telangiectasia (AR3BI, AT5BI). All lines, both normal and ?-sensitive AT, showed a similar resistance to killing by heat alone, suggesting that the defect responsible for the increased radiation sensitivity in AT lines does not confer increased heat sensitivity. Shouldered survival curves were obtained in each case indicating the ability to accumulate sublethal heat damage. All normal and AT cell lines exhibited increased resistance to the lethal effects of heat in response to a thermal stress, indicating that the defect that causes radiosensitivity in AT cell lines does not prevent the induction of thermotolerance. It was hypothesized that in normal cells, this heat treatment inactivates the process which is already defective in AT lines, and that this process may be required for the proper rejoining of double-strand breaks produced during the repair of other radiation-induced lesions

  13. Dandelion Extracts Protect Human Skin Fibroblasts from UVB Damage and Cellular Senescence

    OpenAIRE

    Yafan Yang; Shuangshuang Li

    2015-01-01

    Ultraviolet (UV) irradiation causes damage in skin by generating excessive reactive oxygen species (ROS) and induction of matrix metalloproteinases (MMPs), leading to skin photoageing. Dandelion extracts have long been used for traditional Chinese medicine and native American medicine to treat cancers, hepatitis, and digestive diseases; however, less is known on the effects of dandelion extracts in skin photoageing. Here we found that dandelion leaf and flower extracts significantly protect U...

  14. Heme oxygenase is the major 32-kDa stress protein induced in human skin fibroblasts by UVA radiation, hydrogen peroxide, and sodium arsenite.

    OpenAIRE

    Keyse, S.M.; Tyrrell, R M

    1989-01-01

    We have shown that UVA (320-380 nm) radiation, hydrogen peroxide, and sodium arsenite induce a stress protein of approximately 32 kDa in human skin fibroblasts. The synthesis and cloning of cDNA from arsenite-induced mRNA populations have now allowed us to unequivocally identify the 32-kDa protein as heme oxygenase. By mRNA analysis we have shown that the heme oxygenase gene is also induced in cultured human skin fibroblasts by UVA radiation, hydrogen peroxide, cadmium chloride, iodoacetamide...

  15. L-Lactate Protects Skin Fibroblasts against Aging-Associated Mitochondrial Dysfunction via Mitohormesis

    Czech Academy of Sciences Publication Activity Database

    Zelenka, Jaroslav; Dvořák, Aleš; Alán, Lukáš

    2015-01-01

    Roč. 2015, č. 2015 (2015), ID351698. ISSN 1942-0900 R&D Projects: GA ČR(CZ) GPP305/12/P388 Institutional support: RVO:67985823 Keywords : mitochondria * reactive oxygen species * lactate * fibroblasts Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.516, year: 2014

  16. Reduction of fibroblast size/mechanical force down-regulates TGF-β type II receptor: implications for human skin aging.

    Science.gov (United States)

    Fisher, Gary J; Shao, Yuan; He, Tianyuan; Qin, Zhaoping; Perry, Daniel; Voorhees, John J; Quan, Taihao

    2016-02-01

    The structural integrity of human skin is largely dependent on the quality of the dermal extracellular matrix (ECM), which is produced, organized, and maintained by dermal fibroblasts. Normally, fibroblasts attach to the ECM and thereby achieve stretched, elongated morphology. A prominent characteristic of dermal fibroblasts in aged skin is reduced size, with decreased elongation and a more rounded, collapsed morphology. Here, we show that reduced size of fibroblasts in mechanically unrestrained three-dimensional collagen lattices coincides with reduced mechanical force, measured by atomic force microscopy. Reduced size/mechanical force specifically down-regulates TGF-β type II receptor (TβRII) and thus impairs TGF-β/Smad signaling pathway. Both TβRII mRNA and protein were decreased, resulting in 90% loss of TGF-β binding to fibroblasts. Down-regulation of TβRII was associated with significantly decreased phosphorylation, DNA-binding, and transcriptional activity of its key downstream effector Smad3 and reduced expression of Smad3-regulated essential ECM components type I collagen, fibronectin, and connective tissue growth factor (CTGF/CCN2). Restoration of TβRII significantly increased TGF-β induction of Smad3 phosphorylation and stimulated expression of ECM components. Reduced expression of TβRII and ECM components in response to reduced fibroblast size/mechanical force was fully reversed by restoring size/mechanical force. Reduced fibroblast size was associated with reduced expression of TβRII and diminished ECM production, in aged human skin. Taken together, these data reveal a novel mechanism that provides a molecular basis for loss of dermal ECM, with concomitant increased fragility, which is a prominent feature of human skin aging. PMID:26780887

  17. DNA double strand breaks in fibroblast cell lines from non-Hodgkin's lymphoma patients showing increased sensitivity to chronic gamma irradiation

    International Nuclear Information System (INIS)

    Cultured skin fibroblast cell lines from two non-Hodgkin's lymphoma patients (NHL) and a normal subject were studied for cell killing, chromosomal aberrations (breaks, translocations, dicentrics and rings) and DNA double strand breaks (dsbs) following chronic gamma irradiation. Compared to the cell line from the normal donor, the NHL patients' fibroblasts showed enhanced radiosensitivity for both cell survival and chromosomal aberrations. While spontaneous breaks were observed in both normal and patients' cells, spontaneous translocations and radiation-induced dicentrics and rings were found only in the latter. Radiation-induced DNA double-strand breaks (dsb) were determined by CHEF electrophoresis. After chronic irradiation with gamma rays the fraction of residual dsb was significantly increased from 1.4% in controls to 1.9% in the NHL cell lines. These data, thus suggest that the cellular and chromosomal sensitivity to chronic irradiation observed in NHL patients may be due to a deficiency in the repair of a small fraction of DNA double strand breaks. (author)

  18. Efficient retrovirus-mediated transfer and expression of a human adenosine deaminase gene in diploid skin fibroblasts from an adenosine deaminase-deficient human

    Energy Technology Data Exchange (ETDEWEB)

    Palmer, T.D.; Hock, R.A.; Osborne, W.R.A.; Miller, A.D.

    1987-02-01

    Skin fibroblasts might be considered suitable recipients for therapeutic genes to cure several human genetic diseases; however, these cells are resistant to gene transfer by most methods. The authors studied the ability of retroviral vectors to transfer genes into normal human diploid skin fibroblasts. Retroviruses carrying genes for neomycin or hygromycin B resistance conferred drug resistance to greater than 50% of the human fibroblasts after a single exposure to virus-containing medium. This represents at least a 500-fold increase in efficiency over other methods. Transfer was achieved in the absence of helper virus by using amphotropic retrovirus-packaging cells. A retrovirus vector containing a human adenosine deaminase (ADA) cDNA was constructed and used to infect ADA/sup -/ fibroblasts from a patient with ADA deficiency. The infected cells produced 12-fold more ADA enzyme than fibroblasts from normal individuals and were able to rapidly metabolize exogenous deoxyadenosine and adenosine, metabolites that accumulate in plasma in ADA-deficient patients and are responsible for the severe combined immunodeficiency in these patients. These experiments indicate the potential of retrovirus-mediated gene transfer into human fibroblasts for gene therapy.

  19. Abnormal sensitivity of skin fibroblasts from familial polyposis patients to DNA alkylating agents

    International Nuclear Information System (INIS)

    Fibroblast cell strains derived from different patients all afflicted with genetic predisposing to the development of intestinal polyposis and cancer were tested for their sensitivity to the lethal effects of the DNA alkylating agents methylmethanesulfonate (MMS), ethyl methanesulfonate, N-methyl-N'-nitro-N-nitrosoguanidine, and 4-nitroquinoline 1-oxide. The genetic syndromes studied were: (a) adenomatosis of the colon and rectum only, an autosomal dominant trait; (b) Turcot's syndrome, a rare autosomal recessive polyposis syndrome also characterized by central nervous system tumors; and (c) Gardner's syndrome, an autosomal dominant syndrome which, in addition to intestinal polyposis, is also clinically characterized by osteomas and soft tissue tumors. Fibroblasts from a patient with Turcot's syndrome were hypersensitive to MMS, having a D0 value of 0.24 mM (p less than 0.01) versus the normal average D0 of 0.36 mM and a D10 value of 0.95 mM (p less than 0.01) compared with the normal average value of 1.3 mM. Fibroblasts from the Gardner's syndrome proband were moderately sensitive to MMS, ethyl methanesulfonate, and N-methyl-N'-nitro-N-nitrosoguanidine due to significant differences of D10 values of 0.60 mM (p less than 0.01), 15 mM (p less than 0.01), and 4.8 microM (p less than 0.025), respectively, versus the normal average values of 1.3 mM, 28 mM, and 9.4 microM. Fibroblasts from the clinically affected Gardner's syndrome daughter of the proband were significantly more sensitive to MMS treatment, D0 of 0.22 mM (p less than 0.01) versus the normal average D0 of 0.36 mM and a D10 of 0.97 mM (p less than 0.01) versus the normal average. This differential sensitivity to the several DNA alkylating agents suggests that different mechanisms of hypersensitivity to these chemicals may be associated with fibroblasts from the various forms of familial polyposis

  20. Whole venom of Loxosceles similis activates caspases-3, -6, -7, and -9 in human primary skin fibroblasts

    Science.gov (United States)

    Dantas, Arthur Estanislau; Horta, Carolina Campolina Rebello; Martins, Thais M. M.; do Carmo, Anderson Oliveira; de Oliveira Mendes, Bárbara Bruna Ribeiro; Goes, Alfredo M.; Kalapothakis, Evanguedes; Gomes, Dawidson A.

    2014-01-01

    Spiders of the Loxosceles genus represent a risk to human health due to the systemic and necrotic effects of their bites. The main symptoms of these bites vary from dermonecrosis, observed in the majority of cases, to occasional systemic hemolysis and coagulopathy. Although the systemic effects are well characterized, the mechanisms of cell death triggered by the venom of these spiders are poorly characterized. In this study, we investigated the cell death mechanisms induced by the whole venom of the spider Loxosceles similis in human skin fibroblasts. Our results show that the venom initiates an apoptotic process and a caspase cascade involving the initiator caspase-9 and the effector caspases-3, -6, and -7. PMID:24726468

  1. Altered chaperone and protein turnover regulators expression in cultured skin fibroblasts from type 1 diabetes mellitus with nephropathy.

    Science.gov (United States)

    Tessari, Paolo; Puricelli, Lucia; Iori, Elisabetta; Arrigoni, Giorgio; Vedovato, Monica; James, Peter; Coracina, Anna; Millioni, Renato

    2007-03-01

    In type-1 diabetes mellitus (T1DM) with diabetic nephropathy (DN), accumulation of abnormal proteins in the kidney and other tissues may derive from constitutive alterations of intracellular protein recognition, assembly, and turnover. We characterized the proteins involved in these functions in cultured skin fibroblasts from long-term T1DM patients with [DN+] or without [DN-] nephropathy but similar metabolic control, and from matched healthy subjects. 2-D gel electrophoresis and MS-MALDI analysis were employed. The [DN+] T1DM patients, compared with the two other groups, exhibited increased abundance of a high-molecular weight isoform of protein disulphide-isomerase A3 and a decrease of two low-molecular weight isoforms. They also had increased levels of heat shock protein (HSP) 60 kDa isoform #A4, of HSP71 kDa isoform #A30, and of HSP27 kDa isoform #6, whereas the HSP27 kDa isoforms #A90 and #A71 were decreased. Cathepsin beta-2 (#40), the cation-independent mannose 6-phosphate receptor binding protein 1 (CIMPR) (#A27), and annexin 2 (#A9) were also decreased in the [DN+] T1DM patients, whereas the RNA-binding protein regulatory subunity (#38) and the translationally-controlled tumor protein (TCTP) (#A45) were increased. These changes of chaperone-like proteins in fibroblasts may highlight those of the kidney and be patho-physiologically related to the development of nephropathy in T1DM. PMID:17330940

  2. Cyclobutane-type pyrimidine photodimer formation and induction of ornithine decarboxylase in human skin fibroblasts after UV irradiation

    International Nuclear Information System (INIS)

    Cyclobutane-type pyrimidine photodimers as well as the induction of ornithine decarboxylase (ODC) may serve as biochemical markers of the mutagenic and carcinogenic effects of ultraviolet light (UV). For this reason, it is important to compare the formation of pyrimidine dimers with the induction of ODC in human skin fibroblasts after irradiation with UVC (200-290 nm) and UVB (290-320 nm). In our studies we determined cytosine-thymine (C-T) as well as thymine-thymine dimer yields (T-T) by high-pressure liquid chromatography in cultures of neonatal normal human foreskin-derived fibroblasts after irradiation with UVC and UVB light. It was found that the yield of dimerization and the ratio of T-T/C-T decreased from the UVC to the UVB region. Time-course studies of ODC-induction in the same cells indicated that the maximal activity after UVB irradiation was retarded compared to UVC exposure. For the UV-induced ODC-levels, however, no significant difference in maximal induction could be measured after UVC and UVB irradiation at fluences where comparable yields of thymine dimerization are produced. Similar ODC-maxima were obtained with strains from children, while cells from adults showed significantly less pronounced ODC induction, indicating that ODC-response decreases with age and may therefore be used as a marker of aging

  3. ANALYSIS OF DNA DAMAGE AND REPAIR IN SKIN FIBROBLASTS OF INFANT AND OLDER CHILDREN USING THE IN VITRO ALKALINE COMET ASSAY

    Science.gov (United States)

    ANALYSIS OF DNA DAMAGE AND REPAIR IN SKIN FIBROBLASTS OF INFANT AND OLDER CHILDREN USING THE IN VITRO ALKALINE COMET ASSAY, Alan H. Tennant1, Geremy W. Knapp1 and Andrew D. Kligerman1, 1Environmental Carcinogenesis Division, National Health and Environmental Effects Research Lab...

  4. The role of fibroblast growth factor receptor 2b in skin homeostasis and cancer development

    OpenAIRE

    Grose, Richard; Fantl, Vera; Werner, Sabine; Chioni, Athina-Myrto; Jarosz, Monika; Rudling, Robert; Cross, Barbara; Hart, Ian R.; Dickson, Clive

    2007-01-01

    The epithelial isoform of fibroblast growth factor receptor 2 (Fgfr2b) is essential for embryogenesis, and Fgfr2b-null mice die at birth. Using Cre-Lox transgenics to delete Fgfr2b in cells expressing keratin 5, we show that mice lacking epidermal Fgfr2b survive into adulthood but display striking abnormalities in hair and sebaceous gland development. Epidermal hyperthickening develops with age, and 10% of mutant mice develop spontaneous papillomas, demonstrating the role of Fgfr2b in post-na...

  5. Specific toxicity of aphidicolin to ultraviolet-irradiated excision proficient human skin fibroblasts

    International Nuclear Information System (INIS)

    Aphidicolin, a specific inhibitor of the eucaryotic ? polymerase, has been employed to study the role of this enzyme in repair of potentially lethal damage (PLD) induced by far u.v. (254 nm) radiation in normal and repair defective primary human fibroblasts. There is a strong concentration dependent specific toxicity to cells treated with a fluence of 6 Jm-2 of far-u.v. radiation and incubated with aphidicolin for 2 days over the concentration range 0.0025-2.5 ?g/ml. A similar effect is seen with a xeroderma pigmentosum (XP) variant (excision proficient) strain but there is no specific toxicity to u.v irradiated excision deficient XP cells of complementation group A. Inactivation of irradiated excision proficient fibroblasts is rapid over the first 6 h of aphidicolin (1 ?g/ml) treatment but the reaction takes 2 days or longer to complete depending on the u.v. dose. These results demonstrate that the apparent uncoupling of excision repair seen previously by other investigators prevents repair of PLD and is lethal to the cells. (author)

  6. Specific toxicity of aphidicolin to ultraviolet-irradiated excision proficient human skin fibroblasts

    International Nuclear Information System (INIS)

    Aphidicolin, a specific inhibitor of the eucaryotic alpha polymerase, has been employed to study the role of this enzyme in repair of potentially lethal damage (PLD) induced by far u.v. (254 nm) radiation in normal and repair defective primary human fibroblasts. There is strong concentration dependent specific toxicity to cells treated with a fluence of 6 Jm-2 of far-u.v. radiation and incubated with aphidicolin for 2 days over the concentration range 0.0025-2.5 micrograms/ml. A similar effect is seen with a xeroderma pigmentosum (XP) variant (excision proficient) strain but there is no specific toxicity to u.v. irradiated excision deficient XP cells of complementation group A. Inactivation of irradiated excision proficient fibroblasts is rapid over the first 6 h of aphidicolin (1 microgram/ml) treatment but the reaction takes 2 days or longer to complete depending on the u.v. dose. These results demonstrate that the apparent uncoupling of excision repair seen previously by other investigators prevents repair of PLD and is lethal to the cells

  7. Toxicity evaluation of ZnO nanostructures on L929 fibroblast cell line using MTS assay

    Energy Technology Data Exchange (ETDEWEB)

    Bakhori, Siti Khadijah Mohd; Mahmud, Shahrom; Ann, Ling Chuo [Nano-optoelectronics Research and Technology Laboratory (NOR.), School of Physics, Universiti Sains Malaysia, 11800, USM, Pulau Pinang (Malaysia); Mohamed, Azman Seeni; Saifuddin, Siti Nazmin [Integrative Medicine Cluster, Advanced Medical and Dental Institute, Universiti Sains Malaysia, Bandar Putra Bertam, 13200 Kepala Batas, Pulau Pinang (Malaysia); Masudi, Sam’an Malik; Mohamad, Dasmawati [Craniofacial Science Laboratory, School of Dentistry, Health Campus, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan (Malaysia)

    2015-04-24

    ZnO has wide applications in medical and dentistry apart from being used as optoelectronic devices such as solar cells, photodetectors, sensors and light emitting diodes (LEDs). Therefore, the toxicity evaluation is important to know the toxicity level on normal cell line. The toxicity of two grades ZnO nanostructures, ZnO-4 and ZnO-8 have been carried out using cytotoxicity test of MTS assay on L929 rat fibroblast cell line. Prior to that, ZnO-4 and ZnO-8 were characterized for its morphology, structure and optical properties using FESEM, X-ray diffraction, and Photoluminescence respectively. The two groups revealed difference in morphology and exhibit slightly shifted of near band edge emission of Photoluminescence other than having a similar calculated crystallite size of nanostructures. The viability of cells after 72h were obtained and the statistical significance value was calculated using SPSS v20. The p value is more than 0.05 between untreated and treated cell with ZnO. This insignificant value of p>0.05 can be summarized as a non-toxic level of ZnO-4 and ZnO-8 on the L929 cell line.

  8. Toxicity evaluation of ZnO nanostructures on L929 fibroblast cell line using MTS assay

    Science.gov (United States)

    Bakhori, Siti Khadijah Mohd; Mahmud, Shahrom; Ann, Ling Chuo; Mohamed, Azman Seeni; Saifuddin, Siti Nazmin; Masudi, Sam'an Malik; Mohamad, Dasmawati

    2015-04-01

    ZnO has wide applications in medical and dentistry apart from being used as optoelectronic devices such as solar cells, photodetectors, sensors and light emitting diodes (LEDs). Therefore, the toxicity evaluation is important to know the toxicity level on normal cell line. The toxicity of two grades ZnO nanostructures, ZnO-4 and ZnO-8 have been carried out using cytotoxicity test of MTS assay on L929 rat fibroblast cell line. Prior to that, ZnO-4 and ZnO-8 were characterized for its morphology, structure and optical properties using FESEM, X-ray diffraction, and Photoluminescence respectively. The two groups revealed difference in morphology and exhibit slightly shifted of near band edge emission of Photoluminescence other than having a similar calculated crystallite size of nanostructures. The viability of cells after 72h were obtained and the statistical significance value was calculated using SPSS v20. The p value is more than 0.05 between untreated and treated cell with ZnO. This insignificant value of p>0.05 can be summarized as a non-toxic level of ZnO-4 and ZnO-8 on the L929 cell line.

  9. Toxicity evaluation of ZnO nanostructures on L929 fibroblast cell line using MTS assay

    International Nuclear Information System (INIS)

    ZnO has wide applications in medical and dentistry apart from being used as optoelectronic devices such as solar cells, photodetectors, sensors and light emitting diodes (LEDs). Therefore, the toxicity evaluation is important to know the toxicity level on normal cell line. The toxicity of two grades ZnO nanostructures, ZnO-4 and ZnO-8 have been carried out using cytotoxicity test of MTS assay on L929 rat fibroblast cell line. Prior to that, ZnO-4 and ZnO-8 were characterized for its morphology, structure and optical properties using FESEM, X-ray diffraction, and Photoluminescence respectively. The two groups revealed difference in morphology and exhibit slightly shifted of near band edge emission of Photoluminescence other than having a similar calculated crystallite size of nanostructures. The viability of cells after 72h were obtained and the statistical significance value was calculated using SPSS v20. The p value is more than 0.05 between untreated and treated cell with ZnO. This insignificant value of p>0.05 can be summarized as a non-toxic level of ZnO-4 and ZnO-8 on the L929 cell line

  10. Recovery of fibroblast-like cells from refrigerated goat skin up to 41 d of animal death.

    Science.gov (United States)

    Okonkwo, Charles; Singh, Mahipal

    2015-05-01

    Successful cloning of animals using somatic cell nuclear transfer requires undamaged nuclear DNA from desired donor cell types. In vitro culture of cells is one way of ensuring nuclear integrity. The goal of this study was to evaluate the limits of postmortem cell survival/culture in refrigerated goat ear skin tissues which could be used for long-term storage and cloning of animals in future. To achieve this, 60 explants from 6 different goats were cultured after 0, 3, 6, 9, 13, 16, 20, 23, 27, 30, 33, 37, and 41 d postmortem and observed under inverted microscope for outgrowth of fibroblast-like cells, after 10-12 d of culture. Explants from all time points including 19% from 41-dpm tissues exhibited outgrowth. However, the percentage of outgrowth positive explants, as well as culture confluence, reduced with increasing postmortem time interval. Cell cultures established from primary outgrowth of 41-dpm tissues when compared for their growth profile with similarly obtained 0-dpm cultures revealed similar growth curve and cell morphology. Cytogenetic analysis of 41-dpm tissue-derived cell populations revealed a normal female karyotype with 60 XX homologous chromosomes indicating genetic stability of the cell population. In conclusion, these results show that refrigerated skin tissue remains alive for more than a month and that the cells derived from such tissues are normal and can be cryopreserved for long-term storage and future cloning of animals with desired genetics. PMID:25539865

  11. Generation of human iPS cell line ihFib3.2 from dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    Fernanda Cristina Paccola Mesquita

    2015-11-01

    Full Text Available The human ihFib3.2 iPS cell line was generated from dermal fibroblasts obtained from a healthy donor. Lentiviral particles were produced with the polycistronic hSTEMCCA vector with Oct4, Sox2, cMyc and Klf4 as reprogramming factors.

  12. Effects of plant sterols derived from Aloe vera gel on human dermal fibroblasts in vitro and on skin condition in Japanese women

    Science.gov (United States)

    Tanaka, Miyuki; Misawa, Eriko; Yamauchi, Koji; Abe, Fumiaki; Ishizaki, Chiaki

    2015-01-01

    Background Aloe is known for its topical use for treating wounds and burns. Many previous studies reported the healing effects of Aloe vera. However, there are few clinical studies on the effect of orally administered A. vera gel on the skin. Aloe sterols are a type of plant sterols that have the capability to regulate the metabolism of glucose and lipids. In a recent study, we confirmed that ingested Aloe sterols reached the peripheral tissues through the bloodstream. However, their influence on dermal fibroblasts has not been investigated. Methods First, we investigated the capability of Aloe sterols (cycloartenol and lophenol) to stimulate human dermal fibroblasts in vitro. Then, we investigated the effect of intake of Aloe vera gel powder (AVGP) containing 40 ?g Aloe sterols on the skin conditions in Japanese women with dry skin in a randomized, double-blind, placebo-controlled trial. Results After cocultivation with Aloe sterols, the production of collagen and hyaluronic acid increased by approximately two-fold and 1.5-fold, and gene expression levels of these enzymes responsible for their synthesis were also observed in human dermal fibroblasts. An increase in arm skin hydration was observed at 8 weeks in the AVGP group, whereas a slight decrease in arm skin hydration was noted in the placebo group. However, there was no statistical difference between AVGP and placebo groups in skin moisture. In subgroup analysis, the change in the mean wrinkle depth was significantly lower in the AVGP group than in the control group. In addition, percent body fat after 8 weeks was significantly lower in the AVGP group. No AVGP intake-dependent harmful phenomenon was observed during the intake period. Conclusion The present study confirms that daily oral Aloe sterol-containing AVGP significantly reduced facial wrinkles in women aged ?40 years, and Aloe sterols stimulate collagen and hyaluronic acid production by human dermal fibroblasts. PMID:25759593

  13. Host-cell reactivation of UV-irradiated and chemically-treated herpes simplex virus-1 by xeroderma pigmentosum, xp heterozygotes and normal skin fibroblasts

    International Nuclear Information System (INIS)

    The host-cell reactivation of UV-irradiated and N-acetoxy-2-acetylamino-fluorene-treated herpes simplex virus type 1 strain MP was studied in normal and xeroderma pigmentosum human skin fibroblasts. Virus treated with either agent demonstrated lower survival in XP cells from complementation groups A, B, C and D than in normal fibroblasts. The relative reactivation ability of XP cells from the different genetic complementation groups was found to be the same for both irradiated and chemically treated virus. In addition, the inactivation kinetics for virus treated with either agent in the XP variant were comparable to that seen in normal skin fibroblasts. The addition of 2 or 4 mmoles caffeine to the post-infection assay medium had no effect on the inactivation kinetics of virus treated by either agent in the XP variant or in XP cells from the different genetic complementation groups. Treatment of the virus with nitrogen mustard resulted in equivalent survival in normal and XP genetic complementation group D cells. No apparent defect was observed in the ability of XP heterozygous skin fibroblasts to repair virus damaged with up to 100 μg N-acetoxy-2-acetylaminofluorene per ml. These findings indicate that the repair of UV-irradiated and N-acetoxy-2-acetylaminofluorene-treated virus is accomplished by the same pathway or different pathways sharing a common intermediate step and that the excision defect of XP cells plays little if any role in the reactivation of nitrogen mustard treated virus. (Auth.)

  14. Effect of Microalgal Extracts of Tetraselmis suecica against UVB-Induced Photoaging in Human Skin Fibroblasts

    OpenAIRE

    Jo, Wol Soon; Yang, Kwang Mo; Park, Hee Sung; Kim, Gi Yong; Nam, Byung Hyouk; Jeong, Min Ho; Choi, Yoo Jin

    2012-01-01

    Exposure of cells to ultraviolet B (UVB) radiation can induce production of free radicals and reactive oxygen species (ROS), which damage cellular components. In addition, these agents can stimulate the expression of matrix metalloproteinase (MMP) and decrease collagen synthesis in human skin cells. In this study, we examined the anti-photoaging effects of extracts of Tetraselmis suecica (W-TS). W-TS showed the strongest scavenging activity against 2,2-difenyl-1-picrylhydrazyl (DPPH) and pero...

  15. Cationic star-shaped polymer as an siRNA carrier for reducing MMP-9 expression in skin fibroblast cells and promoting wound healing in diabetic rats

    Directory of Open Access Journals (Sweden)

    Li N

    2014-07-01

    Full Text Available Na Li,1,* Heng-Cong Luo,1,* Chuan Yang,1 Jun-Jie Deng,2 Meng Ren,1 Xiao-Ying Xie,1 Diao-Zhu Lin,1 Li Yan,1 Li-Ming Zhang2 1Department of Endocrinology, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, People’s Republic of China; 2DSAPM Lab and PCFM Lab, Institute of Polymer Science, Department of Polymer and Materials Science, School of Chemistry and Chemical Engineering, Sun Yat-sen University, Guangzhou, People’s Republic of China *These authors contributed equally to this work Background: Excessive expression of matrix metalloproteinase-9 (MMP-9 is deleterious to the cutaneous wound-healing process in the context of diabetes. The aim of the present study was to explore whether a cationic star-shaped polymer consisting of ß-cyclodextrin (ß-CD core and poly(amidoamine dendron arms (ß-CD-[D3]7 could be used as the gene carrier of small interfering RNA (siRNA to reduce MMP-9 expression for enhanced diabetic wound healing. Methods: The cytotoxicity of ß-CD-(D37 was investigated by 3-(4,5-Dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide assay (MMT method in the rat CRL1213 skin fibroblast cell line. The transfection efficiency of ß-CD-(D37/MMP-9-small interfering RNA (siRNA complexes was determined by confocal microscopy and flow cytometry. Quantitative real time (RT polymerase chain reaction was performed to measure the gene expression of MMP-9 after the transfection by ß-CD-(D37/MMP-9-siRNA complexes. The ß-CD-(D37/MMP-9-siRNA complexes were injected on the wounds of streptozocin-induced diabetic rats. Wound closure was measured on days 4 and 7 post-wounding. Results: ß-CD-(D37 exhibited low cytotoxicity in fibroblast cells, and easily formed the complexes with MMP-9-siRNA. The ß-CD-(D37/MMP-9-siRNA complexes were readily taken up by fibroblast cells, resulting in the downregulation of MMP-9 gene expression (P<0.01. Animal experiments revealed that the treatment by ß-CD-(D37/MMP-9-siRNA complexes enhanced wound closure in diabetic rats on day 7 post-wounding (P<0.05. Conclusion: ß-CD-(D37 may be used as an efficient carrier for the delivery of MMP-9-siRNA to reduce MMP-9 expression in skin fibroblast cells and promote wound healing in diabetic rats. Keywords: gene carrier, small interfering RNAs, matrix metalloproteinase-9, diabetic foot ulceration

  16. Dandelion Extracts Protect Human Skin Fibroblasts from UVB Damage and Cellular Senescence.

    Science.gov (United States)

    Yang, Yafan; Li, Shuangshuang

    2015-01-01

    Ultraviolet (UV) irradiation causes damage in skin by generating excessive reactive oxygen species (ROS) and induction of matrix metalloproteinases (MMPs), leading to skin photoageing. Dandelion extracts have long been used for traditional Chinese medicine and native American medicine to treat cancers, hepatitis, and digestive diseases; however, less is known on the effects of dandelion extracts in skin photoageing. Here we found that dandelion leaf and flower extracts significantly protect UVB irradiation-inhibited cell viability when added before UVB irradiation or promptly after irradiation. Dandelion leaf and flower extracts inhibited UVB irradiation-stimulated MMP activity and ROS generation. Dandelion root extracts showed less action on protecting HDFs from UVB irradiation-induced MMP activity, ROS generation, and cell death. Furthermore, dandelion leaf and flower but not root extracts stimulated glutathione generation and glutathione reductase mRNA expression in the presence or absence of UVB irradiation. We also found that dandelion leaf and flower extracts help absorb UVB irradiation. In addition, dandelion extracts significantly protected HDFs from H2O2-induced cellular senescence. In conclusion, dandelion extracts especially leaf and flower extracts are potent protective agents against UVB damage and H2O2-induced cellular senescence in HDFs by suppressing ROS generation and MMP activities and helping UVB absorption. PMID:26576225

  17. Establishment of a pig fibroblast-derived cell line for locus-directed transgene expression in cell cultures and blastocysts

    DEFF Research Database (Denmark)

    Jakobsen, Jannik E; Li, Juan; Moldt, Brian; Kragh, Peter M; Callesen, Henrik; Hertz, Jens Michael; Bolund, Lars; Jørgensen, Arne Lund; Mikkelsen, Jacob Giehm; Nielsen, Anders Lade

    2011-01-01

    We report the establishment of a spontaneously immortalized pig cell line designated Pig Flip-in Visualize (PFV) for locus-directed transgene expression in pig cells and blastocysts. The PFV cell line was isolated from pig ear fibroblasts transfected with a Sleeping Beauty DNA transposon...... transfer. PFV cells supported Flp mediated cassette exchange for transgene substitution of eGFP with dsRED, and the dsRED transgenic PFV cells generated blastocysts with transgene expression. Hence, the PFV cell line constitutes a valuable pig equivalent to transformed cell lines from other mammalian...

  18. Radiosensitivity of skin fibroblasts and lymphocytes from atomic bomb survivors in Hiroshima

    International Nuclear Information System (INIS)

    In the last 30 years or so, the existence of individual differences in in vivo radiation sensitivity has been well recognized in the response of normal tissues, particularly skin tissue, of cancer patients in the course of radiation therapy. If a large variation in radiosensitivity truly exists, it is very important to compare the radiosensitivity between the A-bomb survivors and a general population. If A-bomb survivors include a disproportionately large number of either radioresistant or radiosensitive persons, the surviving population would provide a biased estimate of the true risk of radiogenic cancer. 14 refs., 1 fig., 1 tab

  19. Efficacy of a single high dose versus multiple low doses of LLLT on wounded skin fibroblasts

    Science.gov (United States)

    Hawkins, Denise H.; Abrahamse, Heidi

    2007-07-01

    Background/purpose: In vivo studies have demonstrated that phototherapy accelerates wound healing in the clinical environment; however the exact mechanism is still not completely understood. The main focus of this study was to use in vitro laboratory results to establish an effective treatment regimen that may be practical and applicable to the clinical environment. This in vitro study aimed to compare the cellular responses of wounded fibroblasts following a single exposure of 5 J/cm2 or multiple exposures of low doses (2.5 J/cm2 or 5 J/cm2) on one day of the week to a single application of a higher dose (16 J/cm2) on day 1 and day 4. Methodology: Cellular responses to Helium-Neon (632.8 nm) laser irradiation were evaluated by measuring changes in cell morphology, cell viability, cell proliferation, membrane integrity and DNA damage. Results: Wounded cells exposed to 5 J/cm2 on day 1 and day 4 showed an increase in cell viability, increase in the release of bFGF, increase in cell density, decrease in ALP enzyme activity and decrease in caspase 3/7 activity indicating a stimulatory effect. Wounded cells exposed to three doses of 5 J/cm2 on day 1 showed a decrease in cell viability and cell proliferation and an increase in LDH cytotoxicity and DNA damage indicating an inhibitory effect. Conclusion: Results indicate that cellular responses are influenced by the combination of dose administered, number of exposures and time between exposures. Single doses administered with sufficient time between exposures is more beneficial to restoring cell function than multiple doses within a short period. Although this work confirms previous reports on the cumulative effect of laser irradiation it provides essential information for the initiation of in vivo clinical studies.

  20. Heme oxygenase is the major 32-kDa stress protein induced in human skin fibroblasts by UVA radiation, hydrogen peroxide, and sodium arsenite

    International Nuclear Information System (INIS)

    We have shown that UVA (320-380 nm) radiation, hydrogen peroxide, and sodium arsenite induce a stress protein of approximately 32 kDa in human skin fibroblasts. The synthesis and cloning of cDNA from arsenite-induced mRNA populations have now allowed us to unequivocally identify the 32-kDa protein as heme oxygenase. By mRNA analysis we have shown that the heme oxygenase gene is also induced in cultured human skin fibroblasts by UVA radiation, hydrogen peroxide, cadmium chloride, iodoacetamide, and menadione. The known antioxidant properties of heme catabolites taken together with the observation of a high level of induction of the enzyme in cells from an organ not involved in hemoglobin breakdown strongly supports the proposal that the induction of heme oxygenase may be a general response to oxidant stress and constitutes an important cellular defense mechanism against oxidative damage

  1. Ionising radiation response of mouse embryonic fibroblast cell lines lacking cohesin REC8

    International Nuclear Information System (INIS)

    Full text: The cohesin complex is required for the cohesion of sister chromatids. Cohesin is loaded onto chromosomes during DNA replication and maintains sister chromatid association until anaphase. The cohesin complex is also implicated in homologous recombination, the upkeep of chromosome stability, the repair of double stranded DNA breaks (dsbs) and most recently, chromatin remodelling. REC8 is a key component of the meiotic cohesin complex that we showed was conserved from yeast to humans (Parisi, et. al., 1999). We identified a mouse ortholog of the Rec8 gene and generated Rec8 mutant mice by targeted deletion. To determine the role of REC8 in DNA repair, we derived embryonic fibroblast cell lines from Rec8 mutant mice. By clonogenic assay we are evaluating the resistance of Rec8 -/- cells to ionising (IR) radiation, an indirect measure of repair of dsbs. The study may provide insights into the involvement of cohesins in recombination and IR response. Reference: Parisi, S., McKay, M.J., Molnar, M., Thompson, M.A., van der Spek, P.J., van Drunken-Schoenmaker, E., Kanaar, R., Lehmann, E., Hoeijmakers, J.H.J. and Kohli, J., 1999. Rec8p, a meiotic recombination and sister chromatid cohesion phosphoprotein of the Rad21 family conserved from fission yeast to humans. Mol Cell Biol. 19(5):3515-3528

  2. The effect of Centella asiatica, vitamins, glycolic acid and their mixtures preparations in stimulating collagen and fibronectin synthesis in cultured human skin fibroblast.

    Science.gov (United States)

    Hashim, Puziah

    2014-03-01

    Centella asiatica (Linn.) Urban is well known in promoting wound healing and provides significant benefits in skin care and therapeutic products formulation. Glycolic acid and vitamins also play a role in the enhancement of collagen and fibronectin synthesis. Here, we evaluate the specific effect of Centella asiatica (CA), vitamins, glycolic acid and their mixture preparations to stimulate collagen and fibronectin synthesis in cultured human fibroblast cells. The fibroblast cells are incubated with CA, glycolic acid, vitamins and their mixture preparations for 48 h. The cell lysates were analyzed for protein content and collagen synthesis by direct binding enzyme immunoassay. The fibronectin of the cultured supernatant was measured by sandwich enzyme immunoassay. The results showed that CA, glycolic acid, vitamins A, E and C significantly stimulate collagen and fibronectin synthesis in the fibroblast. Addition of glycolic acid and vitamins to CA further increased the levels of collagen and fibronectin synthesis to 8.55 and 23.75 ?g/100 ?g, respectively. CA, glycolic acid, vitamins A, E, and C, and their mixtures demonstrated stimulatory effect on both extra-cellular matrix synthesis of collagen and fibronectin in in vitro studies on human foreskin fibroblasts, which is beneficial to skin care and therapeutic products formulation. PMID:24577907

  3. Redox-dependent induction of antioxidant defenses by phenolic diterpenes confers stress tolerance in normal human skin fibroblasts: Insights on replicative senescence.

    Science.gov (United States)

    Carvalho, Ana C; Gomes, Andreia C; Pereira-Wilson, Cristina; Lima, Cristovao F

    2015-06-01

    Mild stress-induced hormesis represents a promising strategy for targeting the age-related accumulation of molecular damage and, therefore, for preventing diseases and achieving healthy aging. Fruits, vegetables, and spices contain a wide variety of hormetic phytochemicals, which may explain the beneficial health effects associated with the consumption of these dietary components. In the present study, the induction of cellular antioxidant defenses by the phenolic diterpenes carnosic acid (CA) and carnosol (CS) were studied in normal human skin fibroblasts, and insights into the aging process at the cellular level investigated. We observed that CA and CS induced several cytoprotective enzymes and antioxidant defenses in human fibroblasts, whose induction was dependent on the cellular redox state for CS and associated with Nrf2 signaling for both compounds. The stress response elicited by preincubation with CS conferred a cytoprotective action against a following oxidant challenge with tert-butyl hydroperoxide, confirming its hormetic effect. Preincubation of normal fibroblasts with CS also protected against hydrogen peroxide-induced premature senescence. Furthermore, cultivation of middle passage normal human skin fibroblasts in the presence of CS ameliorated the physiological state of cells during replicative senescence. Our results support the view that mild stress-induced antioxidant defenses by CS can confer stress tolerance in normal cells and may have important implications in the promotion of healthy aging. PMID:25744415

  4. The histamine degradative uptake pathway in human vascular endothelial cells and skin fibroblasts is dependent on extracellular Na+ and Cl-

    International Nuclear Information System (INIS)

    We have previously reported that human vascular endothelial cells and skin fibroblasts carry out degradation of [3H]histamine by a mechanism involving two successive enzymatic steps: imidazole ring tele-methylation by the cells' endogenous methyltransferase and subsequent amine oxidation by an exogenous diamine oxidase. Both histamine and the exogenous second enzyme in the pathway associate with the cells via separate binding sites or receptors. The enzymatic degradation process results in cellular accumulation of the proximal and distal metabolites tele-methylhistamine and 1-methyl-4-imidazoleacetic acid (MIAA). We have now demonstrated that this two-stage histamine degradative pathway is dependent on Na+ and Cl- in the extracellular environment. Accumulation of [3H] histamine-derived products is partially inhibited under conditions of Na+ deprivation and more substantially when Cl- is also withdrawn. The individual tele-methylation and amine oxidation enzymatic reactions themselves are unaffected or actually facilitated under these conditions. This indicates that it is the cellular mechanism for uptake coupled to the degradative pathway which reflects the cation and anion dependency. Restoration of degradative uptake displays a biphasic Na+ concentration curve, suggesting that the uptake process may be driven by multiple components. These findings indicate a role for both inward Na+ and Cl- ion movement in this cellular degradative uptake mechanism

  5. Semi-conservative deoxyribonucleic acid synthesis in unirradiated and ultraviolet-irradiated xeroderma pigmentosum and normal human skin fibroblasts

    International Nuclear Information System (INIS)

    Rates of semiconservative DNA synthesis have been investigated in asynchronous xeroderma pigmentosum (XP), XP variant, and normal human skin fibroblasts using the technique of cellular autoradiography. In unirradiated cells, no differences in DNA synthesis rates were detected among the three cell strains. Exposure to UV radiation caused the rate of DNA synthesis to decrease for at least three hours in all three cell strains. In the normal cell strain, recovery of the DNA synthetic rate occurred at later times following a UV fluence of 5 J/m2. At this same UV fluence, recovery was absent in classical XP cells during a 24 h post-irradiation period while it was slower than normal in XP variant cells. When the UV fluence to classical XP and XP variant cells was reduced so that survival in all three cell strains was approximately the same (25%), recovery of the DNA synthetic rate was similar in all three cell strains. These results are discussed in terms of current models of DNA replication in UV-irradiated cells and indicate: (1) that pyrimidine dimers are very effective blocks to DNA synthesis and (2) that there is no inherent defect in semiconservative DNA synthesis in either classical XP or XP variant cells which is independent of a defect in DNA repair capacity

  6. Activation of NF-κB in human skin fibroblasts by the oxidative stress generated by UVA radiation

    International Nuclear Information System (INIS)

    We have examined the role of the nucleus and the membrane in the activation of nuclear factor (NF)-kB by oxidant stress generated via the UVA (320-380 nm) component of solar radiation. Nuclear extracts from human skin fibroblasts that had been irradiated with UVA at doses that caused little DNA damage contained activated NF-κB that bound to its recognition sequence in DNA. The UVA radiation-dependent activation of NF-κB in enucleated cells confirmed that the nucleus was not involved. On the other hand, UVA radiation-dependent activation of NF-κB appeared to be correlated with membrane damage, and activation could be prevented by α-tocopherol and butylated hydroxytoluene, agents that inhibited UVA radiation-dependent peroxidation of cell membrane lipids. The activation of NF-κB by the DNA damaging agents UVC (200-290 nm) and UVB (290-320 nm) radiation also only occurred at doses where significant membrane damage was induced, and, overall, activation was not correlated with the relative levels of DNA damage induced by UVC/UVB and UVA radiations. We conclude that the oxidative modification of membrane components may be an important factor to consider in the UV radiation-dependent activation of NF-κB over all wavelength ranges examined. (Author)

  7. Low doses of nanodiamonds and silica nanoparticles have beneficial hormetic effects in normal human skin fibroblasts in culture.

    Science.gov (United States)

    Mytych, Jennifer; Wnuk, Maciej; Rattan, Suresh I S

    2016-04-01

    Nanodiamonds (ND) and silica nanoparticles (SiO2-NP) have been much investigated for their toxicity at high doses, little is known about their biological activity at low concentrations. Here we report the biphasic dose response of ND and SiO2-NP in modulating normal human facial skin fibroblasts (FSF1) in culture. ND and SiO2-NP at low concentration (up to 0.5 μg/ml) had beneficial effects on FSF1 in terms of increasing their proliferation and metabolic activity. Exposure of FSF1 cells to low levels of NP enhanced their wound healing ability in vitro and slowed down aging during serial passaging as measured by maintenance of youthful morphology, reduction in the rate of loss of telomeres, and the over all proliferative characteristics. Furthermore, NP treatment induced the activation of Nrf2- and FOXO3A-mediated cellular stress responses, including an increased expression of heme oxygenease (HO-1), sirtuin (SIRT1), and DNA methyltransferase II (DNMT2). These results imply that ND and SiO2-NP at low doses are potential hormetins, which exert mild stress-induced beneficial hormetic effects through improved survival, longevity, maintenance, repair and function of human cells. PMID:26814705

  8. Acyl CoA Binding Domain Containing 3 (ACBD3) Protein in Huntington’s Disease 
Human Skin Fibroblasts

    Czech Academy of Sciences Publication Activity Database

    Kratochvílová, H.; Rodinová, M.; Sládková, J.; Klempíř, J.; Lišková, Irena; Motlík, Jan; Zeman, J.; Hansíková, H.; Tesařová, M.

    2015-01-01

    Roč. 78, Suppl. 2 (2015), s. 34-38. ISSN 1210-7859. [Conference on Animal Models for neurodegenerative Diseases /3./. Liblice, 08.11.2015-10.11.2015] R&D Projects: GA MŠk ED2.1.00/03.0124 Institutional support: RVO:67985904 Keywords : Huntington’s disease * Acyl-CoA binding domain containing 3 protein * human skin fibroblasts Subject RIV: FH - Neurology Impact factor: 0.165, year: 2014

  9. The Apoptotic Effects of the P300 Activator on Breast Cancer and Lung Fibroblast Cell Lines

    Directory of Open Access Journals (Sweden)

    Mohammad Reza Salahshoor

    2013-10-01

    Full Text Available Background: P300 is an enzyme that acetylates histones during stress. It alsoacetylates several non-histone proteins, including P53 which is the most important tumorsuppressor gene. P53 plays an important role in the apoptosis of tumor cells. Hereby,this study describes the potency of cholera toxin B subunit as a P300 activator to induceapoptosis in a breast cancer cell line (MCF-7 and a lung fibroblast cell line (MRC-5as a non-tumorigenic control sample. Methods: MCF-7 and MRC-5 were cultured in RPMI-1640 and treated with orwithout cholera toxin B subunit at the concentration of 85.43 μmol/L, based on the half-maximal inhibitory concentration index at different times (24, 48 and 72 h. Thepercentage of apoptotic cells was measured by flow cytometry. Real-time quantitativeRT-PCR was performed to estimate the mRNA expression of P300 in MCF-7 and MRC-5 with cholera toxin B subunit at different times. We used the ELISA and Bradford proteintechniques to detect levels of total and acetylated P53 protein generated in MCF-7 andMRC-5. Results: Our findings indicated that the cholera toxin B subunit effectively andsignificantly induced more apoptosis in MCF-7 compared to MRC-5. We showed thatexpression of P300 up-regulated by increasing the time of the cholera toxin B subunittreatment in MCF-7 but not in MRC-5. In addition, the acetylated and total P53protein levels increased more in MCF-7 cells than in MRC-5 cells.Conclusion: Cholera toxin B subunit induced significant cell death in MCF-7, butit could be well tolerated in MRC-5. Therefore, cholera toxin B subunit can besuggested as an anti-cancer agent.

  10. Altered binding of 125I-labeled calmodulin to a 46.5-kilodalton protein in skin fibroblasts cultured from patients with cystic fibrosis

    International Nuclear Information System (INIS)

    The levels of calmodulin and calmodulin-binding proteins have been determined in cultured skin fibroblasts from patients with cystic fibrosis (CF) and age- and sex-matched controls. Calmodulin ranged from 0.20 to 0.76 microgram/mg protein; there was no difference between calmodulin concentration in fibroblasts from CF patients and controls. Calmodulin-binding proteins of 230, 212, 204, 164, 139, 70, 59, 46.5, and 41 kD were identified. A protein with a mobility identical to the 59-kD calmodulin-binding protein was labeled by antiserum against calmodulin-dependent phosphatase. Although Ca2+/calmodulin-dependent phosphatase activity was detected, there was no different in activity between control and CF fibroblasts or in the level of phosphatase protein as determined by radioimmunoassay. Lower amounts of 125I-calmodulin were bound to the 46.5-kD calmodulin-binding protein in CF fibroblasts as compared with controls. The 46.5-kD calmodulin-binding protein may be reduced in CF fibroblasts or its structure may be altered resulting in a reduced binding capacity and/or affinity for calmodulin and perhaps reflecting, either directly or indirectly, the genetic defect responsible for cystic fibrosis

  11. Germline-Competent Mouse-Induced Pluripotent Stem Cell Lines Generated on Human Fibroblasts without Exogenous Leukemia Inhibitory Factor

    OpenAIRE

    Li, Chunliang; Yu, Hongyao; Ma, Yu; Shi, Guilai; Jiang, Jing; Gu, Junjie; Yang, Ying; Jin, Shibo; Wei, Zhe; Jiang, Hua; Li, Jinsong; JIN, YING

    2009-01-01

    Induced pluripotent stem (iPS) cells have attracted enormous attention due to their vast potential in regenerative medicine, pharmaceutical screening and basic research. Most prior established iPS cell lines were derived and maintained on mouse embryonic fibroblast (MEF) cells supplemented with exogenous leukemia inhibitory factor (LIF). Drawbacks of MEF cells impede optimization as well as dissection of reprogramming events and limit the usage of iPS cell derivatives in therapeutic applicati...

  12. Effects of plant sterols derived from Aloe vera gel on human dermal fibroblasts in vitro and on skin condition in Japanese women

    Directory of Open Access Journals (Sweden)

    Tanaka M

    2015-02-01

    Full Text Available Miyuki Tanaka,1 Eriko Misawa,1 Koji Yamauchi,1 Fumiaki Abe,1 Chiaki Ishizaki2 1Functional Food Research Department, Food Science and Technology Institute, Morinaga Milk Industry Co, Ltd, Zama, Kanagawa, 2Ebisu Skin Research Center, Inforward, Inc., Tokyo, Japan Background: Aloe is known for its topical use for treating wounds and burns. Many previous studies reported the healing effects of Aloe vera. However, there are few clinical studies on the effect of orally administered A. vera gel on the skin. Aloe sterols are a type of plant sterols that have the capability to regulate the metabolism of glucose and lipids. In a recent study, we confirmed that ingested Aloe sterols reached the peripheral tissues through the bloodstream. However, their influence on dermal fibroblasts has not been investigated. Methods: First, we investigated the capability of Aloe sterols (cycloartenol and lophenol to stimulate human dermal fibroblasts in vitro. Then, we investigated the effect of intake of Aloe vera gel powder (AVGP containing 40 g Aloe sterols on the skin conditions in Japanese women with dry skin in a randomized, double-blind, placebo-controlled trial. Results: After cocultivation with Aloe sterols, the production of collagen and hyaluronic acid increased by approximately two-fold and 1.5-fold, and gene expression levels of these enzymes responsible for their synthesis were also observed in human dermal fibroblasts. An increase in arm skin hydration was observed at 8 weeks in the AVGP group, whereas a slight decrease in arm skin hydration was noted in the placebo group. However, there was no statistical difference between AVGP and placebo groups in skin moisture. In subgroup analysis, the change in the mean wrinkle depth was significantly lower in the AVGP group than in the control group. In addition, percent body fat after 8 weeks was significantly lower in the AVGP group. No AVGP intake-dependent harmful phenomenon was observed during the intake period. Conclusion: The present study confirms that daily oral Aloe sterol-containing AVGP significantly reduced facial wrinkles in women aged ?40 years, and Aloe sterols stimulate collagen and hyaluronic acid production by human dermal fibroblasts. Keywords: aloe sterol, collagen, wrinkle

  13. Selection of reference genes for gene expression studies in ultraviolet B-irradiated human skin fibroblasts using quantitative real-time PCR

    Directory of Open Access Journals (Sweden)

    Qu Tao

    2011-02-01

    Full Text Available Abstract Background Reference genes are frequently used to normalise mRNA levels between different samples. The expression level of these genes, however, may vary between tissues or cells and may change under certain circumstances. Cytoskeleton genes have served as multifunctional tools for experimental studies as reference genes. Our previous studies have demonstrated that the expression of vimentin, one cytoskeletal protein, was increased in ultraviolet B (UVB-irradiated fibroblasts. Thus, we examined the expression of other cytoskeleton protein genes, ACTB (actin, beta, TUBA1A (tubulin, alpha 1a, and TUBB1 (tubulin, beta 1, in human dermal fibroblasts irradiated by UVB to determine which of these candidates were the most appropriate reference genes. Results Quantitative real-time PCR followed by analysis with the NormFinder and geNorm software programmes was performed. The initial screening of the expression patterns demonstrated that the expression of VIM was suppressed after UVB irradiation at doses ?25 mJ/cm2 and that the expression of TUBA1A was significantly reduced by UVB doses ?75 mJ/cm2 in cultured human dermal fibroblasts. The analysis of the experimental data revealed ACTB to be the most stably expressed gene, followed by GAPDH (aglyceraldehyde-3-phosphate dehydrogenase, under these experimental conditions. By contrast, VIM was found to be the least stable gene. The combination of ACTB and TUBB1 was revealed to be the gene pair that introduced the least systematic error into the data normalisation. Conclusion The data herein provide evidence that ACTB and TUBB1 are suitable reference genes in human skin fibroblasts irradiated by UVB, whereas VIM and TUBA1A are not and should therefore be excluded as reference genes in any gene expression studies involving UVB-irradiated human skin fibroblasts.

  14. Formation and repair of DNA damage induced by indirect action of ultraviolet light in normal and xeroderma pigmentosum skin fibroblasts

    International Nuclear Information System (INIS)

    The definitions of direct and indirect DNA damage are as follows: direct action, ''the attack of DNA occurs by a primary agent or a chemical derivative of the primary agent'', and indirect action, ''the attack of DNA occurs by active-O2 species which are formed by the reaction of a primary agent with a non-DNA target''. Comparative studies were performed on the formation and repair of DNA lesions induced by monochromatic light at 254, 265 and 313 nm. Attention was focused on human cells in culture, in particular on the skin fibroblasts from normal individuals and the patients with the autosomal recessive disease, Xerodermal pigmentosum (XP). The DNA lesions induced by the indirect action of active O2 species (superoxide-radicals, OH-radicals, singlet oxygen) become important. The most remarkable observation after the 313 nm irradiation on XP strains at 37 deg is the increased immediate fragmentation of pre-existing, parental DNA in the XP-variant strains. A late step in excision repair such as the ligation of parental DNA fragments may be deficient in the XP variants. The nuclease function involved in the removal of radiation lesions could be more active in the XP variants, or glycosylase function could be more active. The abnormality in de novo DNA synthesis in the XP variants may be the reflection of the primary defects in the repair of parental DNA templates rather than the defects in ''post-replication repair''. The abnormal function in DNA metabolism in the XP variants affects in parallel the repair of parental DNA and de novo DNA synthesis. (Yamashita, S.)

  15. Fibroblast sources: Where can we get them?

    Science.gov (United States)

    Fernandes, I R; Russo, F B; Pignatari, G C; Evangelinellis, M M; Tavolari, S; Muotri, A R; Beltro-Braga, P C B

    2016-03-01

    Fibroblasts are cells widely used in cell culture, both for transient primary cell culture or permanent as transformed cell lines. Lately, fibroblasts become cell sources for use in disease modeling after cell reprogramming because it is easily accessible in the body. Fibroblasts in patients will maintain all genetic background during reprogramming into induced pluripotent stem cells. In spite of their large use, fibroblasts are obtained after an invasive procedure, a superficial punch skin biopsy, collected under patient's local anesthesia. Taking into consideration the minimum patient's discomfort during and after the biopsy procedure, as well as the aesthetics aspect, it is essential to reflect on the best site of the body for the biopsy procedure combined with the success of getting robust fibroblast cultures in the lab. For this purpose, we compared the efficiency of four biopsy sites of the body (skin from eyelid, back of the ear, abdominal cesarean scar and groin). Cell proliferation assays and viability after cryopreservation were measured. Our results revealed that scar tissue provided fibroblasts with higher proliferative rates. Also, fibroblasts from scar tissues presented a higher viability after the thawing process. PMID:25060709

  16. CopA3 Peptide Prevents Ultraviolet-Induced Inhibition of Type-I Procollagen and Induction of Matrix Metalloproteinase-1 in Human Skin Fibroblasts

    Directory of Open Access Journals (Sweden)

    Dong-Hee Kim

    2014-05-01

    Full Text Available Ultraviolet (UV exposure is well-known to induce premature aging, which is mediated by matrix metalloproteinase-1 (MMP-1 activity. A 9-mer peptide, CopA3 (CopA3 was synthesized from a natural peptide, coprisin, which is isolated from the dung beetle Copris tripartitus. As part of our continuing search for novel bioactive natural products, CopA3 was investigated for its in vitro anti-skin photoaging activity. UV-induced inhibition of type-I procollagen and induction of MMP-1 were partially prevented in human skin fibroblasts by CopA3 peptide in a dose-dependent manner. At a concentration of 25 μM, CopA3 nearly completely inhibited MMP-1 expression. These results suggest that CopA3, an insect peptide, is a potential candidate for the prevention and treatment of skin aging.

  17. Host-cell reactivation of uv-irradiated and chemically treated Herpes simplex virus type 1 strain MP in normal and xeroderma pigmentosum skin fibroblasts

    International Nuclear Information System (INIS)

    The host-cell reactivation of UV-irradiated and N-acetoxy-2-acetylaminofluorene-treated herpes simplex virus type 1 strain mp was studied in normal human skin fibroblasts and xeroderma pigmentosum skin fibroblasts from XP genetic complementation groups A-D and in an XP variant. The increasing relative order for the host-cell reactivation of both types of damaged virus in the different complementation groups is A = D < B < C; XP variant = normal controls. XP complementation group D cells, which manifest the most severe inhibition of her ability for both UV-irradiated and N-acetoxy-2-acetylaminofluorene-treated virus, can reactivate nitrogen mustard treated HSV-1 mp to the same extent as normal cells. Together, these results indicate that (1) Excision repair of UV and N-acetoxy-2-acetylaminofluorene DNA damaged viruses share a common rate limiting enzymatic step and (2) The repair defect in xeroderma pigmentosum cells plays little or no role in the recovery of nitrogen mustard treated virus. The results of studies on the effect of caffeine on the survival of both UV- and N-acetoxy-2-acetylaminofluorene-treated virus in normal and XP cells imply that the reactivation of HSV-1 mp is mediated by an excision repair process with little if any recovery contributed by post-replication repair mechanisms. The host-cell reactivation of N-acetoxy-2-acetylaminofluorene-treated HSV-1 mp was also correlated with the defective UV-induced unscheduled DNA synthesis in two skin fibroblast strains established from a skin biopsy obtained from each of two juvenile females who had been clinically diagnosed as xeroderma pigmentosum. These findings are discussed in relation to the further characterization of the xeroderma pigmentosum phenotype and their possible utilization for the selection and isolation of new mammalian cell DNA repair mutants

  18. Absence of correlations between the radiosensitivity of human T-lymphocytes at G0 and skin fibroblasts at log phase from the same individuals

    International Nuclear Information System (INIS)

    Matched samples of peripheral T-lymphocytes and skin fibroblasts from a total of 22 patients who underwent various surgical procedures were tested for a dose-survival study using loss of colony-forming ability as the end point. The results showed that the mean D10 (the dose required to kill 90 % of the cells) SD was 3.58 0.21 Gy for T-lymphocytes irradiated at G0 and 3.19 0.37 Gy for skin fibroblasts irradiated at log phase. The coefficient of variation was found to be 6 % and 11 %, respectively. Contrary to expectation, regression analysis of the D10 values for the two cell types revealed no significant correlations. The absence of correlation is most probably derived from the fact that the apparent interindividual variability of dose-survival curves is largely caused by random experimental fluctuations, at least for lymphocytes. Possible reasons for the greater variability observed in the fibroblast assay are discussed. (author)

  19. Enhanced in vitro radiosensitivity of skin fibroblasts in two patients developing brain necrosis following AVM radiosurgery: a new risk factor with potential for a predictive assay

    International Nuclear Information System (INIS)

    Purpose: Radiosurgery is an effective treatment for arteriovenous malformations (AVM) with a low risk of developing brain necrosis. Models have been developed to predict the risk of complications. We postulated that genetic differences in radiosensitivity may also be a risk factor. Methods and Materials: Fibroblast cultures were established from skin biopsies in two AVM patients developing radiation necrosis. The results of clonogenic survival assays were compared to a parallel study with two groups of cancer patients treated with radiation: 1) patients without late side effects; 2) patients experiencing severe late sequelae. Results: The survival fraction at 2 Gy (SF2) of the 2 AVM patients was 0.17 (0.14-0.19) and 0.18 (0.14-0.22). The SF2's of the cancer patients ranged between 0.25-0.38 (mean = 0.31) for the control group, and between 0.10-0.20 (mean = 0.17) for the hypersensitive group. The SF2's of the AVM patients who developed brain necrosis were comparable to that of the hypersensitive group (p = 0.85) but significantly lower than the control group (p = 0.05). Conclusion: The two patients who developed radiation necrosis demonstrate increased fibroblast radiosensitivity. The SF2 of skin fibroblasts may potentially be used as a predictive assay to detect patients at risk for brain necrosis

  20. Correlation between normal tissue complications and in vitro radiosensitivity of skin fibroblasts derived from radiotherapy patients treated for variety of tumors

    International Nuclear Information System (INIS)

    Purpose: To assess the relationship between fibroblast intrinsic radiosensitivity in vitro and late reactions of normal tissues in patients treated by definitive radiotherapy for variety of tumors. Patients and Methods: Ten patients were selected for this study. They were treated by radical radiotherapy for variety of tumors, including non-Hodgkin's lymphoma, prostate, glottic larynx, anal canal, cervix, bladder, thyroid gland, and tonsil pillar. Five patients did not develop any significant late reactions (normally sensitive group, NS). The other five developed late complications in different normal tissues and organs that proved to be fatal in one patient (clinically hyper-sensitive group, HS). Fibroblast cultures were established from punch skin biopsy and radiosensitivity in vitro was measured. The survival fraction at 2 Gy (SF2) was calculated and compared between the two groups. Results: SF2 ranged between 0.10 and 0.38 with a mean of 0.24. The mean SF2 for each of the NS and the HS groups were 0.31 and 0.17, respectively. The non-parametric rank test of Mann-Whitney shows that the difference between the two groups is statistically significant (p = 0.01). Conclusion: This study indicates that the in vitro radiosensitivity of skin fibroblasts is correlated with late complications in different organs and normal tissues following radiotherapy for variety of tumors. It also lends support to the existence of a common genetic component determining the radiosensitivity of cells targeted by the late effects of ionizing radiation. Key words:

  1. The treatment effects of cultured epidermis, basic fibroblast growth factor and the combination of these two treatments in a radiation skin ulcer model (rat)

    International Nuclear Information System (INIS)

    The objective of this study was to evaluate the treatment effects of cultured epidermis, basic fibroblast growth factor (b-FGF) and the combination of these two treatments in a radiation skin ulcer model. The subjects were 9-week-old male inbred line rats and divided into two parts. Rats in one part were applied X-ray and rats in the other part were not. The dose of X-ray was 20 Gy. Wounds were full-thickness wounds. The ways of treatment were divided into four groups: control group, cultured epidermis group, b-FGF group, combination group (cultured epidermis+b-FGF). Wounds were observed on 5, 8, 11, 14, 17, 20, 23, 26 days after treatment. Wound healing rate was calculated and days needed to heal were counted. Relative hardness of scars was measured on the day of epithelization and on 12 and 21 days after epithelization. Wounds applied X-ray: Mean wound healing rate of cultured epidermis group and combination group was significantly higher than that of the two other groups on 8 and 11 days after treatment. Mean relative hardness of scars of cultured epidermis group and combination group was significantly lower than that of the two other groups on all measurement days. Mean days needed to heal of cultured epidermis group were significantly shorter than those of control group and b-FGF group. And those of combination group were significantly shorter than those of b-FGF group. As the shorter the days from making scars became, relative hardness of scars got lower. Wounds without X-ray: Mean wound healing rate of combination group was significantly lower than that of cultured epidermis group and control group on 5 days after treatment. Cultured epidermis graft can be an effective treatment for radiation skin ulcer. b-FGF can weaken the treatment effect of cultured epidermis graft depending on its density. There can be a positive correlation between relative hardness of scars and the days from making scars. (author)

  2. Effects of macelignan isolated from Myristica fragrans (nutmeg) on expression of matrix metalloproteinase-1 and type I procollagen in UVB-irradiated human skin fibroblasts

    International Nuclear Information System (INIS)

    Exposure to ultraviolet (UV) light causes premature skin aging that is associated with upregulated matrix metalloproteinases (MMPs) and decreased collagen synthesis. Macelignan, a natural lignan compound isolated from Myristica fragrans HOUTT. (nutmeg), has been reported to possess antioxidant and antiinflammatory activities. This study assessed the effects of macelignan on photoaging and investigated its mechanisms of action in UV-irradiated human skin fibroblasts (Hs68) by reverse transcription-polymerase chain reaction, Western blot analysis, 2', 7'-dichlorofluorescein diacetate assay, and enzyme-linked immunosorbent assay. Our results show that macelignan attenuated UV-induced MMP-1 expression by suppressing phosphorylation of mitogen-activated protein kinases (MAPKs) induced by reactive oxygen species. Macelignan also increased type I procollagen expression and secretion through transforming growth factor β (TGF-β)/Smad signaling. These findings indicate that macelignan regulates the expression of MMP-1 and type I procollagen in UV-irradiated human skin fibroblasts by modulating MAPK and TGF-β/Smad signaling, suggesting its potential as an efficacious antiphotoaging agent. (author)

  3. Effects of macelignan isolated from Myristica fragrans (Nutmeg) on expression of matrix metalloproteinase-1 and type I procollagen in UVB-irradiated human skin fibroblasts.

    Science.gov (United States)

    Lee, Kyung-Eun; Mun, Sukyeong; Pyun, Hee-Bong; Kim, Myung-Suk; Hwang, Jae-Kwan

    2012-01-01

    Exposure to ultraviolet (UV) light causes premature skin aging that is associated with upregulated matrix metalloproteinases (MMPs) and decreased collagen synthesis. Macelignan, a natural lignan compound isolated from Myristica fragrans HOUTT. (nutmeg), has been reported to possess antioxidant and antiinflammatory activities. This study assessed the effects of macelignan on photoaging and investigated its mechanisms of action in UV-irradiated human skin fibroblasts (Hs68) by reverse transcription-polymerase chain reaction, Western blot analysis, 2',7'-dichlorofluorescein diacetate assay, and enzyme-linked immunosorbent assay. Our results show that macelignan attenuated UV-induced MMP-1 expression by suppressing phosphorylation of mitogen-activated protein kinases (MAPKs) induced by reactive oxygen species. Macelignan also increased type I procollagen expression and secretion through transforming growth factor ? (TGF-?)/Smad signaling. These findings indicate that macelignan regulates the expression of MMP-1 and type I procollagen in UV-irradiated human skin fibroblasts by modulating MAPK and TGF-?/Smad signaling, suggesting its potential as an efficacious antiphotoaging agent. PMID:23037157

  4. Evidence for a positive correlation between in vitro radiosensitivity of normal human skin fibroblasts and the occurrence of subcutaneous fibrosis after radiotherapy

    International Nuclear Information System (INIS)

    A colony-forming assay of human skin fibroblast radiosensitivity was established in our laboratory and applied to primary skin biopsies from 12 women belonging to an unselected group of patients who received postmastectomy radiotherapy 10-12 years prior to this study. The aim was to investigate the relationship between in vitro radiosensitivity and the occurrence of subcutaneous fibrosis after radiotherapy. Early generations of normal skin fibroblasts in exponential growth were irradiated at room temperature at a high dose-rate to estimate the surviving fraction of colony-forming cells after single doses ranging from 1 to 8 Gy. A linear-quadratic cell survival curve was fitted to the data and from these fits the surviving fraction at 3.5 Gy (SF3.5) was estimated. Replicate experiments of different cell generations were made to validate the assay, and the between-patients variability was significantly larger than the assay variability for both SF2(p=0.002) and SF3.5(p=0.04). Patients were treated in the period 1978-1982 with a dose per fraction between 2.7 and 3.9 Gy, a total of 12 fractions at two fractions per week. They were evaluated with respect to the occurence of marked subcutaneous fibrosis in a total of 36 independent treatment fields. In each treatment field the total dose and dose per fraction at the relevant dosimetric reference depth as well as the length of follow-up were recorded. (Author)

  5. Reversible rearrangement of vimentin-type intermediate filaments in cultured human skin fibroblasts from patients with lysosomal storage diseases.

    Science.gov (United States)

    Tatiana, I; German, W

    1994-06-01

    Immunofluorescence microscopy shows that unlike cytoplasmic microtubules (MT), vimentin-type intermediate filaments (IF) are collected into ring-shaped structures in affected fibroblasts. The altered IF organization could be observed in monolayers of polarized fibroblasts in the prolonged stationary growth phase and-after replating-upon their initial spreading. Transition from a discoid to an extended cellular form is accompanied by centrifugal dislocation of ring-shaped IF structures towards the cell's active edge with gradual restoration of the radial fibrillar vimentin network. Spreading of affected cells occurred more slowly than that of control fibroblasts. PMID:8075625

  6. Development of a cell line from skin of goldfish, Carassius auratus, and effects of ascorbic acid on collagen deposition.

    Science.gov (United States)

    Lee, L E; Caldwell, S J; Gibbons, J

    1997-01-01

    Growth characteristics and collagen expression were investigated in GFSk-S1, a cell line derived from the skin of an adult goldfish (Carassius auratus). These cells are anchorage dependent, grow well in Leibovitz-15 medium with 10% fetal bovine serum, and have been subcultured routinely for 5 years. Cells at various passages have been successfully cryopreserved and thawed. GFSk-S1 cells show mainly a fibroblastic morphology at low density, but at confluence islands of epithelial-shaped cells appear among the fibroblastic cells. The cells require little maintenance, and cultures have been kept viable for more than 3 months without medium changes. Although best growth was observed at room temperature, cell proliferation still occurred at 28 degrees C, and a subline was maintained and passaged for over a year at 25 degrees C. Cells were exposed to various concentrations of ascorbic acid, and its effects on collagen secretion were monitored by light and electron microscopy. Under phase-contrast microscopy, confluent GFSk-S1 cells exposed to ascorbic acid at 50 micrograms/ml showed distinct development of fibres as early as 3 days after treatment. Histochemical staining for collagen demonstrated a thick network of fibres under a monolayer of ascorbic acid-treated GFSk-S1 cells, and observation by transmission electron microscopy showed collagen fibres with typical banding pattern. This cell line appears to show a stable genotype, as collagen expression was induced at all passages. GFSk-S1 could be useful for studies not only of regulation of protein synthesis, but also of cell differentiation and wound healing. PMID:9088943

  7. Altered protein expression pattern in skin fibroblasts from parkin-mutant early-onset Parkinson's disease patients.

    Science.gov (United States)

    Lippolis, Rosa; Siciliano, Rosa Anna; Pacelli, Consiglia; Ferretta, Anna; Mazzeo, Maria Fiorella; Scacco, Salvatore; Papa, Francesco; Gaballo, Antonio; Dell'Aquila, Claudia; De Mari, Michele; Papa, Sergio; Cocco, Tiziana

    2015-09-01

    Parkinson's disease (PD) is the most common neurodegenerative movement disorder caused primarily by selective degeneration of the dopaminergic neurons in substantia nigra. In this work the proteomes extracted from primary fibroblasts of two unrelated, hereditary cases of PD patients, with different parkin mutations, were compared with the proteomes extracted from commercial adult normal human dermal fibroblasts (NHDF) and primary fibroblasts from the healthy mother of one of the two patients. The results show that the fibroblasts from the two different cases of parkin-mutant patients display analogous alterations in the expression level of proteins involved in different cellular functions, like cytoskeleton structure-dynamics, calcium homeostasis, oxidative stress response, protein and RNA processing. PMID:26096686

  8. Establishment of immortal normal and ataxia telangiectasia fibroblast cell lines by introduction of the hTERT gene

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, Hideaki; Fukami, Hiroko; Hayashi, Yuko; Kiyono, Tohru; Ishizaki, Kanji [Aichi Cancer Center, Nagoya (Japan). Research Inst.; Nakatsugawa, Shigekazu; Hamaguchi, Michinari [Nagoya Univ. (Japan). School of Medicine

    2002-06-01

    To establish immortal human cells, we introduced the human catalytic subunit of telomerase (hTERT) gene into skin fibroblast cells obtained from normal and ataxia telangiectasia (AT) individuals of Japanese origin. After hTERT introduction, these cells continue to grow beyond a population doubling number of 200 while maintaining their original radiosensitivity. Inductions of p53, phosphorylation of Serl5 in p53, and induction of p21 by X-ray irradiation in immortal cells derived from normal individual were not affected by the hTERT introduction. Both normal and AT immortal cells exhibited an apparent inhibition of growth as original primary cells when they reached confluence. Karyotype analysis has revealed that they are in a diploid range. These results suggest that cells immortalized by hTERT introduction retain their original characteristics except for immortalization, and that they may be useful for analyzing various effects of radiation on human cells. (author)

  9. Low-dose radiation activates Nrf1/2 through reactive species and the Ca2+/ERK1/2 signaling pathway in human skin fibroblast cells

    OpenAIRE

    Eun Kyeong Lee; Jin-Ah Kim; Seong Joon Park; Jeung Ki Kim; Kyu Heo; Kwang Mo Yang; Tae Gen Son

    2013-01-01

    In the current study, we explored the effect of LDR on the activation of Nrfs transcription factor involved in cellular redox events. Experiments were carried out utilizing 0.05 and 0.5 Gy X-ray irradiated normal human skin fibroblast HS27 cells. The results showed LDR induced Nrf1 and Nrf2 activation and expression of antioxidant genes HO-1, Mn-SOD, and NQO1. In particular, 0.05 Gy-irradiation increased only Nrf1 activation, but 0.5 Gy induced both Nrf1 and Nrf2 activation. LDR-mediated Nrf1...

  10. DEF-1, a Novel Src SH3 Binding Protein That Promotes Adipogenesis in Fibroblastic Cell Lines

    OpenAIRE

    King, Frederick J.; Hu, Erding; Harris, David F.; Sarraf, Pasha; Spiegelman, Bruce M.; Roberts, Thomas M.

    1999-01-01

    The Src homology 3 (SH3) motif is found in numerous signal transduction proteins involved in cellular growth and differentiation. We have purified and cloned a novel protein, DEF-1 (differentiation-enhancing factor), from bovine brain by using a Src SH3 affinity column. Ectopic expression of DEF-1 in fibroblasts resulted in the differentiation of a significant fraction of the culture into adipocytes. This phenotype appears to be related to the induction of the transcription factor peroxisome ...

  11. Studies of DNA and chromosome damage in skin fibroblasts and blood lymphocytes from psoriasis patients treated with 8-methoxypsoralen and UVA irradiation

    International Nuclear Information System (INIS)

    Exposure of human lymphocytes and skin fibroblasts in vitro to a single, clinically used dose of PUVA, i.e., 0.1 micrograms/ml of 8-methoxypsoralen (8-MOP) plus 0.9-4 J/cm2 of longwave ultraviolet radiation (UVA), lead to the formation of DNA damage as determined by alkaline elution, and to chromosome aberrations and sister chromatid exchanges (SCE). When lymphocyte-enriched plasma was obtained from psoriasis patients 2 h after oral intake of 8-MOP and then UVA irradiated (1.8-3.6 J/cm2) in vitro, an increased frequency of chromosome aberrations and SCE was observed. Normal levels of chromosome aberrations and SCE were found in lymphocytes of psoriasis patients after 3-30 weeks of PUVA treatment in vivo. A small but statistically significant increase in the SCE frequency was observed in the lymphocytes of psoriasis patients treated for 1-6 years with PUVA (mean 18.0 SCE/cell) as compared with before PUVA (mean 15.8, p less than 0.05). Skin fibroblasts of psoriasis patients analyzed 5 years after the start of PUVA treatment showed a normal number of SCE but a high fraction of filter-retained DNA in the alkaline elution assay, suggesting the presence of cross-linked DNA

  12. Mécano-Stimulation™ of the skin improves sagging score and induces beneficial functional modification of the fibroblasts: clinical, biological, and histological evaluations

    Directory of Open Access Journals (Sweden)

    Humbert P

    2015-02-01

    Full Text Available Philippe Humbert,1,2 Ferial Fanian,1,2 Thomas Lihoreau,1,2 Adeline Jeudy,1,2 Ahmed Elkhyat,1,2 Sophie Robin,3 Carol Courderot-Masuyer,3 Hélène Tauzin,3 Christine Lafforgue,1,2,4 Marek Haftek5 1Research and Studies Center on the Integument (CERT, Department of Dermatology, Clinical Investigation Center (CIC 1431, Besançon University Hospital; 2INSERM UMR1098, FED4234 IBCT, University of Franche-Comté, Besançon, France; 3SARL BIOEXIGENCE, Besançon, France; 4Dermopharmacology and Cosmetology Unit, University of Paris Sud, France; 5University of Lyon 1, EA4169, Experimental, clinical and therapeutic aspects of the skin barrier function, INSERM US7 – CNRS UMS3453, Lyon, France Background: Loss of mechanical tension appears to be the major factor underlying decreased collagen synthesis in aged skin. Numerous in vitro studies have shown the impact of mechanical forces on fibroblasts through mechanotransduction, which consists of the conversion of mechanical signals to biochemical responses. Such responses are characterized by the modulation of gene expression coding not only for extracellular matrix components (collagens, elastin, etc. but also for degradation enzymes (matrix metalloproteinases [MMPs] and their inhibitors (tissue inhibitors of metalloproteinases [TIMPs]. A new device providing a mechanical stimulation of the cutaneous and subcutaneous tissue has been used in a simple, blinded, controlled, and randomized study. Materials and methods: Thirty subjects (aged between 35 years and 50 years, with clinical signs of skin sagging, were randomly assigned to have a treatment on hemiface. After a total of 24 sessions with Mécano-Stimulation™, biopsies were performed on the treated side and control area for in vitro analysis (dosage of hyaluronic acid, elastin, type I collagen, MMP9; equivalent dermis retraction; GlaSbox®; n=10 and electron microscopy (n=10. Furthermore, before and after the treatment, clinical evaluations and self-assessment questionnaire were done.Results: In vitro analysis showed increases in hyaluronic acid, elastin, type I collagen, and MMP9 content along with an improvement of the migratory capacity of the fibroblasts on the treated side. Electron microscopy evaluations showed a clear dermal remodeling in relation with the activation of fibroblast activity. A significant improvement of different clinical signs associated with skin aging and the satisfaction of the subjects were observed, correlated with an improvement of the sagging cheek.Conclusion: Mécano-Stimulation is a noninvasive and safe technique delivered by flaps microbeats at various frequencies, which can significantly improve the skin trophicity. Results observed with objective measurements, ie, in vitro assessments and electron microscopy, confirm the firming and restructuring effect clinically observed. Keywords: skin rejuvenation, skin sagging, mechanical stimulation, fibroblast synthesis

  13. Production of a cloned calf from a fetal fibroblast cell line

    Directory of Open Access Journals (Sweden)

    Mello M.R.B.

    2003-01-01

    Full Text Available The present study examined the in vitro and in vivo development of bovine nuclear-transferred embryos. A bovine fetal fibroblast culture was established and used as nucleus donor. Slaughterhouse oocytes were matured in vitro for 18 h before enucleation. Enucleated oocytes were fused with fetal fibroblasts with an electric stimulus and treated with cytochalasin D and cycloheximide for 1 h followed by cycloheximide alone for 4 h. Reconstructed embryos were cultured for 7-9 days and those which developed to blastocysts were transferred to recipient cows. Of 191 enucleated oocytes, 83 (43.5% were successfully fused and 24 (28.9% developed to blastocysts. Eighteen freshly cloned blastocysts were transferred to 14 recipients, 5 (27.8% of which were pregnant on day 35 and 3 (16.7% on day 90. Of the three cows that reached the third trimester, one recipient died of hydrallantois 2 months before term, one aborted fetus was recovered at 8 months of gestation, and one delivered by cesarian section a healthy cloned calf. Today, the cloned calf is 15 months old and presents normal body development (378 kg and sexual behavior (libido and semen characteristics.

  14. Ex-vivo transduced autologous skin fibroblasts expressing human Lim Mineralization Protein-3 efficiently form new bone in animal models

    Science.gov (United States)

    Lattanzi, Wanda; Parrilla, Claudio; Fetoni, Annarita; Logroscino, Giandomenico; Straface, Giuseppe; Pecorini, Giovanni; Stigliano, Egidio; Tampieri, Anna; Bedini, Rossella; Pecci, Raffaella; Michetti, Fabrizio; Gambotto, Andrea; Robbins, Paul D.; Pola, Enrico

    2012-01-01

    Local gene transfer of the human LIM Mineralization Protein (LMP), a novel intracellular positive regulator of the osteoblast differentiation program, can induce efficient bone formation in rodents. In order to develop a clinically relevant gene therapy approach to facilitate bone healing, we have used primary dermal fibroblasts transduced ex vivo with Ad.LMP3 and seeded on an hydroxyapatite/collagen matrix prior to autologous implantation. Here we demonstrate that genetically modified autologous dermal fibroblasts expressing Ad.LMP-3 are able to induce ectopic bone formation following implantation of the matrix into the mouse triceps and paravertebral muscles. Moreover, implantation of the Ad.LMP-3-modified dermal fibroblasts into a rat mandibular bone critical size defect model results in efficient healing as determined by X-ray, histology and three dimensional micro computed tomography (3D?CT). These results demonstrate the effectiveness of the non-secreted intracellular osteogenic factor LMP-3, in inducing bone formation in vivo. Moreover, the utilization of autologous dermal fibroblasts implanted on a biomaterial represents a promising approach for possible future clinical applications aimed at inducing new bone formation. PMID:18633445

  15. Adaptive response to ionizing radiation in normal human skin fibroblasts. Enhancement of DNA repair rate and modulation of gene expression

    International Nuclear Information System (INIS)

    Low doses and dose rates of ionizing radiation enhance the rate of DNA repair in human fibroblasts and protect the cells against radiation-induced micronucleus formation. Chronic exposures reduce the mRNA levels of the genes topoisomerase II and FACC-1 (Fanconi's anemia, group C). (authors). 11 refs., 1 tab., 2 figs

  16. HPV-positive HNSCC cell lines but not primary human fibroblasts are radiosensitized by the inhibition of Chk1

    International Nuclear Information System (INIS)

    Purpose: Despite the comparably high cure rates observed for HPV-positive HNSCC, there is still a great need for specific tumor radiosensitization due to the often severe side effects resulting from intense radiochemotherapy. We recently demonstrated that HPV-positive HNSCC cell lines are characterized by a defect in DNA double-strand break repair associated with a pronounced G2-arrest. Here we tested whether abrogation of this radiation-induced G2-arrest by the inhibition of Chk1 results in specific radiosensitization of HPV-positive HNSCC cells. Materials and methods: Experiments were performed with five HPV and p16-positive (93-VU-147T, UM-SCC-47, UT-SCC-45, UD-SCC-2, UPCI-SCC-154) and two HPV and p16-negative HNSCC cell lines, as well as two normal human fibroblast strains. Chk1 was inhibited by the selective inhibitor PF-00477736. Cell cycle distribution was determined by flow cytometry, Chk1-activity via Western blot and cell survival by colony formation assay. Results: With the exception of UPCI-SCC-154, the inhibition of Chk1 was found to abolish the pronounced radiation-induced G2-arrest in all HPV-positive cells utilized. All tumor cell lines that demonstrated the abrogation of G2-arrest also demonstrated radiosensitization. Notably, in G1-arrest-proficient normal human fibroblasts no radiosensitization was induced. Conclusion: Abrogation of the G2 checkpoint through the inhibition of Chk1 may be used to selectively increase the cellular radiosensitivity of HPV-positive HNSCC without affecting the surrounding normal tissue

  17. Effects of PUVA on a human skin epithelial cell line

    International Nuclear Information System (INIS)

    An established human epithelial cell line was exposed to photoactivated 8-methoxy psoralen (PUVA) during exponential growth. Effects of PUVA treatment on cell growth were measured by cell kinetic methods (counting of cell numbers, flow cytometric measurements (FCM) of DNA and calculations of labelling indices (LI)). Doses of 8-methoxy psoralen and UVA were comparable to those used in patients. The cell number in PUVA treated cultures remained almost constant, and very few mitoses were seen for 144 h. About 9 h after PUVA, both FCM and LI showed an increase in the fraction of cells in S-phase, reaching a maximum of 85-90% after 24 h. DNA synthesis took place at a low rate in these cells. FCM showed an increasing fraction of polyploid cells after PUVA treatment. The possibility that inhibition of cell proliferation is one of the main effects of PUVA, is discussed. (orig.)

  18. Collagen expression in fibroblasts with a novel LMNA mutation

    International Nuclear Information System (INIS)

    Laminopathies are a group of genetic disorders caused by LMNA mutations; they include muscular dystrophies, lipodystrophies, and progeroid syndromes. We identified a novel heterozygous LMNA mutation, L59R, in a patient with the general appearance of mandibuloacral dysplasia and progeroid features. Examination of the nuclei of dermal fibroblasts revealed the irregular morphology characteristic of LMNA mutant cells. The nuclear morphological abnormalities of LMNA mutant lymphoblastoid cell lines were less prominent compared to those of primary fibroblasts. Since it has been reported that progeroid features are associated with increased extracellular matrix in dermal tissues, we compared a subset of these components in fibroblast cultures from LMNA mutants with those of control fibroblasts. There was no evidence of intracellular accumulation or altered mobility of collagen chains, or altered conversion of procollagen to collagen, suggesting that skin fibroblast-mediated matrix production may not play a significant role in the pathogenesis of this particular laminopathy

  19. Evidence for the existence of hypothetical proteins in human bronchial epithelial, fibroblast, amnion, lymphocyte, mesothelial and kidney cell lines.

    Science.gov (United States)

    Oh, J E; Krapfenbauer, K; Fountoulakis, M; Frischer, Th; Lubec, G

    2004-02-01

    The human genome maybe limited to about 30000 genes whereas the human proteome may be represented by a rough estimate of one million proteins. A legion of proteins have been described and information about these structures are readily available in data banks. There remains, however, a large series of unknown or hypothetical proteins (HPs). Many of them have been predicted from nucleic acid sequences only and are therefore named predicted or HPs. Carrying out "protein hunting" by generating large maps of human cell lines, we aimed to find and identify HPs and provide an analytical tool thereof. Cell lysates from human bronchial epithelial, fibroblast, amnion, lymphocyte, mesothelial and kidney cell lines were prepared and proteins run on two-dimensional gel-electrophoresis (2DE) with in-gel digestion and mass spectrometrical analysis using the MALDI-TOF principle.16 HPs were found in these cell lines and some show cell-specific expressional patterns. HPs belong to several protein classes including structural, signaling, transcriptional/translational, chaperone-related and others. We furthermore provide analytical data i.e. pIs that were often different from predicted values in data banks.A list of HPs has been shown to really exist in several human cell lines thus contributing to knowledge on protein machineries and cascades. Observed and predicted pI values are given representing an analytical tool along with unambiguous identification of protein spots by mass spectrometry independent of antibody availability and specificity thus complementing established methods. PMID:14752611

  20. Antiageing Mechanisms of a Standardized Supercritical CO2 Preparation of Black Jack (Bidens pilosa L.) in Human Fibroblasts and Skin Fragments

    Science.gov (United States)

    Dieamant, Gustavo; Pereda, Maria Del Carmen V.; Nogueira, Cecília; Eberlin, Samara; Facchini, Gustavo; Checon, Juliana Tibério; Cesar, Camila Kappke; Mussi, Lilian; Polezel, Márcio Antonio; Martins-Oliveira, Divino; Di Stasi, Luiz Claudio

    2015-01-01

    The use of topical retinoids to treat skin disorders and ageing can induce local reactions, while oral retinoids are potent teratogens and produce several unwanted effects. This way, efforts to explore complementary care resources should be supported. Based on this, we evaluate the antiageing effects of a supercritical CO2 extract from Bidens pilosa L. (BPE-CO2A) containing a standardized multicomponent mixture of phytol, linolenic, palmitic, linoleic, and oleic acids. BPE-CO2A was assessed for its effects on human dermal fibroblasts (TGF-β1 and FGF levels using ELISA; collagen, elastin, and glycosaminoglycan by colorimetric assays, and mRNA expression of RXR, RAR, and EGFr by qRT-PCR) and human skin fragments (RAR, RXR, collagen, elastin, and glycosaminoglycan by immunohistochemical analysis). Levels of extracellular matrix elements, TGF-β1 and FGF, and EGFr gene expression were significantly increased by BPE-CO2A. The modulation of RXR and RAR was positively demonstrated after the treatment with BPE-CO2A or phytol, a component of BPE-CO2A. The effects produced by BPE-CO2A were similar to or better than those produced by retinol and retinoic acid. The ability to stimulate extracellular matrix elements, increase growth factors, and modulate retinoid and rexinoid receptors provides a basis for the development of preparation containing BPE-CO2A as an antiageing/skin-repair agent. PMID:25883669

  1. Antiageing Mechanisms of a Standardized Supercritical CO 2 Preparation of Black Jack (Bidens pilosa L.) in Human Fibroblasts and Skin Fragments.

    Science.gov (United States)

    Dieamant G; Pereda Mdel C; Nogueira C; Eberlin S; Facchini G; Checon JT; Cesar CK; Mussi L; Polezel MA; Martins-Oliveira D Jr; Di Stasi LC

    2015-01-01

    The use of topical retinoids to treat skin disorders and ageing can induce local reactions, while oral retinoids are potent teratogens and produce several unwanted effects. This way, efforts to explore complementary care resources should be supported. Based on this, we evaluate the antiageing effects of a supercritical CO2 extract from Bidens pilosa L. (BPE-CO2A) containing a standardized multicomponent mixture of phytol, linolenic, palmitic, linoleic, and oleic acids. BPE-CO2A was assessed for its effects on human dermal fibroblasts (TGF-?1 and FGF levels using ELISA; collagen, elastin, and glycosaminoglycan by colorimetric assays, and mRNA expression of RXR, RAR, and EGFr by qRT-PCR) and human skin fragments (RAR, RXR, collagen, elastin, and glycosaminoglycan by immunohistochemical analysis). Levels of extracellular matrix elements, TGF-?1 and FGF, and EGFr gene expression were significantly increased by BPE-CO2A. The modulation of RXR and RAR was positively demonstrated after the treatment with BPE-CO2A or phytol, a component of BPE-CO2A. The effects produced by BPE-CO2A were similar to or better than those produced by retinol and retinoic acid. The ability to stimulate extracellular matrix elements, increase growth factors, and modulate retinoid and rexinoid receptors provides a basis for the development of preparation containing BPE-CO2A as an antiageing/skin-repair agent.

  2. Quercetin enhances UVA-induced DNA damage in a rat fibroblast cell line

    OpenAIRE

    De Castro, M. F.; Basto, Diana; Silva, João P.; Coutinho, O P

    2007-01-01

    Ultraviolet A (UVA) radiation from sunlight induces the production of reactive oxygen species (ROS), affecting a variety of cellular targets including the DNA. Quercetin, a flavonol present in many fruits, vegetables and beverages has been reported as a powerful antioxidant with an important role in prevention of carcinogenesis. The use of this compound, in topical formulations, could be of benefit in the prevention of skin damage produced by sunlight exposure. We investigated the effec...

  3. Decreased mitochondrial density and ultrastructural changes of mitochondria in cultivated skin fibroblasts of patients with Huntington´s disease

    Czech Academy of Sciences Publication Activity Database

    Rodinová, M.; Marková, M.; Kratochvílová, H.; Kučerová, I.; Tesařová, M.; Lišková, Irena; Klempíř, J.; Roth, J.; Zeman, J.; Hansíková, H.

    2015-01-01

    Roč. 78, Suppl 2 (2015), s. 20-21. ISSN 1210-7859. [Conference on Animal Models for neurodegenerative Diseases /3./. 08.11.2015-10.11.2015, Liblice] R&D Projects: GA MŠk ED2.1.00/03.0124; GA MŠk(CZ) 7F14308 Institutional support: RVO:67985904 Keywords : Huntington´s disease * fibroblasts * mitochondrial ultrastructure Subject RIV: FH - Neurology

  4. Ex-vivo transduced autologous skin fibroblasts expressing human Lim Mineralization Protein-3 efficiently form new bone in animal models

    OpenAIRE

    Lattanzi, Wanda; Parrilla, Claudio; Fetoni, Annarita; Logroscino, Giandomenico; Straface, Giuseppe; Pecorini, Giovanni; Stigliano, Egidio; Tampieri, Anna; Bedini, Rossella; Pecci, Raffaella; Michetti, Fabrizio; Gambotto, Andrea; Robbins, Paul D.; Pola, Enrico

    2008-01-01

    Local gene transfer of the human LIM Mineralization Protein (LMP), a novel intracellular positive regulator of the osteoblast differentiation program, can induce efficient bone formation in rodents. In order to develop a clinically relevant gene therapy approach to facilitate bone healing, we have used primary dermal fibroblasts transduced ex vivo with Ad.LMP3 and seeded on an hydroxyapatite/collagen matrix prior to autologous implantation. Here we demonstrate that genetically modified autolo...

  5. In vitro study of RRS HA injectable mesotherapy/biorevitalization product on human skin fibroblasts and its clinical utilization

    OpenAIRE

    Deglesne PA; Arroyo R; Ranneva E; Deprez P

    2016-01-01

    Pierre-Antoine Deglesne,* Rodrigo Arroyo,* Evgeniya Ranneva, Philippe Deprez Research and Development, SKIN TECH PHARMA GROUP, Castelló d'Empúries, Spain  *These authors contributed equally to this work Abstract: Mesotherapy/biorevitalization with hyaluronic acid (HA) is a treatment approach currently used for skin rejuvenation. Various products with a wide range of polycomponent formulations are available on the market. Most of these formulations contain noncross-...

  6. Evidence that novobiocin and nalidixic acid do not inhibit excision repair in u.v.-irradiated human skin fibroblasts at a pre-incision

    International Nuclear Information System (INIS)

    The effects of novobiocin and nalidixic acid on the specific toxicity of aphidicolin towards u.v. irradiated arrested human skin fibroblasts have been determined. Contrary to the result expected if either drug were causing inhibition of excision repair at a pre-incision step the sector of toxicity due to a combined treatment of 300 ?g ml-1 nalidixic acid and 1.0 ?g ml-1 aphidicolin is unchanged when compared with that due to treatment with 1.0 ?g ml-1 aphidicolin alone, while that for 150 ?g ml-1 novobiocin + 1.0 ?g ml-1 aphidicolin was slightly increased. In parallel measurements of the inhibition of u.v.-induced DNA repair synthesis in arrested fibroblasts by these drugs, 150 ?g ml-1 novobiocin inhibited repair synthesis by approx.60% over the fluence range employed. Nalidixic acid (300 ?g ml-1) caused no detectable inhibition of repair synthesis. It was concluded that the mode of action of novobiocin in the inhibition of DNA excision repair is not via the inhibition of a pre-incision step and the data do not support the hypothesis that a type II topoisomerase mediated change in DNA supercoiling is an essential early step in excision repair of u.v.-induced damage. (author)

  7. Evidence that novobiocin and nalidixic acid do not inhibit excision repair in u.v.-irradiated human skin fibroblasts at a pre-incision step

    International Nuclear Information System (INIS)

    The effects of novobiocin and nalidixic acid on the specific toxicity of aphidicolin towards u.v. irradiated arrested (nondividing) human skin fibroblasts have been determined. Contrary to the result expected if either drug were causing inhibition of excision repair at a pre-incision step the sector of toxicity due to a combined treatment of 300 micrograms ml-1 nalidixic acid and 1.0 micrograms ml-1 aphidicolin is unchanged when compared with that due to treatment with 1.0 micrograms ml-1 aphidicolin alone, while that for 150 micrograms ml-1 novobiocin + 1.0 micrograms ml-1 aphidicolin was slightly increased. In parallel measurements of the inhibition of u.v.-induced DNA repair synthesis in arrested fibroblasts by these drugs, 150 micrograms ml-1 novobiocin inhibited repair synthesis by approximately 60% over the fluence range employed. Nalidixic acid at a concentration of 300 micrograms ml-1 caused no detectable inhibition of repair synthesis. The authors conclude that the mode of action of novobiocin in the inhibition of DNA excision repair is not via the inhibition of a pre-incision step and the data do not support the hypothesis that a type II topoisomerase mediated change in DNA supercoiling is an essential early step in excision repair of u.v.-induced damage

  8. Microflora of the penile skin-lined neovagina of transsexual women

    OpenAIRE

    Claeys Geert; De Backer Ellen; Saerens Bart; dos Santos Lopes Santiago Guido; Monstrey Stan; Gerris Jan; Verstraelen Hans; Weyers Steven; Vaneechoutte Mario; Verhelst Rita

    2009-01-01

    Abstract Background The microflora of the penile skin-lined neovagina in male-to-female transsexuals is a recently created microbial niche which thus far has been characterized only to a very limited extent. Yet the knowledge of this microflora can be considered as essential to the follow-up of transsexual women. The primary objective of this study was to map the neo-vaginal microflora in a group of 50 transsexual women for whom a neovagina was constructed by means of the inverted penile skin...

  9. Secretion of cyclic GMP by cultured epithelial and fibroblast cell lines in response to nitric oxide.

    Science.gov (United States)

    Patel, M J; Wypij, D M; Rose, D A; Rimele, T J; Wiseman, J S

    1995-04-01

    LLC-PK1 epithelial cells and RFL-6 fibroblasts secreted both cyclic AMP (cAMP) and cyclic GMP (cGMP) when costimulated with forskolin and 3-morpholinosydnonimine (a chemical nitric oxide generator). Intracellular cAMP levels as high as 1100 and 12,000 pmol/10(6) cells were achieved for the two cell types, respectively. These levels were high enough to reach approximately 50% saturation of the cAMP transporter and inhibited transport of cGMP to an equal extent, suggesting that the two cyclic nucleotides compete for a common transport system. The rates of secretion of cGMP and cAMP from LLC-PK1 cells increased in proportion to their rates of synthesis as concentrations of stimulant were varied, but increased only 25% relative to intracellular concentrations in response to inhibition of phosphodiesterases by 3-isobutylmethylxanthine. It is proposed that secretion of cyclic nucleotides is not simply proportional to the total intracellular pool in these cells, but rather is coupled to synthesis. In support of this model, oxyhemoglobin was used to trap nitric oxide and block activity of guanylate cyclase in cells treated with 3-morpholinosydnonimine. As a result, secretion of cGMP ceased within 1 min, whereas intracellular levels decreased slowly over 60 min. Probenecid [p-(dipropylsulfamoyl)benzoic acid] is a nonselective antagonist of anion transport that inhibited secretion of cAMP in both cell types but, unexpectedly, blocked synthesis of cGMP, and this was reflected in direct inhibition of soluble guanylate cyclase in cell lysates. Two heat-stable, high molecular weight factors that confer sensitivity to probenecid were identified, and these factors increased the sensitivity of guanylate cyclase to nitric acid by an order of magnitude. PMID:7536242

  10. Interaction between Leptospiral Lipopolysaccharide and Toll-like Receptor 2 in Pig Fibroblast Cell Line, and Inhibitory Effect of Antibody against Leptospiral Lipopolysaccharide on Interaction

    OpenAIRE

    Guo, Yijie; Fukuda, Tomokazu; Nakamura, Shuichi; Bai, Lanlan; Xu, Jun; Kuroda, Kengo; Tomioka, Rintaro; Yoneyama, Hiroshi(Department, of, Physics,, Saga, University,, Saga, 840-8502,, Japan); Isogai, Emiko

    2015-01-01

    Leptospiral lipopolysaccharide (L-LPS) has shown potency in activating toll-like receptor 2 (TLR2) in pig fibroblasts (PEFs_NCC1), and causes the expression of proinflammatory cytokines. However, the stimulation by L-LPS was weak eliciting the function of TLR2 sufficiently in pig innate immunity responses during Leptospira infection. In this study, the immune response of pig embryonic fibroblast cell line (PEFs_SV40) was investigated and was found to be the high immune response, thus TLR2 is ...

  11. Replacement of primary chicken embryonic fibroblasts (CEF) by the DF-1 cell line for detection of avian leucosis viruses.

    Science.gov (United States)

    Maas, Riks; van Zoelen, Diana; Oei, Hok; Claassen, Ivo

    2006-09-01

    International regulations prescribe that the absence of avian leucosis viruses (ALV) in avian live virus vaccines has to be demonstrated. Primary chicken embryo fibroblasts (CEF) from special SPF chicken lines are normally used for detection of ALV. The suitability of the DF-1 cell line for ALV-detection, as alternative for primary CEF, was studied in three types of experiments: (1) in titration experiments without cell passage, (2) in experiments with passages in cell cultures according to European Pharmacopoeia requirements, and (3) in experiments with commercial live avian vaccines that had been spiked with known amounts of ALV. In all tests the sensitivity of ALV-A and ALV-J detections on DF-1 cells was at least as high as on primary CEF. The sensitivity of ALV-B detection was always superior when DF-1 cells were used. ALV were detected earlier in all comparative tests when DF-1 cells were used. ALV-A, ALV-B and ALV-J all induced CPE on DF-1 cells, whereas no clear CPE was seen on CEF-cells. For reasons of sensitivity, standardisation as well as reduction of animal use, the data support the use of DF-1 cells to monitor absence of ALV in vaccine virus seed lots or finished products. PMID:16257542

  12. Characterization of a novel fibroblast-like cell line from rainbow trout and responses to sublethal anoxia

    DEFF Research Database (Denmark)

    Ossum, Carlo Gunnar; Hoffmann, Else Kay; Vijayan, M.M.; Holt, S.E.; Bols, N.C.

    2004-01-01

    A novel fibroblast-like cell line RTHDF was established from hypodermal connective tissue of rainbow trout Oncorhynchus mykiss and telomerase activity was demonstrated early and late in cell line development. When RTHDF cells were exposed to bioenergetic stress, i.e. anoxia, activation of the...... stress activated member of the mitogen-activated protein kinase family, p38MAPK and induction of heat shock protein (Hsp70) were evident. The time-course of the p38MAPK activation and the induction of Hsp70 expression in RTHDF were studied in response to chemically induced anoxia. p38MAPK was activated...... rapidly, with maximal activity after 10 min of anoxia. Hsp70 was induced after 30 min of anoxia, followed by overnight recovery in growth medium at 21° C. Using the p38MAPK-specific inhibitor SB203580, the enhanced expression of Hsp70 occurred independently of p38MAPK activation in RTHDF. These data...

  13. Growth and Replication of Infectious Bursal Disease Virus in the DF-1 Cell Line and Chicken Embryo Fibroblasts

    Science.gov (United States)

    Rekha, Kaliyaperumal; Sivasubramanian, Chandran; Chung, Ill-Min; Thiruvengadam, Muthu

    2014-01-01

    Infectious bursal disease virus (IBDV) causes a highly contagious disease in young chicks and leads to significant economic losses in the poultry industry. To determine a suitable cell line for IBDV infection, replication, and growth kinetics of the virus, DF-1 cells and chicken embryo fibroblasts (CEF) were used. The population doubling per day (Pd/D) was found to be higher in DF-1 as compared to CEF cells. A suitable time of infection (TOI) was established for increased production of virus and greater infectivity titers. The DF-1 and CEF cells were found to be susceptible to infection by producing marked cytopathic effects (CPEs), and the growth curves of IBDV in DF-1 and CEF cells were evaluated by infectivity assay using tissue culture infectious dose (TCID50). The cytopathic effects of the virus in DF-1 and CEF cells were found to be similar, but higher viral titers were detected in the DF-1 cells as compared to CEF. Thus the DF-1 cell line had a higher growth potential and infectivity, which will be of advantage in vaccine production. PMID:24949455

  14. Control by fibroblast growth factor of differentiation in the BC3H1 muscle cell line

    OpenAIRE

    1985-01-01

    The regulation of creatine phosphokinase (CPK) expression by polypeptide growth factors has been examined in the clonal mouse muscle BC3H1 cell line. After arrest of cell growth by exposure to low concentrations of serum, BC3H1 cells accumulate high levels of muscle- specific proteins including CPK. The induction of this enzyme is reversible in the presence of high concentrations of fetal calf serum, which cause quiescent, differentiated cells to reenter the cell cycle. Under these conditions...

  15. Performance of a novel keratinocyte-based reporter cell line to screen skin sensitizers in vitro

    International Nuclear Information System (INIS)

    In vitro tests are needed to replace animal tests to screen for the skin sensitization potential of chemicals. Skin sensitizers are electrophilic molecules and the Nrf2-electrophile-sensing pathway comprising the repressor protein Keap1, the transcription factor Nrf2 and the antioxidant response element (ARE) is emerging as a toxicity pathway induced by skin sensitizers. Previously, we screened a large set of chemicals in the reporter cell line AREc32, which contains an eight-fold repeat of the rat GSTA2 ARE-sequence upstream of a luciferase reporter gene in the human breast cancer cell line MCF7. This approach was now further developed to bring it closer to the conditions in the human skin and to propose a fully standardized assay. To this end, a luciferase reporter gene under control of a single copy of the ARE-element of the human AKR1C2 gene was stably inserted into HaCaT keratinocytes. A standard operating procedure was developed whereby chemicals are routinely tested at 12 concentrations in triplicate for significant induction of gene activity. We report results from this novel assay on (i) a list of reference chemicals published by ECVAM, (ii) the ICCVAM list of chemicals for validation of alternative endpoints in the LLNA and (iii) on a more general list of 67 chemicals derived from the ICCVAM database. For comparison, peptide reactivity data are presented for the same chemicals. The results indicate a good predictive value of this approach for hazard identification. Its technical simplicity, the high-throughput format and the good predictivity may make this assay a candidate for rapid validation to meet the tight deadline to replace animal tests for skin sensitization by 2013 set by the European authorities.

  16. Metabolism of [1-(14)C] and [2-(14)C] leucine in cultured skin fibroblasts from patients with isovaleric acidemia. Characterization of metabolic defects.

    Science.gov (United States)

    Tanaka, K; Mandell, R; Shih, V E

    1976-01-01

    Leucine metabolism in cultured skin fibroblasts from patients with isovaleric acidemia was compared with that in normal fibroblasts and in cells from patients with maple syrup urine disease using [1-(14)C] and [2-(14)C] leucine as substrates. Inhibitory effects of methylenecyclopropylacetic acid on leucine metabolism in normal cells were also investigated. Production of 14CO2 from [2-(14)C] leucine was very reduced (96-99%) in both types of mutant cells. Radioactive isovaleric acid accumulated in assay media with isovaleric acidemia cells but not in those with maple syrup urine disease cells. Unexpectedly, 14CO2 production from [1-(14)C] leucine was partially depressed (80%) in isovaleric acidemia cells whereas in maple syrup urine disease cells it was strongly depressed (99%) as expected. These two mutant cells were clearly distinguished by detection of 14C-isovaleric acid accumulation after incubation with [2-(14)C] leucine. A pattern of inhibition of leucine oxidation similar to that seen in isovaleric acidemia cells was induced in normal cells by the addition of 0.7 mM methylenecyclopropylacetic acid to the assay medium. The partial inhibition of [1-(14)C] leucine oxidation seen in isovaleric acidemia cells and also in normal cells in the presence of the inhibitor appears to be, at least in part, due to an accumulation of isovalerate in the cells. Isovaleric acid (5-10) mM) inhibited [1-(14)C] leucine oxidation 32-68% when added to the assay medium with normal cells. Addition of flavin adenine dinucleoside to culture medium or assay medium or both did not restore oxidation of either leucine substrate in isovaleric acidemia cells. PMID:932204

  17. Antagonism of phenanthrene cytotoxicity for human embryo lung fibroblast cell line HFL-I by green tea polyphenols

    Energy Technology Data Exchange (ETDEWEB)

    Mei Xin [Department of Tea Science, Zhejiang University, Hangzhou 310029 (China); Key Laboratory of Horticultural Plant Growth Development and Biotechnology of Ministry of Agriculture, Zhejiang University, Hangzhou 310029 (China); Wu Yuanyuan; Mao Xiao [Department of Tea Science, Zhejiang University, Hangzhou 310029 (China); Tu Youying, E-mail: youytu@zju.edu.c [Department of Tea Science, Zhejiang University, Hangzhou 310029 (China)

    2011-01-15

    Polycyclic aromatic hydrocarbons (PAHs) have been detected in some commercial teas around the world and pose a threat to tea consumers. However, green tea polyphenols (GTP) possess remarkable antioxidant and anticancer effects. In this study, the potential of GTP to block the toxicity of the model PAH phenanthrene was examined in human embryo lung fibroblast cell line HFL-I. Both GTP and phenanthrene treatment individually caused dose-dependent inhibition of cell growth. A full factorial design experiment demonstrated that the interaction of phenanthrene and GTP significantly reduced growth inhibition. Using the median effect method showed that phenanthrene and GTP were antagonistic when the inhibitory levels were less than about 50%. Apoptosis and cell cycle detection suggested that only phenanthrene affected cell cycle significantly and caused cell death; GTP lowered the mortality of HFL-I cells exposed to phenanthrene; However, GTP did not affect modulation of the cell cycle by phenanthrene. - Green tea polyphenols antagonised cytotoxicity of a low-ring PAH phenanthrene.

  18. Antagonism of phenanthrene cytotoxicity for human embryo lung fibroblast cell line HFL-I by green tea polyphenols

    International Nuclear Information System (INIS)

    Polycyclic aromatic hydrocarbons (PAHs) have been detected in some commercial teas around the world and pose a threat to tea consumers. However, green tea polyphenols (GTP) possess remarkable antioxidant and anticancer effects. In this study, the potential of GTP to block the toxicity of the model PAH phenanthrene was examined in human embryo lung fibroblast cell line HFL-I. Both GTP and phenanthrene treatment individually caused dose-dependent inhibition of cell growth. A full factorial design experiment demonstrated that the interaction of phenanthrene and GTP significantly reduced growth inhibition. Using the median effect method showed that phenanthrene and GTP were antagonistic when the inhibitory levels were less than about 50%. Apoptosis and cell cycle detection suggested that only phenanthrene affected cell cycle significantly and caused cell death; GTP lowered the mortality of HFL-I cells exposed to phenanthrene; However, GTP did not affect modulation of the cell cycle by phenanthrene. - Green tea polyphenols antagonised cytotoxicity of a low-ring PAH phenanthrene.

  19. Ispaghula (Plantago ovata) seed husk polysaccharides promote proliferation of human epithelial cells (skin keratinocytes and fibroblasts) via enhanced growth factor receptors and energy production.

    Science.gov (United States)

    Deters, A M; Schröder, K R; Smiatek, T; Hensel, Andreas

    2005-01-01

    Endogenous carbohydrates, especially oligo- and polysaccharides, participate in the regulation of a broad range of biological activities, e. g., signal transduction, inflammation, fertilisation, cell-cell-adhesion and act as in vivo markers for the determination of cell types. In the present study, water-soluble (WS) and gel-forming polysaccharides (GF) of ispaghula seed husk (Plantago ovata Forsskal, Plantaginaceae) were characterised as neutral and acidic arabinoxylans and tested under in vitro conditions for regulating activities on cell physiology of human keratinocytes and human primary fibroblasts. Only water-soluble polysaccharides exhibited strong and significant effects on cell physiology of keratinocytes and fibroblasts. Proliferation of cells of the spontaneously immortalised keratinocyte cell line HaCaT was significantly up-regulated in a dose-independent manner. Analysis of activated signal pathways by RNA analysis proved an effect of the acidic arabinoxylan on the expression of keratinocyte growth factor (KGF) in HaCaT cells. Differentiation behaviour of normal human keratinocytes (NHK) determined by involucrin was slightly influenced, due to the enhanced cell proliferation, leading to a cell-cell-mediated indirect induction of early differentiation. WS did not influence late differentiation, as determined by keratin K1 and K10 titres. PMID:15678371

  20. Comparison of colony-formation efficiency of bovine fetal fibroblast cell lines cultured with low oxygen, hydrocortisone, L-carnosine, bFGF, or different levels of FBS.

    Science.gov (United States)

    Talbot, Neil C; Powell, Anne M; Caperna, Thomas J

    2004-01-01

    A comparison of colony-formation efficiency (CFE) was made between six independent bovine fetal fibroblast (BFF) cell lines used in somatic cell nuclear transfer. Variation in CFE was assessed under different culture conditions. The conditions examined were ambient atmosphere (approximately 20% oxygen) culture versus 5% oxygen culture, three levels of fetal bovine serum (FBS) in the medium (5%, 10% or 20%), and the amendment of 10% FBS medium with basic fibroblast growth factor (1 ng/mL), L-carnosine (20 mM), or hydrocortisone (1 microM). The six BFF cell lines showed significant differences from one another in CFE. No significant difference in CFE was found with reduced oxygen culture. L-Carnosine also had no significant effect on CFE. A FBS concentration of 10% was found to produce the best overall CFE. Hydrocortisone treatment reduced the size of colonies although the number of colonies formed was not affected. Basic FGF increased the size of colonies but the number of colonies formed was not affected. The results showed that different BFF cell lines varied significantly in their CFE. Also, some medium supplements or culture conditions that have shown positive CFE effects on the fibroblasts of other species failed to show significant positive CFE effects on the BFF cell lines tested. PMID:15107245

  1. The effects of hypoxia on human skin, lung and tendon cells in vitro.

    OpenAIRE

    Webster, D. F.; Burry, H C

    1982-01-01

    Fibroblasts derived from human foetal tendon, skin and lung were cultured in media containing dissolved oxygen at concentrations of 20%, 10% and 2.7%. Total protein and collagen synthesis were reduced at 2.7% oxygen, tendon-derived cells proving significantly less sensitive than lung and skin fibroblasts. The effects of hypoxia were more marked in low-density cultures than in confluent cultures. Proliferation rates varied greatly between the 3 cell lines but were not affected by hypoxia. The ...

  2. Exploring the Anticancer Activity of Grape Seed Extract on Skin Cancer Cell Lines A431

    Directory of Open Access Journals (Sweden)

    V. Mohansrinivasan

    2015-08-01

    Full Text Available In this study, grape seeds were extracted using ethyl acetate and petroleum ether by solvent-solvent extraction method. The phytochemical tests were performed to identify different phytochemical compounds present in the grape seed extract (GSE. Antibacterial activity of the GSE was determined using agar diffusion method against Gram- positive and Gram-negative bacteria. Gas chromatography-mass spectrometry (GC-MS and Fourier transform infrared spectroscopy (FTIR analysis was done to identify the presence of bioactive compounds and their functional groups. The GC-MS results revealed a total of four compounds, known to have potent activity against cancer cells, viz, squalene, the most potent compound found in ethyl acetate extract and diethyl phthalate, ethyl-9- cis -11- trans octadecadienoate and (R-(--14,-methyl-8-Hexadecyn-1-ol in petroleum ether extract. Cytotoxic activity of the GSE was observed against skin cancer cell lines A4321 using 3-(4, 5-dimethylthiazol-2-yl-2-5-diphenyl tetrazolium bromide MTT assay. The IC50 value of the GSE against A431 skin cancer cell line was 480 µg/mL. This is first such report against A4321 cell lines. The study gives the overall perception about importance of GSE in medicine and nutraceuticals purposes.

  3. Benzo(a)pyrene metabolism, DNA-binding and UV-induced repair of DNA damage in cultured skin fibroblasts from a patient with unilateral multiple basal cell carcinoma

    International Nuclear Information System (INIS)

    The metabolism of benzo(a)pyrene (BP), and its subsequent binding to DNA, and the repair of UV-induced DNA damage were studied in fibroblasts cultured from the skin of a 61-year-old male who had multiple basal cell carcinoma (BCC) (>100) on his left upper trunk. Results suggest that BP metabolism and repair of DNA are altered in tumor-bearing site (TSB) cells and that patients with this type of metabolic profile may be at higher risk of the development of cutaneous neoplasms. It is also possible that fibroblasts from tumour bearing skin undergo some as yet unexplained alteration in carcinogen metabolism as a consequence of the induction of neoplasia. (author)

  4. Low-dose radiation activates Nrf1/2 through reactive species and the Ca2+/ERK1/2 signaling pathway in human skin fibroblast cells

    Directory of Open Access Journals (Sweden)

    Eun Kyeong Lee

    2013-05-01

    Full Text Available In the current study, we explored the effect of LDR on theactivation of Nrfs transcription factor involved in cellular redoxevents. Experiments were carried out utilizing 0.05 and 0.5 GyX-ray irradiated normal human skin fibroblast HS27 cells. Theresults showed LDR induced Nrf1 and Nrf2 activation andexpression of antioxidant genes HO-1, Mn-SOD, and NQO1.In particular, 0.05 Gy-irradiation increased only Nrf1 activation,but 0.5 Gy induced both Nrf1 and Nrf2 activation.LDR-mediated Nrf1/2 activation was accompanied by reactivespecies (RS generation and Ca2+ flux. This effect was abolishedin the presence of N-acetyl-cysteine and BAPTA- AM.Furthermore, Nrf1/2 activation by LDR was suppressed byPD98059, an inhibitor of ERK1/2. In conclusion, LDR inducesNrf1 and Nrf2 activation and expression of Nrf-regulatedantioxidant defense genes through RS and Ca2+/ERK1/2pathways, suggesting new insights into the molecularmechanism underlying the beneficial role of LDR in HS27cells. [BMB Reports 2013; 46(5: 258-263

  5. Both near ultraviolet radiation and the oxidizing agent hydrogen peroxide induce a 32-kDa stress protein in normal human skin fibroblasts

    International Nuclear Information System (INIS)

    We have analyzed the pattern of protein synthesis in solar near ultraviolet (334 nm, 365 nm) and near visible (405 nm) irradiated normal human skin fibroblasts. Two hours after irradiation we find that one major stress protein of approximately 32 kDa is induced in irradiated cells. This protein is not induced by ultraviolet radiation at wavelengths shorter than 334 nm and is not inducible by heat shock treatment of these cells. Although sodium arsenite, diamide, and menadione all induced a 32-kDa protein, they also induced the major heat shock proteins. In contrast, the oxidizing agent, hydrogen peroxide, induced the low molecular weight stress protein without causing induction of the major heat shock proteins. A comparison of the 32-kDa proteins induced by sodium arsenite, H2O2, and solar near ultraviolet radiation using chemical peptide mapping shows that they are closely related. These results imply that the pathways for induction of the heat shock response and the 32-kDa protein are not identical and suggest that, at least in the case of radiation and treatment with H2O2, the 32-kDa protein might be induced in response to cellular oxidative stress. This conclusion is supported by the observation that depletion of endogenous cellular glutathione prior to solar near ultraviolet irradiation lowers the fluence threshold for induction of the 32-kDa stress protein

  6. A correlation between ultraviolet-induced sister chromatid exchanges and ultraviolet-indced mutagenesis in ''Muntiacus muntjak'' (Indian Muntjac) skin fibroblasts in culture

    International Nuclear Information System (INIS)

    The purpose of this thesis was to develop the capability of simultaneously assaying SCEs and mutations in Indian muntjac cells to determine (1) the relationship between the ultraviolet radiation (UVR) induction of SCEs and the UVR induction of mutations at the (UVR) HGPRT locus in Indian muntjac cells and (2) the possible role of DNA repair in the UVR induction of these two events. Indian muntjac skin fibroblasts were chosen for this study because of a unique karyotype consisting of a diploid chromosome number of 6 in females and 7 in males. An HGPRT mutation assay in Indian muntjac cells was developed by this author since at the time this study was undertaken no mutational assay system utilizing Indian muntjac cells existed. It is concluded from this study that a linear correlation exists betwen the UVR-induction of SCEs and of mutations to 6TG resistance in Indian muntjac cells. As more time is allowed between the UVR-induced DNA damage and onset of DNA replication, more of the lesions leading to both mutations and SCE formation are repaired. The fact that SCE and mutation frequencies are reduced at different rates may indicate that the lesions responsible for SCEs and for mutations are repaired differently

  7. Establishment and characterization of equine fibroblast cell lines transformed in vivo and in vitro by BPV-1: Model systems for equine sarcoids

    International Nuclear Information System (INIS)

    It is now widely recognized that BPV-1 and less commonly BPV-2 are the causative agents of equine sarcoids. Here we present the generation of equine cell lines harboring BPV-1 genomes and expressing viral genes. These lines have been either explanted from sarcoid biopsies or generated in vitro by transfection of primary fibroblasts with BPV-1 DNA. Previously detected BPV-1 genome variations in equine sarcoids are also found in sarcoid cell lines, and only variant BPV-1 genomes can transform equine cells. These equine cell lines are morphologically transformed, proliferate faster than parental cells, have an extended life span and can grow independently of substrate. These characteristics are more marked the higher the level of viral E5, E6 and E7 gene expression. These findings confirm that the virus has an active role in the induction of sarcoids and the lines will be invaluable for further studies on the role of BPV-1 in sarcoid pathology

  8. Characterization of fibroblast growth factor receptor 2 overexpression in the human breast cancer cell line SUM-52PE

    International Nuclear Information System (INIS)

    The fibroblast growth factor receptor (FGFR)2 gene has been shown to be amplified in 5-10% of breast cancer patients. A breast cancer cell line developed in our laboratory, SUM-52PE, was shown to have a 12-fold amplification of the FGFR2 gene, and FGFR2 message was found to be overexpressed 40-fold in SUM-52PE cells as compared with normal human mammary epithelial (HME) cells. Both human breast cancer (HBC) cell lines and HME cells expressed two FGFR2 isoforms, whereas SUM-52PE cells overexpressed those two isoforms, as well as several unique FGFR2 polypeptides. SUM-52PE cells expressed exclusively FGFR2-IIIb isoforms, which are high-affinity receptors for fibroblast growth factor (FGF)-1 and FGF-7. Differences were identified in the expression of the extracellular Ig-like domains, acid box and carboxyl termini, and several variants not previously reported were isolated from these cells. The FGFR family of receptor tyrosine kinases includes four members, all of which are highly alternatively spliced and glycosylated. For FGFR2, alternative splicing of the second half of the third Ig-like domain, involving exons IIIb and IIIc, is a mutually exclusive choice that affects ligand binding specificity and affinity [1,2,3]. It appears that the second half of the third Ig-like domain can dictate high affinity for FGF-2 or keratinocyte growth factor (KGF), whereas affinity for FGF-1 appears to remain the same [3]. Alternative splicing of the carboxyl terminus has been shown to involve at least two different exons that can produce at least three different variants. The C1-type and C2-type carboxyl termini are encoded by the same exon, and have two different splice acceptor sites, whereas the C3-type carboxyl terminus is encoded by a separate exon [4]. The biologic significance of the C1 carboxyl terminus, as compared with the shorter C3 variant found primarily in tumorigenic samples, has been studied in NIH3T3 transfection assays, in which C3 variants were able to produce three times more transformed foci in soft agar than C1 variants (both IIIb), whereas full length FGFR2 and FGFR1 (both IIIc variants) showed no transforming activity [4]. Previous studies [5,6] have found amplification and overexpression of FGFR2 in 5-10% of primary breast cancer specimens. A recent study [7] done using a tissue array consisting of 372 primary breast cancer specimens found a 5% incidence of FGFR2 amplification. To our knowledge, none of the HBC cell lines studied thus far have an FGFR2 gene amplification, although overexpression of FGFR2 message and protein has been documented for some breast cancer cell lines [6,8,9]. SUM-52PE is a breast cancer cell line previously isolated in our laboratory that grows under serum-free and epidermal growth factor-free conditions, has high levels of tyrosine-phosphorylated membrane proteins, and has the capacity to invade and grow under anchorage-independent conditions [10,11,12]. This cell line exhibits all of the important hallmarks of transformed, highly malignant cells. Therefore, SUM-52PE was used as a model to study the diversity of FGFR2 expression in a breast cancer cell line that has true amplification and overexpression of FGFR2. This study was conducted to examine the degree of FGFR2 amplification and overexpression in the breast cancer cell line SUM-52PE. Subsequent sequencing and characterization of individual FGFR2 variants cloned from the SUM-52PE cell line was completed to determine the complexity of FGFR2 alternative splicing in the context of a highly metastatic breast cancer cell line. Southern, Northern and Western blot analyses were done in order to determine the degree of FGFR2 amplification and overexpression in the breast cancer cell line SUM-52PE. Individual FGFR2 variants were cloned out of SUM-52PE using FGFR2-specific primers in a reverse transcription (RT) polymerase chain reaction (PCR). FGFR2 cDNAs were characterized by restriction fragment analysis, sequencing and transient transfection into 293 cells to examine the protein expression of each FGFR2 clone. The results of the Southern blot showed that there was a 12-fold amplification of FGFR2 in the SUM-52PE cell line. Northern blot analysis of SUM-52PE showed FGFR2 transcripts to be highly overexpressed compared with other breast cancer cell lines and normal HME cells. Several overexpressed bands of approximately 6.3, 5.0, 4.0, and 2.8kb were observed in SUM-52PE cells. The most prominent band, at 2.8kb, was so abundant that it was difficult to discern other individual bands clearly. Western blot analysis showed that both normal HME and HBC cells expressed two FGFR2 variants of 95 and 135kDa. The SUM-52PE cell line greatly overexpressed not only these two polypeptides, as compared with HME and HBC cells, but also overexpressed two unique variants of FGFR2 - 85 and 109kDa polypeptides - as well as several smaller polypeptides in the 46-53kDa range. The antibody used in Western blot analysis only recognizes FGFR2 isoforms that express the C1 carboxyl termini, therefore greatly underestimating the actual number of different FGFR2 variants that are overexpressed in this cell line. PCR was performed to determine the proportion of C1/C2 variants as compared with C3 variants in the SUM-52PE cell line. Results of this analysis indicated the presence of all three types of variants in this cell line, although the C1/C2 variants were predominant as compared with the C3 variants in SUM-52PE. Four different FGFR2-C1 clones were isolated and sequenced from SUM-52PE cells, which differed in their signal sequence, first Ig-like loop, and acid box. Two FGFR2-C2 clones were isolated from the SUM-52PE cell line, which were identical to each other except for the variable expression of the number of Ig-like domains (two or three). Three C3 clones were isolated and sequenced, two of which have not previously been described in the literature. Clone C3-#3 contained two Ig-like domains, but no acid box. C3-#5 was missing the first two Ig-like domains and the acid box, but did contain the third Ig-like domain. There is an extensive amount of evidence implicating erbB-2, a gene that is overexpressed in approximately 30% of breast cancer cases, as a breast cancer gene [13]. The identification of other breast oncogenes that function in the remaining 70% of cases is an ongoing challenge, as is establishing a causal role for such oncogenes in HME cell transformation. FGFR1 and FGFR2, previously established oncogenes, were found to be amplified within large amplicons on 8p11 and 10q26, respectively, in the breast cancer cell line SUM-52PE [14]. Previous studies have shown that the FGFR2 gene is amplified in about 5-10% of breast cancer cases. Our results showed that SUM-52PE cells overexpressed many alternatively spliced isoforms of FGFR2 at both the transcript and protein level as compared with normal HME cells. The variability in FGFR2 isoform expression is complex and involves exon IIIb/c, which encodes the second half of the third Ig-like loop; variations in the carboxyl terminal end of the receptor, involving the C1/C2 or C3 domains; and variable expression of the Ig-like loops and acid box in the extracellular portion of the receptor. The characterization of three unique FGFR2 isoforms that were cloned from SUM-52PE may build on the findings of others concerning the transforming potential of FGFR2 variants [4]. In particular, because it has been demonstrated that expression of C3-IIIb variants may have more transforming activity than C1-IIIb variants, differences between the three C3 clones we have isolated may provide information regarding the influence of particular structural domains on transforming potential. Ongoing studies are aimed at characterizing the transforming ability of FGFR2 isoforms obtained from SUM-52PE cells by transducing these genes into normal HME cells. By overexpressing FGFR2 isoforms in a physiologically relevant system, we hope to determine the isoform(s) that acts in a dominant way in the process of cell transformation, and to determine whether different regions present in individual clones drive specific phenotypes associated with transformation

  9. Transformation and radiosensitivity of human diploid skin fibroblasts transfected with activated RAS oncogene and SV40 T-antigen

    Energy Technology Data Exchange (ETDEWEB)

    Su, L.-N.; Little, J.B. (Harvard School of Public Health, Boston, MA (United States))

    1992-08-01

    Three normal human diploid cell strains were transfected with an activated Ha-ras oncogene (EJ ras) or SV40 T-antigen. Multiple clones were examined for morphological alterations, growth requirements, ability to grow under anchorage independent conditions, immortality and tumorigenicity in nude mice. Clones expressing SV40 T-antigen alone or in combination with ras protein p21 were significantly radioresistant as compared with their parent cells or clones transfected with the neo gene only. This radioresistant phenotype persisted in post-crisis, immortalized cell lines. These data suggest that expression of the SV40 T-antigen but not activated Ha-ras plays an important role in the radiosensitivity of human diploid cells. The radioresistant phenotype in SV40 T transfected cells was not related to the enhanced level of genetic instability seen in pre-crisis and newly immortalized cells, nor to the process of immortalization itself. (author).

  10. Stimulation of MMP-11 (stromelysin-3) expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

    International Nuclear Information System (INIS)

    Matrix metalloproteinases (MMPs) are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14) and stromelysin-3 (MMP-11) are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs) were: a) treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b) grown on collagens I, IV and V; c) treated with fibronectin, con-A and matrigel; and d) co-cultured with a range of HBC (human breast cancer) cell lines of varied invasive and metastatic potential. Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms

  11. Stimulation of MMP-11 (stromelysin-3 expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

    Directory of Open Access Journals (Sweden)

    Matthaei Klaus I

    2004-07-01

    Full Text Available Abstract Background Matrix metalloproteinases (MMPs are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14 and stromelysin-3 (MMP-11 are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. Methods To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs were: a treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b grown on collagens I, IV and V; c treated with fibronectin, con-A and matrigel; and d co-cultured with a range of HBC (human breast cancer cell lines of varied invasive and metastatic potential. Results Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. Conclusion We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms.

  12. Microflora of the penile skin-lined neovagina of transsexual women

    Directory of Open Access Journals (Sweden)

    Claeys Geert

    2009-05-01

    Full Text Available Abstract Background The microflora of the penile skin-lined neovagina in male-to-female transsexuals is a recently created microbial niche which thus far has been characterized only to a very limited extent. Yet the knowledge of this microflora can be considered as essential to the follow-up of transsexual women. The primary objective of this study was to map the neo-vaginal microflora in a group of 50 transsexual women for whom a neovagina was constructed by means of the inverted penile skin flap technique. Secondary objectives were to describe possible correlations of this microflora with multiple patients' characteristics, such as sexual orientation, the incidence of vaginal irritation and malodorous vaginal discharge. Results Based on Gram stain the majority of smears revealed a mixed microflora that had some similarity with bacterial vaginosis (BV microflora and that contained various amounts of cocci, polymorphous Gram-negative and Gram-positive rods, often with fusiform and comma-shaped rods, and sometimes even with spirochetes. Candida cells were not seen in any of the smears. On average 8.6 species were cultured per woman. The species most often found were: Staphylococcus epidermidis, Streptococcus anginosus group spp., Enterococcus faecalis, Corynebacterium sp., Mobiluncus curtisii and Bacteroides ureolyticus. Lactobacilli were found in only one of 30 women There was no correlation between dilatation habits, having coitus, rinsing habits and malodorous vaginal discharge on the one hand and the presence of a particular species on the other. There was however a highly significant correlation between the presence of E. faecalis on the one hand and sexual orientation and coitus on the other (p = 0.003 and p = 0.027 respectively. Respectively 82%, 58% and 30% of the samples showed an amplicon after amplification with M. curtisii, Atopobium vaginae and Gardnerella vaginalis primer sets. Conclusion Our study is the first to describe the microflora of the penile skin-lined neovagina of transsexual women. It reveals a mixed microflora of aerobe and anaerobe species usually found either on the skin, in the intestinal microflora or in a BV microflora.

  13. Extracts of Sarcoptes scabiei De Geer Downmodulate Secretion of IL-8 by Skin Keratinocytes and Fibroblasts and of GM-CSF by Fibroblasts in the Presence of Proinflammatory Cytokines

    OpenAIRE

    Mullins, Jeremi S.; Arlian, Larry G.; MORGAN, MARJORIE S.

    2009-01-01

    Previous in vitro studies showed that molecules in an extract of the mite Sarcoptes scabiei variety canis De Geer could modulate the secretion of cytokines from cultured normal human epidermal keratinocytes and dermal fibroblasts in the absence of proinflammatory cytokines in the cell culture media. The purpose of this study was to investigate whether scabies extract could also modulate cytokine and chemokine secretion from epidermal keratinocytes and dermal fibroblasts in the presence of pro...

  14. Estrogens and aging skin

    OpenAIRE

    Thornton, M. Julie

    2013-01-01

    Estrogen deficiency following menopause results in atrophic skin changes and acceleration of skin aging. Estrogens significantly modulate skin physiology, targeting keratinocytes, fibroblasts, melanocytes, hair follicles and sebaceous glands, and improve angiogenesis, wound healing and immune responses. Estrogen insufficiency decreases defense against oxidative stress; skin becomes thinner with less collagen, decreased elasticity, increased wrinkling, increased dryness and reduced vascularity...

  15. Neural cell adhesion molecule mediates contact-dependent inhibition of growth of near-diploid mouse fibroblast cell line m5S/1M

    OpenAIRE

    1991-01-01

    A near-diploid mouse fibroblast cell line m5S/1M used in this study shows a high sensitivity to contact-dependent inhibition of growth, and the addition of EGF causes both morphological change and loss of contact-dependent inhibition of growth. The m5S/1M cell and its transformants obtained by x-ray irradiation have been used to search for the cell surface glycoproteins that are responsible for the growth regulation via cell-cell interactions. Lectin blotting analyses showed that the expressi...

  16. Probing pyruvate metabolism in normal and mutant fibroblast cell lines using 13C-labeled mass isotopomer analysis and mass spectrometry.

    Science.gov (United States)

    Riazi, Roya; Khairallah, Maya; Cameron, Jessie M; Pencharz, Paul B; Des Rosiers, Christine; Robinson, Brian H

    2009-12-01

    Fibroblast cell lines are frequently used to diagnose genetic mitochondrial defects in children. The effect of enzyme deficiency on overall flux rate through metabolic pathways is, however, not generally considered. We have transposed an experimental paradigm that was developed for isolated perfused organs using (13)C-labeled substrates and (13)C-isotopomer analysis to probe pyruvate mitochondrial metabolism in cultured human fibroblast cell lines with normal or genetically mutant pyruvate decarboxylation (PDC) or carboxylation (PC) activity. Cells were incubated with 1mM [U-(13)C]pyruvate, and the (13)C-molar percent enrichment (MPE) of intracellular pyruvate, citrate, malate (as a surrogate of oxaloacetate) and aspartate was assessed by mass spectrometry. We estimated various flux ratios relevant to metabolic pathways involved in energy production, namely pyruvate formation, PDC, PC, and citrate recycling in the citric acid cycle (CAC). In all cell lines, exogenous pyruvate was predominately decarboxylated (PC/PDC ratios 0.01-0.3). PC-deficient cell lines displayed an expected negligible contribution of PC flux to oxaloacetate formation for citrate synthesis (PC/CS), which was associated with a greater contribution of PDC to acetyl-CoA formation (PDC/CS), and greater recycling of (13)C-labeled citrate into the CAC. In PDH-deficient cell lines, metabolic flux alterations were most apparent in cells with more than 50% reduction in enzyme activity. This led to an unexpected lower PC/CS flux ratio, while the PDC/CS flux ratio was unchanged. These data illustrate the usefulness of this approach in identifying unexpected metabolic consequences of genetic defects related to pyruvate metabolism. PMID:19640754

  17. Monoclonal antibody to the type I insulin-like growth factor (IGF-I) receptor blocks IGF-I receptor-mediated DNA synthesis: clarification of the mitogenic mechanisms of IGF-I and insulin in human skin fibroblasts

    International Nuclear Information System (INIS)

    Insulin and insulin-like growth factor type I (IGF-I) stimulate an overlapping spectrum of biological responses in human skin fibroblasts. Although insulin and IGF-I are known to stimulate the incorporation of [3H]thymidine into DNA in these cells, the identify of the receptor(s) that mediates this effect has not been fully clarified. The mouse anti-human IGF-I receptor antibody ?IR-3 binds with specificity to IGF-I but not to insulin receptors in human placental membranes; it also specifically inhibits the binding of 125I-labeled IGF-I but not 125I-labeled insulin to suspensions of human skin fibroblasts in a dose-dependent manner. ?IR-3 competitively inhibits IGF-I-mediated stimulation of [3H]thymidine incorporation into DNA. This inhibition is dependent on the concentration of ?IR-3 and in the presence of a fixed antibody concentration can be partially overcome by high concentrations of IGF-I. In contrast, at concentrations of 3H]thymidine incorporation is not inhibited by ?IR-3. However, the incremental effects of higher concentrations (> 1 ?g/ml) of insulin on [3H]thymidine incorporation are inhibited by ?IR-3. ?IR-3 is a highly specific antagonist of IGF-I receptor-mediated mitogenesis in human skin fibroblasts. By using this antibody, it is shown directly that insulin can act through the IGF-I receptor to stimulate DNA synthesis but can also activate this effect through the insulin receptor itself

  18. Cytotoxicity and Proliferation Studies with Arsenic in Established Human Cell Lines: Keratinocytes, Melanocytes, Dendritic Cells, Dermal Fibroblasts, Microvascular Endothelial Cells, Monocytes and T-Cells

    Directory of Open Access Journals (Sweden)

    Hari H. P. Cohly

    2003-01-01

    Full Text Available Abstract: Based on the hypothesis that arsenic exposure results in toxicity and mitogenecity, this study examined the dose-response of arsenic in established human cell lines of keratinocytes (HaCaT, melanocytes (1675, dendritic cells (THP-1/A23187, dermal fibroblasts (CRL1904, microvascular endothelial cells (HMEC, monocytes (THP-1, and T cells (Jurkat. Cytotoxicity was determined by incubating THP-1, THP-1+ A23187 and JKT cells in RPMI 1640, 1675 in Vitacell, HMEC in EBM, and dermal fibroblasts and HaCaT in DMEM with 10% fetal bovine serum, 1% streptomycin and penicillin for 72 hrs in 96-well microtiter plates, at 37oC in a 5% CO2 incubator with different concentrations of arsenic using fluorescein diacetate (FDA. Cell proliferation in 96-well plates was determined in cultured cells starved by prior incubation for 24 hrs in 1% FBS and exposed for 72 hours, using the 96 cell titer proliferation solution (Promega assay. Cytotoxicity assays yielded LD50s of 9 μg/mL for HaCaT, 1.5 μg/mL for CRL 1675, 1.5 μg/mL for dendritic cells, 37 μg/mL for dermal fibroblasts, 0.48 μg/mL for HMEC, 50 μg/mL for THP-1 cells and 50 μg/mL for JKT-T cells. The peak proliferation was observed at 6 μg/mL for HaCaT and THP-1 cells, 0.19 μg/mL for CRL 1675, dendritic cells, and HMEC, and 1.5 μg/mL for dermal fibroblasts and Jurkat T cells. These results show that arsenic is toxic at high doses to keratinocytes, fibroblasts, monocytes and T cells, and toxic at lower doses to melanocytes, microvascular endothelial cells and dendritic cells. Proliferation studies showed sub-lethal doses of arsenic to be mitogenic.

  19. Degradation of blood-group A glycolipids in cultures of human skin fibroblasts; an approach to investigation of some lysosomal enzyme defects

    International Nuclear Information System (INIS)

    Degradation of blood group A glycosphingolipid A-6-2 has been studied by loading experiments in cultured human skin from controls and from patients with inherited lysosomal enzymopathies (?-N-acetylgactosaminidase, ?-fucosidase, ?-glucocerebrosidase, GM1 ?-galactosidase, ceramidase, sap B and sap-precursor deficiencies). In the cell lines from the most unrelated enzymopathies, accumulation of the glycosphingolipids was found corresponding to the inherent enzyme defect. The technique of feeding of radiolabelled glycolipids to cells in culture and analysis of their metabolism was used to examine whether cells from patients with deficiency of ?-N-acetylgactosaminidase (?-NAGA deficiency, Schindler disease). Further we have extended our study with the same probe to other lysosomal enzymophatic cell lines. For this purpose we isolated glycosphingolipid A-6-2 (IV2-?-fucosyl-IV3-?-N-acetylgactosaminylnelactotetraosylceramide) from erythrocyte membrane and labelled it on the ceramide moiety with tritium. The results of this study clearly show that loading experiments in cell culture are a valuable tool to analyze general degradation pathways, assess the physiological significance and the substrate specificity in vivo of the enzymes involved and estimate their residual activities or that of alternative degradation pathways. (authors)

  20. Laser Doppler line scanner for monitoring skin perfusion changes of port wine stains during vascular-targeted photodynamic therapy

    Science.gov (United States)

    Chen, Defu; Ren, Jie; Wang, Ying; Gu, Ying

    2014-11-01

    Vascular-targeted photodynamic therapy (V-PDT) is known to be an effective therapeutic modality for the treatment of port wine stains (PWS). Monitoring the PWS microvascular response to the V-PDT is crucial for improving the effectiveness of PWS treatment. The objective of this study was to use laser Doppler technique to directly assess the skin perfusion in PWS before and during V-PDT. In this study, 30 patients with PWS were treated with V-PDT. A commercially laser Doppler line scanner (LDLS) was used to record the skin perfusion of PWS immediately before; and at 1, 3, 5, 7, 10, 15 and 20 minutes during V-PDT treatment. Our results showed that there was substantial inter- and intra-patient perfusion heterogeneity in PWS lesion. Before V-PDT, the comparison of skin perfusion in PWS and contralateral healthy control normal skin indicated that PWS skin perfusion could be larger than, or occasionally equivalent to, that of control normal skin. During V-PDT, the skin perfusion in PWS significantly increased after the initiation of V-PDT treatment, then reached a peak within 10 minutes, followed by a slowly decrease to a relatively lower level. Furthermore, the time for reaching peak and the subsequent magnitude of decrease in skin perfusion varied with different patients, as well as different PWS lesion locations. In conclusion, the LDLS system is capable of assessing skin perfusion changes in PWS during V-PDT, and has potential for elucidating the mechanisms of PWS microvascular response to V-PDT.

  1. A New Type of Signaling Pathways as Pilomotor Lines along Skin for Transmitting Acupuncture Signals to Produce Acupuncture Effects.

    Science.gov (United States)

    Liu, Li-Yuan

    2015-06-30

    In our previous study, we observed a linear system consisted of sympathetic endings in the arrector pili muscles (AP muscles) along the rat skin termed sympathetic substance lines, or SSLs. After shaving the hair of the rats, the first wave of hair re-growth was not evenly distributed, but followed specific hair loop lines (HLLs). The patterns of HLL and SSL correspond with each other and also with the "Meridians" described in Chinese traditional medicine (CTM). Here I investigated in rabbits and rats whether the acupuncture signals are transmitted via the SSL/HLL, and whether the acupuncture analgesia (AA) is dependent on any peripheral mechanism. Firstly, when acupuncture was operated or phenylephrine, an agonist for ? receptor, was injected into the dermis at an acupoint, a pilomotor line occurred. The course of the pilomotor line coincided with the SSL/HLL. When the skin was incised or regitin, an antagonist for ? receptor, was injected into the dermis, the pilomotor line did not cross the site of incision or injection. These results directly demonstrated the process of transmission of acupuncture signals involving the pilomotor line and the sympathetic. Secondly, AA produced by acupuncture at an acupoint was significantly blocked when the skin was incised or regitin was injected into the dermis along the SSL/HLL or the Meridian. These results suggest that the factor that blocked the pilomotor line also blocked the AA and the pilomotor line related to the AA. Lastly, noradrenaline was shown to be released from the skin along the Meridian line after acupuncture; when phenylephrine was injected into an acupoint, AA was strongly simulated. All these results indicate that: 1. the transmission pathway of acupuncture signals exists in the skin, just as the Meridians described in the CTM; 2. these pathways are the SSLs/HLLs and the pilomotor lines; and 3. the pilomotor line is just for the transmission of acupuncture signals and the transmission is dependent on the ? receptor in the AP muscles, specifically the contraction of the AP muscles. Moreover, these findings suggest a new system and a new type of signal transmission in the physiology. PMID:26014122

  2. Liposomes encapsulating Aloe vera leaf gel extract significantly enhance proliferation and collagen synthesis in human skin cell lines.

    Science.gov (United States)

    Takahashi, Makoto; Kitamoto, Dai; Asikin, Yonathan; Takara, Kensaku; Wada, Koji

    2009-01-01

    Aloe vela leaf gel extract (AGE) are widely used as cosmetic and pharmaceutical ingredients because of its versatile skin care properties. In order to enhance the bioavailability of AGE, liposomes encapsulating AGE were prepared and examined for their interfacial and biochemical properties. The liposomes prepared from a soybean lecithin (SLP-WHITE, 1.0 wt%) by the Bangham method gave relatively a good trapping efficiency up to the AGE concentration of 0.5 wt%. The stable liposomes were then prepared from 1.0 wt% of SLP-WHITE and different concentrations of AGE by the mechanochemical method using a homogenizer and microfluidizer. The liposomes obtained from 0.25 wt% of AGE were confirmed to be small unilamellar vesicles with a diameter of less than 200 nm, and remained well dispersed for at least two weeks. The obtained liposomes encapsulating AGE were further examined for the effects on proliferation and type I collagen synthesis in normal human neonatal skin fibroblasts, NB1RGB cells. Liposomal AGE clearly showed higher proliferation rate than that of AGE alone. In addition, compared to the control, liposomal AGE significantly increased the collagen synthesis by 23%, while AGE alone showed a small effect. Liposomal AGE was also assayed for the effect on proliferation in normal human epidermal keratinocytes, NHEK(F) cells. Interestingly, liposomal AGE fractions containing 4 and 20 microg/mL of the extract considerably increased the proliferation rate by 77% and 101%, respectively. In contrast, AGE alone fractions containing 4 and 20 microg/mL of the extract increased the rate by 41% and 60%, respectively. Accordingly, the bioavailability and skin care properties of AGE will be significantly enhanced by liposome encapsulation, and the present liposomal AGE should have a great potential as an effective skin care formulation. PMID:19915322

  3. GLUT10 deficiency leads to oxidative stress and non-canonical ?v?3 integrin-mediated TGF? signalling associated with extracellular matrix disarray in arterial tortuosity syndrome skin fibroblasts.

    Science.gov (United States)

    Zoppi, Nicoletta; Chiarelli, Nicola; Cinquina, Valeria; Ritelli, Marco; Colombi, Marina

    2015-12-01

    Arterial tortuosity syndrome (ATS) is an autosomal recessive connective tissue disorder caused by loss-of-function mutations in SLC2A10, which encodes facilitative glucose transporter 10 (GLUT10). The role of GLUT10 in ATS pathogenesis remains an enigma, and the transported metabolite(s), i.e. glucose and/or dehydroascorbic acid, have not been clearly elucidated. To discern the molecular mechanisms underlying the ATS aetiology, we performed gene expression profiling and biochemical studies on skin fibroblasts. Transcriptome analyses revealed the dysregulation of several genes involved in TGF? signalling and extracellular matrix (ECM) homeostasis as well as the perturbation of specific pathways that control both the cell energy balance and the oxidative stress response. Biochemical and functional studies showed a marked increase in ROS-induced lipid peroxidation sustained by altered PPAR? function, which contributes to the redox imbalance and the compensatory antioxidant activity of ALDH1A1. ATS fibroblasts also showed activation of a non-canonical TGF? signalling due to TGFBRI disorganization, the upregulation of TGFBRII and connective tissue growth factor, and the activation of the ?v?3 integrin transduction pathway, which involves p125FAK, p60Src and p38 MAPK. Stable GLUT10 expression in patients' fibroblasts normalized redox homeostasis and PPAR? activity, rescued canonical TGF? signalling and induced partial ECM re-organization. These data add new insights into the ATS dysregulated biological pathways and definition of the pathomechanisms involved in this disorder. PMID:26376865

  4. Injury induces in vivo expression of platelet-derived growth factor (PDGF) and PDGF receptor mRNAs in skin epithelial cells and PDGF mRNA in connective tissue fibroblasts

    International Nuclear Information System (INIS)

    Platelet-derived growth factor (PDGF) stimulates many of the processes important in tissue repair, including proliferation of fibroblasts and synthesis of extracellular matrices. In this study, the authors have demonstrated with in situ hydridization and immunocytochemistry the reversible expression of 3-sis/PDGF-2 and PDGF receptor (PDGF-R) b mRNAs and their respective protein products in epithelial cells and fibroblasts following cutaneous injury in pigs. Epithelial cells in control, unwounded skin did not express c-sis and PDGF-R mRNAs, and fibroblasts expressed only PDGF-R mRNA. The expression levels in the injured site were correlated with the stage of tissue repair, being highest during the initial stages of the repair process and declining at the time of complete re-epithelialization and tissue remodeling. These studies provide a mulecular basis for understanding the mechanisms contributing to normal tissue repair. They suggest the possibility that a defect in these mechanisms may be associated with defective wound healing. It is also conceivable that chronic injury may induce irreversible gene expression leading to pathologic, unregulated cell growth

  5. Ochratoxin A and T-2 Toxin Induce Clonogenicity and Cell Migration in Human Colon Carcinoma and Fetal Lung Fibroblast Cell Lines.

    Science.gov (United States)

    Abassi, Haila; Ayed-Boussema, Imen; Shirley, Sarah; Abid, Salwa; Bacha, Hassen

    2016-03-01

    T-2 toxin and Ochratoxin A (OTA) are toxic secondary metabolites produced by various fungi, and together they contaminate feedstuffs worldwide. T-2 toxin and OTA may exert carcinogenic action in rodent. Despite the various in vivo experiments, carcinogenicity of these two mycotoxins has not yet been proven for human. In this current study, we proposed to investigate, in Human colon carcinoma cells and fetal lung fibroblast-like cells transfected with MYC, the effect of T-2 toxin and OTA on cell clonogenicity and cell migration. Results of the present investigation showed that T2-toxin as well as OTA has an important clonogenic effect in all cell lines, suggesting that these mycotoxins could promote the transcription of c-myc gene. Furthermore, T-2 toxin and OTA enhanced the migration effect of HCT116 cells at very low concentrations, proposing that these mycotoxins may exhibit carcinogenesis-like properties in the studied cells. PMID:26849850

  6. 3D flow investigation near the denticles of biomimetic shark skin model using Digital In-line Holographic Microscopy

    Science.gov (United States)

    Toloui, Mostafa; Hong, Jiarong

    2013-11-01

    It has been hypothesised that the complex microscopic denticles on a shark skin reduce the total drag for a swimming shark. However, the fundamental mechanism of this hydrodynamic function is not fully understood due to the inability to reproduce the complex shark surface and resolve the detailed flow around the skin denticles. Here we report a preliminary experiment using a 3D printed transparent rough surface replicating the morphological features of real shark skin. The model skin consists of closely-packed denticles of 2 mm in scale, i.e. ~ 10 times of the real size. Particle image velocimetry based on digital in-line holography is employed to measure 3D flow structures. To reduce optical abberration and enable imaging around the denticles, we use a fluid medium that has the same optical refractive index as that of the skin model. The experiment is conducted in 2'' 2'' square channel at a moderate Re number matching the general flow around a cruising shark. Several samples of the 3D velocity field amid and above the denticles are obtained. The follow-up research envisions a large dataset of these samples over the rigid/deformable model operated in stationary/undulating mode to ellucidate the dominant flow structures generated by the denticals. This research is collaborated with Prof. George Lauder's group.

  7. Development of an artificial lock for the skin-pass section in a hot dip galvanising line

    International Nuclear Information System (INIS)

    In this paper, we present the application of data mining techniques to develop an artificial lock for the skin-pass in an attempt to solve a problem that can arise during the galvanising manufacturing process:the wrong labelling of the steel grade of a coil. In order to detect these errors and thus to avoid that coils with different properties than expected end up with a client, we propose neural network-based models for on-line predicting the strip elongation in the skin-pass section according to the manufacturing conditions and its chemical composition. thus, a significant difference between estimated and measured elongation would mean that the coil must be removed from the line for further analyses. (Author) 14 refs

  8. Differentiation of normal and transformed human fibroblasts in vitro is influenced by electromagnetic fields

    International Nuclear Information System (INIS)

    We studied the effect of symmetric, biphasic sinusoidal electromagnetic fields (EMF) (20 Hz, 6 mT) on the differentiation of normal human skin fibroblasts (HH-8), normal human lung fibroblasts (WI38), and SV40-transformed human lung fibroblasts (WI38SV40) in in vitro cultures. Cells were exposed up to 21 days for 2 x 6 h per day to EMF. Normal mitotic human skin and lung fibroblasts could be induced to differentiate into postmitotic cells upon exposure to EMF. Concomitantly, the synthesis of total collagen as well as total cellular protein increased significantly by a factor of 5-13 in EMF-induced postmitotic cells. As analyzed by two-dimensional gel electrophoresis of [35S]methionine-labeled polypeptides, EMF-induced postmitotic cells express the same differentiation-dependent and cell type-specific marker proteins as their spontaneously arising counterparts. In SV40-transformed human lung fibroblasts (cell line WI38SV40) the exposure to EMF induced the differentiation of mitotic WI38SV40 cells into postmitotic and degenerating cells in subpopulations of WI38SV40 cell cultures. Other subpopulations of WI38SV40 cells did not show any effect of EMF on cell proliferation and differentiation. These results indicate that long-term EMF exposure of fibroblasts in vitro induces the differentiation of mitotic to postmitotic cells that are characterized by differentiation-specific proteins and differentiation-dependent enhanced metabolic activities

  9. Chemically induced skin carcinogenesis in a transgenic mouse line (TG.AC) carrying a v-Ha-ras gene.

    Science.gov (United States)

    Spalding, J W; Momma, J; Elwell, M R; Tennant, R W

    1993-07-01

    A transgenic mouse line (TG.AC) created in the FVB/N strain, carries a v-Ha-ras gene fused to a zeta-globin promoter gene. These trangenic mice have the properties of genetically initiated skin and have been shown to be sensitive to 12-O-tetradecanoylphorbol-13-acetate (TPA), a well-described promoter of skin papillomas in the two-stage mouse skin tumorigenesis model. It was of interest to determine whether the TG.AC mouse strain was also responsive to other known promoters. Groups of heterozygous or homozygous TG.AC mice were treated topically, 2x/week, for up to 20 weeks with benzoyl peroxide (BPO), 2-butanol peroxide (2-BUP), phenol (PH), acetic acid (AA), TPA and acetone (ACN), the vehicle control. Skin papillomas were induced in all groups treated with TPA, BPO and 2-BUP. Papillomas were observed in some treatment groups as early as 3 weeks. The relative activity of the promoters was TPA > 2-BUP > BPO > PH = AA = ACN. No papillomas were observed in any of the uninitiated FVB/N mice treated in a similar manner and which served as treatment control groups. Studies to determine the sensitivity of TG.AC mice to TPA, indicated that a total dose of 25-30 micrograms of TPA administered in 3 or 10 applications, was sufficient to induce an average incidence of 11-15 papillomas per mouse. The papilloma incidence continued to increase and was maintained up to 15 weeks after TPA treatment was terminated. The short latency period and high incidence of papilloma induction indicate that TG.AC mice have a high sensitivity to known skin promoters. The TG.AC line should prove to be a sensitive model for identifying putative tumor promoters or complete carcinogens. PMID:8330346

  10. Cytotoxic Evaluation of Elastomeric Dental Impression Materials on a Permanent Mouse Cell Line and on a Primary Human Gingival Fibroblast Culture

    Directory of Open Access Journals (Sweden)

    Roberta Tiozzo

    2009-08-01

    Full Text Available The need for clinically relevant in vitro tests of dental materials is widely recognized. Nearly all dental impression materials are introduced into the mouth just after mixing and allowed to set in contact with the oral tissues. Under these conditions, the materials may be toxic to cells or may sensitize the tissues. The aim of the present study is to evaluate the potential cytotoxicity of new preparations of elastomeric dental impression materials: A four vinylpolysiloxanes: Elite H-D Putty and Elite H-D Light Body (Zhermack, Badia Polesine, Rovigo, Italy; Express Putty and Express Light Body (3M ESPE AG Seefeld, Germany and B two polyethers: Impregum Penta and Permadyne Penta L (3M ESPE AG Seefeld, Germany. The cytotoxicity of these impression materials were examined using two different cell lines: Balb/c 3T3 (permanent cell line and human gingival fibroblasts (primary cell line and their effects were studied by indirect and direct tests. The direct tests are performed by placing one sample of the impression materials in the centre of the Petri dishes at the time of the seeding of cells. The cell growth was evaluated at the 12th and 24th hours by cell number. The indirect tests were performed by incubating a square of 1 cm diameter impression material in 5 mL of medium at 37 C for 24 hours (eluates. Subconfluent cultures are incubated with eluates for 24 hours. The MTT-formazan production is the method used for measuring the cell viability. The results indicate that: a polyether materials are cytotoxic under both experimental conditions; b among vinylpolysiloxanes, only Express Light Body (3M ESPE AG Seefeld, Germany induces clear inhibition of cellular viability of Balb/c 3T3 evaluated by direct and indirect tests and c the primary cell line is less sensitive to the toxic effect than the permanent cell line.

  11. Increased transcription of the c-myc oncogene in two methylcholanthrene-induced quail fibroblastic cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Saule, S.; Martin, P.; Gegonne, A.; Begue, A.; Lagrou, C.; Stehelin, D.

    1984-12-01

    The expression of three c-onc genes (c-erb, c-myc, c-myb) was investigated in five cell lines established from fibrosarcomas induced with 20-methylcholanthrene (MCA) of Japanese quails. These cell lines showed low levels of the three c-onc genes, with the exception of two cell lines that accumulated moderate (MCAQ 1-4) and large amounts (MCAQ 3-5) of c-myc RNA. Molecular cloning and restriction endonuclease analyses indicated that expression of c-myc in these two cell lines were not associated with detectable rearrangements in the c-myc locus, that the size of the c-myc transcript (2.7 kb) in MCAQ 3-5 was similar to that of the normal c-myc messenger RNAs (mRNA) and that the transcriptional activatin observed in MCAQ 3-5 was not mediated by the LTR (long terminal repeat) of a proximate ALV (avian leukosis virus) provirus. Finally, when analyzed with the restriction enzymes Msp I and Hpa II, the c-myc locus of MCAQ 3-5 and MCAQ 1-4 was found hypomethylated as compared with that of the other cell lines tested that show low levels of c-myc transcripts. Results suggest that one of the ways methylcholanthrene could mediate transformation is by inducing an abnormal regulation of the c-myc gene.

  12. Local dose assessment by PCC-FISH in skin fibroblasts: application to the case of one of the victims of the 2001 georgian radiological accident

    International Nuclear Information System (INIS)

    A new method of biodosimetry is proposed that could be applied in case of localized irradiation. The approach is based on excess chromosome segments determination by the PCC-FISH technique in fibroplasts isolated from skin biopsy

  13. Increased susceptibility of spinal muscular atrophy fibroblasts to camptothecin is p53-independent

    Directory of Open Access Journals (Sweden)

    Funanage Vicky L

    2009-05-01

    Full Text Available Abstract Background Deletion or mutation(s of the survival motor neuron 1 (SMN1 gene causes spinal muscular atrophy (SMA. The SMN protein is known to play a role in RNA metabolism, neurite outgrowth, and cell survival. Yet, it remains unclear how SMN deficiency causes selective motor neuron death and muscle atrophy seen in SMA. Previously, we have shown that skin fibroblasts from SMA patients are more sensitive to the DNA topoisomerase I inhibitor camptothecin, supporting a role for SMN in cell survival. Here, we examine the potential mechanism of camptothecin sensitivity in SMA fibroblasts. Results Camptothecin treatment reduced the DNA relaxation activity of DNA topoisomerase I in human fibroblasts. In contrast, kinase activity of DNA topoisomerase I was not affected by camptothecin, because levels of phosphorylated SR proteins were not decreased. Upon camptothecin treatment, levels of p53 were markedly increased. To determine if p53 plays a role in the increased sensitivity of SMA fibroblasts to camptothecin, we analyzed the sensitivity of SMA fibroblasts to another DNA topoisomerase I inhibitor, β-lapachone. This compound is known to induce death via a p53-independent pathway in several cancer cell lines. We found that β-lapachone did not induce p53 activation in human fibroblasts. In addition, SMA and control fibroblasts showed essentially identical sensitivity to this compound. By immunofluorescence staining, SMN and p53 co-localized in gems within the nucleus, and this co-localization was overall reduced in SMA fibroblasts. However, depletion of p53 by siRNA did not lessen the camptothecin sensitivity in SMA fibroblasts. Conclusion Even though p53 and SMN are associated, the increased sensitivity of SMA fibroblasts to camptothecin does not occur through a p53-dependent mechanism.

  14. Modulation of radio-induced oxidative damage by the combination of pentoxifylline and γ-tocopherol in skin fibroblasts and microvascular endothelial cells

    International Nuclear Information System (INIS)

    Clinical or accidental localized ionizing radiation exposure can induce severe skin damage constituting the cutaneous radiological syndrome which is divided in acute and late phases. The combination of pentoxifylline (PTX), antioxidant phytochemical, and γ-tocopherol, antioxidant nutrient shows effectiveness in reducing the late radio-induced skin damage with a long period. This work aims to investigate the molecular and cellular mechanisms involved in the effects of this combination

  15. Whole-exome sequencing of fibroblast and its iPS cell lines derived from a patient diagnosed with xeroderma pigmentosum

    Directory of Open Access Journals (Sweden)

    Kohji Okamura

    2015-12-01

    Full Text Available Cells from a patient with a DNA repair-deficiency disorder are anticipated to bear a large number of somatic mutations. Because such mutations occur independently in each cell, there is a high degree of mosaicism in patients' tissues. While major mutations that have been expanded in many cognate cells are readily detected by sequencing, minor ones are overlaid with a large depth of non-mutated alleles and are not detected. However, cell cloning enables us to observe such cryptic mutations as well as major mutations. In the present study, we focused on a fibroblastic cell line that is derived from a patient diagnosed with xeroderma pigmentosum (XP, which is an autosomal recessive disorder caused by a deficiency in nucleotide excision repair. By making a list of somatic mutations, we can expect to see a characteristic pattern of mutations caused by the hereditary disorder. We cloned a cell by generating an iPS cell line and performed a whole-exome sequencing analysis of the progenitor and its iPS cell lines. Unexpectedly, we failed to find causal mutations in the XP-related genes, but we identified many other mutations including homozygous deletion of GSTM1 and GSTT1. In addition, we found that the long arm of chromosome 9 formed uniparental disomy in the iPS cell line, which was also confirmed by a structural mutation analysis using a SNP array. Type and number of somatic mutations were different from those observed in XP patients. Taken together, we conclude that the patient might be affected by a different type of the disorder and that some of the mutations that we identified here may be responsible for exhibiting the phenotype. Sequencing and SNP-array data have been submitted to SRA and GEO under accession numbers SRP059858 and GSE55520, respectively.

  16. Whole-exome sequencing of fibroblast and its iPS cell lines derived from a patient diagnosed with xeroderma pigmentosum.

    Science.gov (United States)

    Okamura, Kohji; Toyoda, Masashi; Hata, Kenichiro; Nakabayashi, Kazuhiko; Umezawa, Akihiro

    2015-12-01

    Cells from a patient with a DNA repair-deficiency disorder are anticipated to bear a large number of somatic mutations. Because such mutations occur independently in each cell, there is a high degree of mosaicism in patients' tissues. While major mutations that have been expanded in many cognate cells are readily detected by sequencing, minor ones are overlaid with a large depth of non-mutated alleles and are not detected. However, cell cloning enables us to observe such cryptic mutations as well as major mutations. In the present study, we focused on a fibroblastic cell line that is derived from a patient diagnosed with xeroderma pigmentosum (XP), which is an autosomal recessive disorder caused by a deficiency in nucleotide excision repair. By making a list of somatic mutations, we can expect to see a characteristic pattern of mutations caused by the hereditary disorder. We cloned a cell by generating an iPS cell line and performed a whole-exome sequencing analysis of the progenitor and its iPS cell lines. Unexpectedly, we failed to find causal mutations in the XP-related genes, but we identified many other mutations including homozygous deletion of GSTM1 and GSTT1. In addition, we found that the long arm of chromosome 9 formed uniparental disomy in the iPS cell line, which was also confirmed by a structural mutation analysis using a SNP array. Type and number of somatic mutations were different from those observed in XP patients. Taken together, we conclude that the patient might be affected by a different type of the disorder and that some of the mutations that we identified here may be responsible for exhibiting the phenotype. Sequencing and SNP-array data have been submitted to SRA and GEO under accession numbers SRP059858 and GSE55520, respectively. PMID:26697316

  17. Characteristics of changes in the number of yH2AX and Rad51 protein foci in human skin fibroblasts after prolonged exposure to low-dose rate X-ray radiation

    Directory of Open Access Journals (Sweden)

    Ozerov I.V.

    2014-12-01

    Full Text Available Aim: to compare the repair process of DNA double-strand breaks in mammalian cells after acute versus prolonged exposure to X-ray irradiation with different dose rates. Material and methods. Studies were performed on primary human fibroblasts isolated from skin biopsies of healthy volunteers (women, 29 and 30 years. Cells were irradiated using an X-ray machine RUB RUST-M1 (JSC "Ruselectronics", Moscow, Russia at 37C temperature with a dose rate of 400 mGy/min (200 kV, 2*2.4 mA, a filter of 1.5mm AI or 4 mGy/min (50 kV, 2*0.4 mA, a filter of 1.5 mm AI. Immuno-cytochemical protein staining was utilized for yH2AX and Rad51 foci analysis. Results. Phosphorylated histone H2AX (yH2AX and the key protein of homologous recombination Rad51 foci formation and disappearance kinetics were investigated simultaneously in primary human dermal fibroblasts after acute and prolonged exposure to X-ray radiation at a same dose. It was shown that the relative yield of yH2AX foci per dose reduces with decrease in dose rate, while the relative yield of Rad51 foci conversely increases. Conclusion. Our findings suggest the fundamental differences in the ratio of non-homologous end joining and homologous recombination DNA repair in acute versus prolonged irradiated cells.

  18. Mouse fibroblast L929 cells are less permissive to infection by Nelson Bay orthoreovirus compared to other mammalian cell lines.

    Science.gov (United States)

    Mok, Lawrence; Wynne, James W; Grimley, Samantha; Shiell, Brian; Green, Diane; Monaghan, Paul; Pallister, Jackie; Bacic, Antony; Michalski, Wojtek P

    2015-07-01

    In recent years, bats have been identified as a natural reservoir for a diverse range of viruses. Nelson Bay orthoreovirus (NBV) was first isolated from the heart blood of a fruit bat (Pteropus poliocephalus) in 1968. While the pathogenesis of NBV remains unknown, other related members of this group have caused acute respiratory disease in humans. Thus the potential for NBV to impact human health appears plausible. Here, to increase our knowledge of NBV, we examined the replication and infectivity of NBV using different mammalian cell lines derived from bat, human, mouse and monkey. All cell lines supported the replication of NBV; however, L929 cells showed a greater than 2 log reduction in virus titre compared with the other cell lines. Furthermore, NBV did not induce major cytopathic effects in the L929 cells, as was observed in other cell lines. Interestingly, the related Pteropine orthoreoviruses, Pulau virus (PulV) and Melaka virus (MelV) were able to replicate to high titres in L929 cells but infection resulted in reduced cytopathic effect. Our study demonstrates a unique virus-host interaction between NBV and L929 cells, where cells effectively control viral infection/replication and limit the formation of syncytia. By elucidating the molecular mechanisms that control this unique relationship, important insights will be made into the biology of this fusogenic virus. PMID:25748429

  19. Protective Effect of Boric Acid on Oxidative DNA Damage In Chinese Hamster Lung Fibroblast V79 Cell Lines

    Directory of Open Access Journals (Sweden)

    SezenY?lmaz

    2016-02-01

    Full Text Available Objective: Many studies have been published on the antioxidative effects of boric acid (BA and sodium borates in in vitro studies. However, the boron (B concentrations tested in these in vitro studies have not been selected by taking into account the realistic blood B concentrations in humans due to the lack of comprehensive epidemiological studies. The recently published epidemiological studies on B exposure conducted in China and Turkey provided blood B concentrations for both humans in daily life and workers under extreme exposure conditions in occupational setting. The results of these studies have made it possible to test antioxidative effects of BA in in vitro studies within the concentration range relevant to humans. The aim of this study was to investigate the protective effects of BA against oxidative DNA damage in V79 (Chinese hamster lung fibroblast cells. The concentrations of BA tested for its protective effect was selected by taking the blood B concentrations into account reported in previously published epidemiological studies. Therefore, the concentrations of BA tested in this study represent the exposure levels for humans in both daily life and occupational settings. Materials and Methods: In this experimental study, comet assay and neutral red uptake (NRU assay methods were used to determinacy to toxicity and genotoxicity of BA and hydrogen peroxide (H2O2. Results: The results of the NRU assay showed that BA was not cytotoxic within the tested concentrations (3, 10, 30, 100 and 200 ?M. These non-cytotoxic concentrations were used for comet assay. BA pre-treatment significantly reduced (P<0.05, one-way ANOVA the DNA damaging capacity of H2O2 at each tested BA concentrations in V79 cells. Conclusion: Consequently, pre-incubation of V79 cells with BA has significantly reduced the H2O2-induced oxidative DNA damage in V79 cells. The protective effect of BA against oxidative DNA damage in V79 cells at 5, 10, 50, 100 and 200 ?M (54, 108, 540, 1080, and 2161 ng/ml B equivalents concentrations was proved in this in vitro study.

  20. Abnormal response to DNA crosslinking agents of Fanconi anemia fibroblasts can be corrected by transfection with normal human DNA.

    OpenAIRE

    Diatloff-Zito, C; Papadopoulo, D.; Averbeck, D; Moustacchi, E.

    1986-01-01

    Primary skin fibroblast cell lines from patients with Fanconi anemia were cotransfected with UV-irradiated pSV2neo plasmids and high molecular weight DNA from normal human cells. Restoration of a normal cellular resistance to mitomycin C (MMC) was observed provided that a Fanconi anemia cell line is selected for DNA-mediated transformation (neo gene) and that at least two successive rounds of transfection are performed. Cells were selected by taking advantage of the higher proliferation rate ...

  1. Cytotoxic and mutagenic effects of carcinogenic aromatic amides and polycyclic hydrocarbons and ultraviolet irradiation in normally repairing and repair-deficient (xeroderma pigmentosum) diploid human skin fibroblasts

    International Nuclear Information System (INIS)

    The cloning ability of fibroblasts taken from a xeroderma pigmentosum patient proved 2.5 to 3.5 times more sensitive to the cytotoxic effect of active derivatives of carcinogens or to uv irradiation than that of normal cells. They also exhibited a corresponding 2.5- to 3.5-fold greater increase in the frequency of induced mutations to 8-azaguanine resistance per survivor, which might have been expected since these XP cells exhibit less than 20 percent of the DNA-repairing capacity of the normal cells following exposure to such DNA-damaging agents

  2. Three-dimensional culture environment increases the efficacy of platelet rich plasma releasate in prompting skin fibroblast differentiation and extracellular matrix formation.

    Science.gov (United States)

    Rothan, Hussin A; Djordjevic, Ivan; Bahrani, Hirbod; Paydar, Mohammadjavad; Ibrahim, Fatimah; Abd Rahmanh, Noorsaadah; Yusof, Rohana

    2014-01-01

    Platelet rich plasma clot- releasate (PRCR) shows significant influence on tissue regeneration in clinical trials. Although, the mechanism of PRCR effect on fibroblast differentiation has been studied on 2D culture system, a detailed investigation is needed to establish the role of PRCR in cell seeded in 3D scaffolds. Therefore, a study was conducted to evaluate the influence of PRCR in fibroblasts (DFB) differentiation and extracellular matrix formation on both 3D and 2D culture systems. Cell viability was measured using MTT assay and DFB differentiation was evaluated by determining the expression levels of nucleostamin and alpha smooth muscle actin (?-SMA), using indirect immunostaining and Western blotting. The expression levels of extracellular matrix genes (collagen-I, collagen-III, fibronectin and laminin) and focal adhesion formation gene (integrin beta-1) were measured using Real-time PCR. The PRCR at 10% showed significant effect on cells viability compared with 5% and 20% in both culture environments. The decrease in the expression levels of nucleostamin and the increase in ?-SMA signify the DFB differentiation to myofibroblast-like cells that was prominently greater in 3D compared to 2D culture. In 3D culture systems, the total collage production, expression levels of the extracellular matrix gene and the focal adhesion gene were increased significantly compared to 2D culture. In conclusion, 3D culture environments enhances the proliferative and differentiation effects of PRCR on DFB, thereby potentially increases the efficacy of DFB for future tissue engineering clinical application. PMID:25136258

  3. Evaluation of the effect of laser radiation on fibroblast proliferation in repair of skin wounds of rats with iron deficiency anemia

    Science.gov (United States)

    DeCastro, Isabele C. V.; Oliveira-Sampaio, Susana C. P.; Monteiro, Juliana S. de C.; Ferreira, Maria de Fátima L.; Cangussu, Maria T.; N. dos Santos, Jean; Pinheiro, Antonio Luiz B.

    2011-03-01

    The aim of this study was to assess the effect of low- level laser therapy (LLLT) on fibroblast proliferation on wound repair of rats with Iron deficiency anemia since there is no reports on literature about this subject. Iron deficiency anemia was induced on 36 newborn rats then an excisional wound was created on the dorsum of the animals which were divided into four groups: (I) - non-anemic, (II) - Anemic, (III) - non-anemic + LLLT, (IV) Anemic+ LLLT. The animals in each group were sacrificed at 7, 14 and 21 days. Laser irradiation was performed on each group (λ660nm,40Mw,CW) by contact mode with a dose of 2,5J/ cm2 in four points on the area of the wound and total of 10J/cm2 per session. Data were evaluated by analysis of variance (ANOVA) followed by Paired t-test. The results showed LLLT was able to stimulate fibroblastic proliferation in rats with iron deficiency anemia at the 21st day while at control group (III) no statistically significant differences was found.

  4. Application of collagen-chitosan/fibrin glue asymmetric scaffolds in skin tissue engineering*

    OpenAIRE

    Han, Chun-mao; Zhang, Li-Ping; Sun, Jin-zhang; Shi, Hai-Fei; Zhou, Jie; Gao, Chang-you

    2010-01-01

    To create a scaffold that is suitable for the construction of tissue-engineered skin, a novel asymmetric porous scaffold with different pore sizes on either side was prepared by combining a collagen-chitosan porous membrane with fibrin glue. Tissue-engineered skin was fabricated using this asymmetric scaffold, fibroblasts, and a human keratinocyte line (HaCaT). Epidermal cells could be seen growing easily and achieved confluence on the fibrin glue on the upper surface of the scaffold. Scannin...

  5. Tropoelastin regulates chemokine expression in fibroblasts in Costello syndrome

    International Nuclear Information System (INIS)

    Costello syndrome is a multiple congenital anomaly associated with growth and mental retardation, cardiac and skeletal anomalies, and a predisposition to develop neoplasia. Comprehensive expression analysis revealed remarkable up-regulation of several cytokines and chemokines including Gro family proteins, interleukin-1β (IL-1β), IL-8 and MCP-1 but down-regulation of extracellular matrix components including collagens and proteoglycans of skin fibroblasts derived from a Japanese Costello syndrome patient characterized by significantly reduced tropoelastin mRNA, impaired elastogenesis and enhanced cell proliferation. In contrast, decreases in these chemokines and IL-1β expression were observed in Costello fibroblastic cell lines stably expressing the bovine tropoelastin (btEln) gene and in restored elastic fibers. These results strongly suggest that the human TE gene (ELN) transfer could be applicable for the gene therapy of a group of Costello syndrome patients with reduced ELN gene expression

  6. Potentiation by caffeine of x-ray damage to cultured human skin fibroblasts from normal subjects and ataxia-telangiectasia patients

    International Nuclear Information System (INIS)

    Caffeine was found to potentiate x-ray-induced killing of human diploid fibroblasts from a normal subject and an ataxia-telangiectasia (AT) patient when it was present at 2 mM concentration for 30 to 66 h postirradiation. The dose-modifying factor for caffeine-treated normal cells had an average value of 1.26 +- 0.13 which did not vary significantly with treatment time or x-ray dose. The dose-modifying factor for caffeine-treated AT cells was 1.12 +- 0.12 at 30 h, rose to 1.66 +- 0.17 at 41 h, and decreased to 1.31 +- 0.13 at 66 h. Thus no clear difference was observed between these two cell strains' susceptibility to postirradiation caffeine treatment

  7. The potentiation by caffeine of X-ray damage to cultured human skin fibroblasts from normal subjects and ataxia-telangiectasia patients

    International Nuclear Information System (INIS)

    Caffeine was found to potentiate X-ray-induced killing of human diploid fibroblasts from a normal subject and an ataxia-telangiectasia (AT) patient when it was present at 2 mM concentration for 30 to 66 hr postirradiation. The dose-modifying factor for caffeine-treated normal cells had an average value of 1.26 +/- 0.13 which did not vary significantly with treatment time or X-ray dose. The dose-modifying factor for caffeine-treated AT cells was 1.12 +/- 0.12 at 30 hr, rose to 1.66 +/- 0.17 at 41 hr, and decreased to 1.31 +/- 0.13 at 66 hr. Thus no clear difference was observed between these two cell strains' susceptibility to postirradiation caffeine treatment

  8. Microprobe analysis of human fibroblasts

    International Nuclear Information System (INIS)

    The Melbourne Proton Microprobe has been used to study the copper content in human skin fibroblast cells derived from patients with the genetic disease Menkes Syndrome. Both normal and diseased cells have been studied to investigate any elemental differences occurring between the two cell types. This paper details the preparatory techniques necessary for individual cell analysis and presents the elemental information with a new three dimensional contour mapping technique. These maps are used to highlight elemental differences between normal and mutant fibroblasts. The work also confirms the expected copper excess found in the Menkes cell and indicates that the microprobe can be used for rapid identification of a Menkes carrier

  9. Granulocyte macrophage colony stimulating factor (GM-CSF biological actions on human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    S Montagnani

    2009-12-01

    Full Text Available Fibroblasts are involved in all pathologies characterized by increased ExtraCellularMatrix synthesis, from wound healing to fibrosis. Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF is a cytokine isolated as an hemopoietic growth factor but recently indicated as a differentiative agent on endothelial cells. In this work we demonstrated the expression of the receptor for GM-CSF (GMCSFR on human normal skin fibroblasts from healthy subjects (NFPC and on a human normal fibroblast cell line (NHDF and we try to investigate the biological effects of this cytokine. Human normal fibroblasts were cultured with different doses of GM-CSF to study the effects of this factor on GMCSFR expression, on cell proliferation and adhesion structures. In addition we studied the production of some Extra-Cellular Matrix (ECM components such as Fibronectin, Tenascin and Collagen I. The growth rate of fibroblasts from healthy donors (NFPC is not augmented by GM-CSF stimulation in spite of increased expression of the GM-CSFR. On the contrary, the proliferation of normal human dermal fibroblasts (NHDF cell line seems more influenced by high concentration of GM-CSF in the culture medium. The adhesion structures and the ECM components appear variously influenced by GM-CSF treatment as compared to fibroblasts cultured in basal condition, but newly only NHDF cells are really induced to increase their synthesis activity. We suggest that the in vitro treatment with GM-CSF can shift human normal fibroblasts towards a more differentiated state, due or accompanied by an increased expression of GM-CSFR and that such “differentiation” is an important event induced by such cytokine.

  10. Deterministic late effects of skin irradiation: in vitro studies of experimental radiation fibrosis

    International Nuclear Information System (INIS)

    After radiotherapy or radiation accidents, large doses of gamma irradiation can induce necrosis and fibrosis, and secondary cancers are occasionally observed. The identity of the target cells in such irradiated tissues is still controversial. As the role of the fibroblast in wound repair after acute irradiation is not well documented, this role was investigated in an experimental model. Pigs were gamma irradiated with an 192Ir source on the thigh, which resulted in necrosis and then fibrosis of the skin and muscular tissues. In primary cultures, fibroblasts isolated from the fibrotic tissue exhibited an activated phenotype for up to two years after irradiation. They had a myofibroblastic morphology and their proliferation was activated. The fibroblasts synthesised an extracellular matrix similar to that of immature scars or developing tissues. Most fibroblasts carried numerous chromosome anomalies. In long-term cultures, fibrotic fibroblasts gave rise to established cell lines, in which the morphology of the cells resembled that of transformed cells. Such highly modified cells observed in vitro might be related to the appearance of secondary sarcoma in vivo. We conclude from these results that the fibroblast is clearly an important target cell in irradiated skin. The difficult healing of late radiation damage could be related to chronic inflammation and to the long-term activation of abnormal fibroblasts in irradiated tissues. (author)

  11. Defective glycosylation of decorin and biglycan, altered collagen structure, and abnormal phenotype of the skin fibroblasts of an Ehlers-Danlos syndrome patient carrying the novel Arg270Cys substitution in galactosyltransferase I (beta4GalT-7).

    Science.gov (United States)

    Seidler, Daniela G; Faiyaz-Ul-Haque, Muhammad; Hansen, Uwe; Yip, George W; Zaidi, Syed H E; Teebi, Ahmad S; Kiesel, Ludwig; Götte, Martin

    2006-07-01

    The Ehlers-Danlos syndrome (EDS) is a heterogeneous group of connective tissue disorders affecting skin and joint function. Molecular defects in extracellular matrix proteins, including collagen (type I, III, and V) and tenascin X are associated with different forms of EDS. Compound heterozygous mutations in the B4GALT7 gene, resulting in aberrant glycosylation of the dermatan sulfate proteoglycan decorin, had been described in a single patient affected with the progeroid form of EDS. We have studied the molecular phenotype of decorin, biglycan, and collagen type I containing fibrils in skin fibroblasts of a patient carrying the novel homozygous C808T point mutation in the B4GALT7 gene, which causes an Arg270Cys substitution in beta4GalT-7. Compared to control fibroblasts, galactosyltransferase activity in beta4GalT-7(Arg270Cys) cells was approximately three times reduced over a temperature range of 25-41 degrees C. Pulse-chase experiments and confocal microscopy demonstrated that synthesis and secretion of decorin were normal in beta4GalT-7(Arg270Cys) cells. However, about 50% of decorin were synthesized as a protein core in addition to its proteoglycan form. Biglycan was found in a monoglycanated form in addition to its mature form. Glycosaminoglycan chains were of the dermatan/chondroitin sulfate type both in beta4GalT-7(Arg270Cys) and control cells, and epimerization was reduced for decorin and biglycan. Compared to control cells, beta4GalT-7(Arg270Cys) cells showed altered, highly spread or stretched phenotypes and decreased proliferation rates. At the ultrastructural level, an intracellular accumulation of multiple secondary lysosomes and degenerative vacuoles was seen in beta4GalT-7(Arg270Cys) cells. Furthermore, the collagen suprastructures were altered in the beta4GalT-7(Arg270Cys) cells. The reduced beta4GalT-7 activity resulting in defective glycosylation of decorin and biglycan may be responsible for the complex molecular pathology in beta4GalT-7 deficient EDS patients, given the role of these proteoglycans in bone formation, collagen fibrillogenesis, and skeletal muscle development. PMID:16583246

  12. Prolidase-dependent mechanism of (Z)-8,9-epoxyheptadeca-1,11,14-triene-induced inhibition of collagen biosynthesis in cultured human skin fibroblasts.

    Science.gov (United States)

    Szoka, Lukasz; Karna, Ewa; Nazaruk, Jolanta; Palka, Jerzy A

    2016-03-01

    The effects of polyolefinic compound from roots of Cirsium palustre, (Z)-8,9-epoxyheptadeca-1,11,14-triene (EHT) on collagen biosynthesis, prolidase activity, expression of insulin-like growth factor receptor (IGF-IR), ?1 integrin, MAP kinases (pERK1/2), the transcription factors such as nuclear factor kappa B (NF-?B) and hypoxia-inducible factor-1? (HIF-1?) were evaluated in human dermal fibroblasts treated with micromolar concentrations (40-200?M) for 24h. It was found that EHT-dependent inhibition of collagen biosynthesis was accompanied by parallel inhibition in prolidase activity. Since IGF-I is the most potent regulator of both processes and prolidase is regulated by ?1 integrin signalling, the effect of EHT on IGF-IR and ?1 integrin receptor expressions were evaluated. Exposure of the cells to EHT contributed to distinct increase in IGF-IR and slight increase in ?1 integrin receptor expressions. It was accompanied by decrease in expression of pERK1/2, HIF-1? and NF-?B. EHT-dependent inhibition of collagen biosynthesis results from inhibition of prolidase activity, the enzyme involved in collagen biosynthesis. PMID:25982243

  13. Fibroblast cultures in duchenne muscular dystrophy

    International Nuclear Information System (INIS)

    Primary skin fibroblast cultures were grown from forearm pinch skin biopsies obtained from 24 patients with Duchenne muscular dystrophy (DMD) and ten normal controls matched for sex and age. The first subcultures were grown for 7 days and incubated with L-(3H)-proline for 24 hours. Intracellular collagen incoption was significantly decreased (2.2 X) and extracellular collagen incorporation significantly increased (1.8 X) in fibroblast cultures from patients with DMD by both collagenase assay and polyacrylamide gel electrophoresis. The synthesis of noncollagen proteins showed low values from the DMD fibroblast cultures. The alterations in synthesis and secretion of collagen and noncollagen proteins were characteristic only for the log phase of DMD fibroblasts. (author)

  14. Mitogenic activity of fetal bovine serum, fish fry extract, insulin-like growth factor-I, and fibroblast growth factor on brown bullhead catfish cells - BB line

    Directory of Open Access Journals (Sweden)

    CYRINO J. E. P.

    1999-01-01

    Full Text Available Biossays were performed to assess the effects of different levels of growth medium supplementation with fetal bovine serum (FBS, fish fry extract (FE, combinations of FBS and FE, and addition of insulin-like growth factor I (IGF-I and fibroblast growth factor (FGF on the proliferation of brown bullhead catfish cells (BB line. Treatments (n = 4 were: 2.5, 5, 10, and 15.0% FBS or FE and 5/2.5, 5/5, 10/2.5, and 10/5 of a FBS/FE combination as supplement to the growth medium, or the addition of 0.1, 1, 2.5, 10, 25, and 75 ng/ml of either IGF-I or FGF to the growth media. Initial cell density was 1.1 x 10(6 cells per well on uncoated 24-well plates. Incubation temperature was 29.5 0.7C. Six hours after plating, initial culture medium was removed, plates rinsed with Dulbecco?s phosphate buffered saline, treatment media added, and cells allowed to proliferate for 24 hours. Another bioassay was performed with rat myoblast omega cells (RMo using the same levels of growth medium supplemented with FBS, FE and FBS/FE. Base growth medium was Dulbecco?s MEM. The initial cell density was 7.2 x 10(6 cells per well, and the bioassay was carried out at 36.0 0.5C, on a 95% air, 5% CO2 incubator. Increasing levels of FBS had a positive effect (P < 0.05 on the proliferation of both BB and RMo cells. Increasing levels of FE had a negative effect (P < 0.05 on the proliferation of BB cells and totally inhibited the proliferation of RMo cells at any level of supplementation. Higher levels of FE on the FBS/FE combinations presented a negative effect on the proliferation of both BB and RMo cells (P < 0.05. Insulin-like growth factor I had a positive quadratic effect (P < 0.05 on the proliferation of BB cells. Apparently, mammalian growth factors slightly stimulated mitogenic activity in fish cells, while FE contained factors which inhibited the mitogenic activity of RMo and BB cell lines.

  15. Comparative study on cytotoxicity effect of biological and commercial synthesized nanosilver on human gastric carcinoma and normal lung fibroblast cell lines

    Directory of Open Access Journals (Sweden)

    Mohammad Ali Rashmezad

    2015-03-01

    Full Text Available Background: Biosynthesis of nanoparticles has attracted the attention of the scientific community in nanotechnology and biotechnology due to their extensive application in the area of material sciences and medicine. Nowadays, despite a various application of nanomaterial’s, there is a little information about their impact on human health. In this study, we investigated the comparative study on cytotoxicity effect of biological and commercial synthesized nanosilver on human gastric carcinoma (AGS and normal lung fibroblast (MRC-5 cell lines. Methods: The current experimental study was carried out in Islamic Azad University, East Tehran Branch, from April to November 2014. The biological synthesis of nanosilver was obtained from Eucalyptus plant extract as a reducing agent. Further to more analysis, morphological study on size and shape of developed biological nanosilver was characterized by performing scanning electron microscopy and dynamic light scattering. AGS and MCR-5 cell lines were treated with various concentration of nanosilver for 24, 48 and 72 hours. Finally, the cell viability was evaluated by using MTT assay. Results: The results show that the nanosilver exerts a dose-dependent inhibitory effect on viability of cells. At 100µg/mL of commercial and biological synthesized nanosilver, the viability of AGS was reduced to 7.47±0.002% (P=0.002 and 3.65±0.01% (P=0.003 after 72 hours, respectively. In addition, the viability of MRC-5 at the same condition was reduced to 10.27±0.19% (P=0.001 and 9.16±1.53% (P=0.002, respectively. Conclusion: Based on a thorough literature surveys, the present study is the first research about biosynthesis of nanosilver using Eucalyptus plant extract. This eco-friendly and cost effective method can be used for large scale production of silver nanoparticle. In addition, based on the current obtained data, commercial and biological synthesized nanosilver can more inhibitory effect on cancer cells compared to the normal cells. Hence, silver nanoparticles might be used as a new strategy for treating many human cancers. However, further studies are necessary to ascertain their potential as anticancer agents.

  16. Use of metabolic inhibitors to compare the excision repair of pyrimidine dimers and nondimer DNA damages in human skin fibroblasts exposed to 254-nm and sunlamp-produced > 310-nm ultraviolet radiation

    International Nuclear Information System (INIS)

    Normal human skin fibroblasts were exposed to either 0-5 J/m2 of 254-nm ultraviolet (UV) radiation or 0-50 kJ/m2 of the Mylar-filtered UV (>310 nm) produced by a fluorescent sunlamp. These cells were then incubated for 0-20 min in medium containing 10 mM hydroxyurea (HU) and 0.1 mM 1-?-D-arabinofuranosyl cytosine (ara C), and the yield of DNA strand breaks was measured by means of the alkaline elution technique. For cells irradiated with 254-nm UV, which results primarily in the formation of cyclobutane pyrimidine dimers, a rapid increase in DNA strand breaks was detected following incubation with these metabolic inhibitors. In contrast, only a low level of strand breaks formed in cells incubated with HU and ara C after irradiation with approximately equitoxic fluences of sunlamp UV >310 nm, which mainly causes the induction of nondimer DNA lesions. Hence, these results are consistent with the conclusion that the pathways involved in the repair of nondimer DNA damages induced by UV wavelengths >310 nm differ from the repair of pyrimidine dimers

  17. Performance of line-scanning confocal microscopy in human skin: investigation of potential for clinical translation

    Science.gov (United States)

    Larson, Bjorg; Peterson, Gary; Abeytunge, Sanjeewa; Rajadhyaksha, Milind

    2011-03-01

    Line-scanning, using 8-10 optical components, linear-array detectors and custom-FPGA electronics, may enable smaller, simpler and lower-cost confocal microscopes to accelerate translation to the clinic. The adaptability of commercially available low-cost array detectors for confocal microscopy is being investigated. Measurements of optical sectioning and lateral resolution showed good agreement with theory, and are comparable to that of point-scanning systems. LSFs through full thickness of human epidermis show a two-fold degradation in sectioning performance. Imaging of human epidermis in vivo demonstrates nuclear and cellular detail down to the basal layer with a bench top setup and also a compact clinical prototype. Blood flow in oral mucosa can be imaged using the clinical prototype. However, speckle and background noise degrade contrast and resolution of the image.

  18. A Comparison between the Colony Formation of Adult Mouse Spermatogonial Stem Cells in Co cultures with Sertoli and STO (Mouse Embryonic Fibroblast Cell Line

    Directory of Open Access Journals (Sweden)

    Seyed Morteza Koruji

    2010-01-01

    Full Text Available Objective: The aim of this study was to compare the colony formation of spermatogonialstem cells (SSCs on sertoli and STO (Mouse embryonic fibroblast cell line feeder celllayers during a two-week period.Materials and Methods: Initially, sertoli cells and SSCs were isolated from adultmouse testes using a two-step enzymatic digestion and lectin immobilization. Characteristicsof the isolated cells were immunocytochemically confirmed by examiningfor the presence of Oct-4, CDH1, promyelocytic leukaemia zinc finger factor (PLZF,SSC C-kit, and the distribution of Sertoli cell vimentin. SSCs were then cultured abovethe Sertoli, STO and the control (without co-culture separately for two weeks. In allthree groups, the number and diameter of colonies were evaluated using an invert microscopeon the 3rd, 7th, 10th and 14th day. ?1 and ?6-integrin m-RNA expressions wereassessed using a reverse transcription polymerase chain reaction (RT-PCR and realtimePCR. Furthermore, Oct-4 m RNA expression was assessed using real time PCR.Statistical analysis was performed using ANOVA; and the paired two-sample t test andTukeys test were used as post-hoc tests for the data analysis of the three sertoli, STOand control cocultures.Results: At the four specified time points, our results showed significant differences (p<0.05in colony numbers and diameters among the sertoli, STO and control groups. The numberand diameter of colonies increased more rapidly in the sertoli coculture than in the othertwo Our results at all four time points also showed significant differences (p<0.05 in themean colony numbers and diameters between the three groups, with the Sertoli coculturehaving the highest mean values for colony numbers and diameters. The RT-PCR results,after two-weeks of culturing, showed that ?1-integrin was expressed in all three groups cocultures,but ?6-integrin was not expressed. Additionally, based on real time PCR results,the three genes (?1-integrin, ?6-integrin, Oct-4 mentioned were also expressed in all threeco cultures groups.Conclusion: Based on the optimal effects of sertoli feeder cells on spermatogonial stemcells in a co culture system, as also confirmed by several other studies, their use is suggestedto achieve better colonization of SSCs.

  19. Application of Allogeneic Fibroblast Cells in Cellular Therapy of Recessive Dystrophic Epidermolysis Bullosa

    Directory of Open Access Journals (Sweden)

    Zare

    2015-09-01

    Full Text Available Context Connective tissue cells include fibroblasts, chondrocytes, adipocyte, and osteocytes. These cells are specialized for the secretion of collagenous extracellular matrix and are responsible for the architectural framework of the human body. Evidence Acquisition Connective tissue cells play a central role in supporting as well as repairing tissues and organs. Fibroblast cell therapy could be used for the treatment of burn wounds, scars, diabetic foot ulcers, acne scars and skin aging. This review focused on biology of fibroblasts and their role in cell therapy of recessive dystrophic epidermolysis bullosa (RDEB. Results Fibroblasts are known to play a pivotal role in skin structure and integrity, and dermal fibroblasts are believed to promote skin regeneration and rejuvenation via collagen production. Conclusions Fibroblasts can be used in transplantations to ameliorate an immune system response, in order to reduce antigen production. Human fibroblasts suppress ongoing mixed lymphocyte reactions (MLRs between lymphocyte cells from two individuals, and supernatant materials from fibroblast cultures suppress MLRs.

  20. Platelet gel stimulates proliferation of human dermal fibroblasts in vitro:

    OpenAIRE

    Domanovič, Dragoslav; Jeras, Matjaž; Krašna, Metka; Švajger, Urban; Tomšič, Ana

    2007-01-01

    Background: Platelet gel is a source of many soluble proteins that can modulate tissue regeneration. Dermal fibroblasts play an important role in tissue remodeling and wound healing. Objective: The study was performed to investigate the influence of platelet gel and its active ingredients on the proliferation of human dermal fibroblasts in vitro. Methods: A fibroblast culture was established from a normal human skin biopsy. Platelet gel was prepared from platelet-rich plasma (PRP) following t...

  1. Scorbutic and fasted guinea pig sera contain an insulin-like growth factor I-reversible inhibitor of proteoglycan and collagen synthesis in chick embryo chondrocytes and adult human skin fibroblasts.

    Science.gov (United States)

    Oyamada, I; Palka, J; Schalk, E M; Takeda, K; Peterkofsky, B

    1990-01-01

    Chick embryo chondrocytes cultured in sera from scorbutic and fasted guinea pigs exhibited decreases in collagen and proteoglycan production to about 30-50% of control values (I. Oyamada et al., 1988, Biochem. Biophys. Res. Commun. 152, 1490-1496). Here we show by pulse-chase labeling experiments that in the chondrocyte system, as in the cartilage of scorbutic and fasted guinea pigs, decreased incorporation of precursor into collagen was due to decreased synthesis rather than to increased degradation. There was a concomitant decrease in type II procollagen mRNA to about 32% of the control level. As in scorbutic cartilage, proteoglycan synthesis by chondrocytes in scorbutic serum was blocked at the stage of glycosaminoglycan chain initiation. Scorbutic and fasted guinea pig sera also caused a 50-60% decrease in the rates of collagen and proteoglycan synthesis in adult human skin fibroblasts, which synthesize mainly type I collagen. Decreased matrix synthesis in both cell types resulted from the presence of an inhibitor in scorbutic and fasted sera. Elevated cortisol levels in these sera were not responsible for inhibition, as determined by the addition of dexamethasone to chondrocytes cultured in normal serum. Insulin-like growth factor I (IGF-I, 300-350 ng/ml) reversed the inhibition of extracellular matrix synthesis by scorbutic and fasted guinea pig sera in both cell types and prevented the decrease in type II procollagen mRNA in chondrocytes. Therefore, in addition to its established role in proteoglycan metabolism, IGF-I also regulates the synthesis of several collagen types. An increase in the circulating inhibitor of IGF-I action thus could lead to the negative regulation of collagen and cartilage proteoglycan synthesis that occurs in ascorbate-deficient and fasted guinea pigs. PMID:2297232

  2. Ultraviolet exposure of melanoma cells induces fibroblast activation protein-alpha in fibroblasts: Implications for melanoma invasion

    OpenAIRE

    Wster, Petra; Rosdahl, Inger; Gilmore, Brendan F.; Seifert, Oliver

    2011-01-01

    Fibroblast activation protein-alpha (FAP-alpha) promotes tumor growth and cell invasiveness through extracellular matrix degradation. How ultraviolet radiation (UVR), the major risk factor for malignant melanoma, influences the expression of FAP-alpha is unknown. We examined the effect of UVR on FAP-alpha expression in melanocytes, keratinocytes and fibroblasts from the skin and in melanoma cells. UVR induces upregulation of FAP-alpha in fibroblasts, melanocytes and primary melanoma cells (PM...

  3. Parental somatic and germ-line mosaicism for a FBN2 mutation and analysis of FBN2 transcript levels in dermal fibroblasts.

    OpenAIRE

    Putnam, E A; Park, E S; Aalfs, C.M.; Hennekam, R.C.; Milewicz, D. M.

    1997-01-01

    Congenital contractural arachnodactyly (CCA) is an autosomal dominant disorder that is phenotypically related to the Marfan syndrome. CCA has recently been shown to result from mutations in the FBN2 gene, which encodes an elastin-associated microfibrillar protein called fibrillin-2. Two siblings are reported here with classic manifestations of CCA with unaffected parents. Analysis of the FBN2 cDNA from dermal fibroblasts from one of the affected siblings revealed a heterozygous exon splicing ...

  4. In vitro gene manipulation of spinal muscular atrophy fibroblast cell line using gene-targeting fragment for restoration of SMN protein expression.

    Science.gov (United States)

    Rashnonejad, A; Gndz, C; Ssler, S Y; Onay, H; Durmaz, B; Bandehpour, M; zk?nay, F

    2016-01-01

    The reduced level of survival motor neuron (SMN) protein, caused by homozygous deletions in the SMN gene, led to a common neurodegenerative disorder known as spinal muscular atrophy (SMA). In spite of extensive efforts to find a cure for SMA, there is currently no effective treatment available for this devastating disease. In this study, restoration of SMN expression through 'gene-targeting' method in SMA fibroblast cells was attempted. We designed a 2697-bp gene-targeting cassette; it consisted of an SMN1 open reading frame expressing 38?kD SMN protein and the upstream and downstream regions of exon 1 of SMN1 gene at the ends as the homology arms. SMA fibroblast cells were transfected by gene-targeting cassette using Lipofectamine LTX-PLUS reagent. Occurrence of homologous recombination in selected cells was investigated by PCR analysis. Increased expression of SMN protein was shown by real-time PCR and western blotting analysis. The immunofluorescence analysis results demonstrated that the number of SMN nuclear structures, Gems, was the same as or greater than the number of Gems found in normal fibroblasts. The results of this study indicate that gene-targeting methods do, in fact, present as an alternative for restoration of SMN expression in SMA patients-derived cells in vitro. PMID:26331341

  5. Preparation of Proper Immunogen by Cloning and Stable Expression of cDNA coding for Human Hematopoietic Stem Cell Marker CD34 in NIH-3T3 Mouse Fibroblast Cell Line

    Directory of Open Access Journals (Sweden)

    Farzaneh Shafaghat

    2015-03-01

    Full Text Available Purpose: Transmembrane CD34 glycoprotein is the most important marker for identification, isolation and enumeration of hematopoietic stem cells (HSCs. We aimed in this study to clone the cDNA coding for human CD34 from KG1a cell line and stably express in mouse fibroblast cell line NIH-3T3. Such artificial cell line could be useful as proper immunogen for production of mouse monoclonal antibodies. Methods: CD34 cDNA was cloned from KG1a cell line after total RNA extraction and cDNA synthesis. Pfu DNA polymerase-amplified specific band was ligated to pGEMT-easy TA-cloning vector and sub-cloned in pCMV6-Neo expression vector. After transfection of NIH-3T3 cells using 3 ?g of recombinant construct and 6 ?l of JetPEI transfection reagent, stable expression was obtained by selection of cells by G418 antibiotic and confirmed by surface flow cytometry. Results: 1158 bp specific band was aligned completely to reference sequence in NCBI database corresponding to long isoform of human CD34. Transient and stable expression of human CD34 on transfected NIH-3T3 mouse fibroblast cells was achieved (25% and 95%, respectively as shown by flow cytometry. Conclusion: Cloning and stable expression of human CD34 cDNA was successfully performed and validated by standard flow cytometric analysis. Due to murine origin of NIH-3T3 cell line, CD34-expressing NIH-3T3 cells could be useful as immunogen in production of diagnostic monoclonal antibodies against human CD34. This approach could bypass the need for purification of recombinant proteins produced in eukaryotic expression systems.

  6. Skin pigmentary anomalies and mosaicism for an acentric marker chromosome originating from 3q

    OpenAIRE

    Portnoi, M.; Boutchnei, S.; Bouscarat, F.; Morlier, G.; Nizard, S.; Dersarkissian, H.; Crickx, B.; Nouchy, M.; Taillemite, J.; Belaich, S.

    1999-01-01

    We report on a 22 year old man with hyperpigmentation distributed along the lines of Blaschko in whom cytogenetic analysis showed mosaicism for an unusual supernumerary marker chromosome. The patient was of normal intelligence and was not dysmorphic. The marker was present in 30% of his lymphocytes and in 6% of his skin fibroblasts from a dark area, while fibroblasts from a light area showed a normal karyotype, 46,XY.We have identified the origin of the marker using fluorescence in situ hybri...

  7. Role of oxidative stress in ERK and p38 MAPK activation induced by the chemical sensitizer DNFB in a fetal skin dendritic cell line

    OpenAIRE

    Matos, MT; Duarte, CB; Gonçalo, Margarida; Lopes, Mc

    2005-01-01

    The intracellular mechanisms involved in the early phase of dendritic cell (DC) activation upon contact with chemical sensitizers are not well known. The strong skin sensitizer 2,4-dinitrofluorobenzene (DNFB) was shown to induce the activation of mitogen-activated protein kinases (MAPK) in DC. In the present study, we investigated a putative role for oxidative stress in DNFB-induced MAPK activation and upregulation of the costimulatory molecule CD40. In a DC line generated from fetal mouse sk...

  8. Differentiation of human labia minora dermis-derived fibroblasts into insulin-producing cells

    OpenAIRE

    Kim, Bona; Yoon, Byung Sun; Moon, Jai-Hee; Kim, Jonggun; Jun, Eun Kyoung; Lee, Jung Han; Kim, Jun Sung; Baik, Cheong Soon; Kim, AeRee; Whang, Kwang Youn; You, Seungkwon

    2011-01-01

    Recent evidence has suggested that human skin fibroblasts may represent a novel source of therapeutic stem cells. In this study, we report a 3-stage method to induce the differentiation of skin fibroblasts into insulin-producing cells (IPCs). In stage 1, we establish the isolation, expansion and characterization of mesenchymal stem cells from human labia minora dermis-derived fibroblasts (hLMDFs) (stage 1: MSC expansion). hLMDFs express the typical mesenchymal stem cell marker proteins and ca...

  9. Abnormal proliferation and aging of cultured fibroblasts from pigs with subcutaneous fibrosis induced by gamma irradiation

    International Nuclear Information System (INIS)

    In vivo, fibrotic disorders, which may be due either to injury or disease, are characterized by overproliferation of fibroblasts and overproduction of connective tissue. In vitro, however, most of the fibrotic cell lines studied exhibited no differences in growth potential compared with control cell lines derived from normal skin. In the present study, we investigated the in vitro behavior of fibroblasts derived from pigs with subcutaneous fibrosis induced by gamma irradiation. The cells were isolated from the scar tissue six to 20 months after irradiation. In primary cultures, the cells derived from the fibrotic lesions exhibited greater attachment efficiency and faster proliferation than those of the cells derived from the normal skin of the same animal. In long-term cultures, the differences between normal and fibrotic cells were still greater: the normal skin cells underwent 17 population doublings and then died, whereas the fibrotic cells exhibited a prolonged life span, and were still actively proliferating after 80 population doublings. Cell morphology and the number of chromosomes were modified throughout subcultures. These results imply that in the scar tissue active fibrotic cell proliferation continued for years after irradiation and that this activation was expressed in vitro. Therefore, in this pig fibrosis model, the data acquired in the present in vitro studies closely resemble that obtained from earlier in vivo observations

  10. Effects of Ginger Phenylpropanoids and Quercetin on Nrf2-ARE Pathway in Human BJ Fibroblasts and HaCaT Keratinocytes

    OpenAIRE

    Schadich, Ermin; Hlav?, Jan; Voln, Tereza; Varanasi, Lakshman; Hajdch, Marin; Dubk, Petr

    2016-01-01

    Quercetin and phenylpropanoids are well known chemoprotective compounds identified in many plants. This study was aimed at determining their effects on activation of Nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant response element (Nrf2-ARE) signalling pathway and expression of its important downstream effector phase II detoxification enzyme glutathione-S-transferase P1 (GSTP1) in BJ foreskin fibroblasts and skin HaCaT keratinocytes. Cell lines and their corresponding Nrf2-ARE ...

  11. Polycomponent mesotherapy formulations for the treatment of skin aging and improvement of skin quality

    OpenAIRE

    Prikhnenko S

    2015-01-01

    Sergey Prikhnenko Private Practice, Novosibirsk, Russia Abstract: Skin aging can largely be attributed to dermal fibroblast dysfunction and a decrease in their biosynthetic activity. Regardless of the underlying causes, aging fibroblasts begin to produce elements of the extracellular matrix in amounts that are insufficient to maintain the youthful appearance of skin. The goal of mesopreparations is primarily to slow down and correct changes in skin due to aging. The rationale for developing ...

  12. Bioactive Constituents of Zanthoxylum rhetsa Bark and Its Cytotoxic Potential against B16-F10 Melanoma Cancer and Normal Human Dermal Fibroblast (HDF Cell Lines

    Directory of Open Access Journals (Sweden)

    Ramesh Kumar Santhanam

    2016-05-01

    Full Text Available Zanthoxylum rhetsa is an aromatic tree, known vernacularly as “Indian Prickly Ash”. It has been predominantly used by Indian tribes for the treatment of many infirmities like diabetes, inflammation, rheumatism, toothache and diarrhea. In this study, we identified major volatile constituents present in different solvent fractions of Z. rhetsa bark using GC-MS analysis and isolated two tetrahydrofuran lignans (yangambin and kobusin, a berberine alkaloid (columbamine and a triterpenoid (lupeol from the bioactive chloroform fraction. The solvent fractions and purified compounds were tested for their cytotoxic potential against human dermal fibroblasts (HDF and mouse melanoma (B16-F10 cells, using the MTT assay. All the solvent fractions and purified compounds were found to be non-cytotoxic to HDF cells. However, the chloroform fraction and kobusin exhibited cytotoxic effect against B16-F10 melanoma cells. The presence of bioactive lignans and alkaloids were suggested to be responsible for the cytotoxic property of Z. rhetsa bark against B16-F10 cells.

  13. Age-related disruption of autophagy in dermal fibroblasts modulates extracellular matrix components

    Energy Technology Data Exchange (ETDEWEB)

    Tashiro, Kanae [Skin Research Department, POLA Chemical Industries, Inc., Yokohama (Japan); Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Shishido, Mayumi [Skin Research Department, POLA Chemical Industries, Inc., Yokohama (Japan); Fujimoto, Keiko [Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Organelle Homeostasis Research Center, Kyushu University, Fukuoka (Japan); Hirota, Yuko [Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Yo, Kazuyuki; Gomi, Takamasa [Skin Research Department, POLA Chemical Industries, Inc., Yokohama (Japan); Tanaka, Yoshitaka, E-mail: tanakay@bioc.phar.kyushu-u.ac.jp [Division of Pharmaceutical Cell Biology, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka (Japan); Organelle Homeostasis Research Center, Kyushu University, Fukuoka (Japan)

    2014-01-03

    Highlights: •Autophagosomes accumulate in aged dermal fibroblasts. •Autophagic degradation is impaired in aged dermal fibroblasts. •Autophagy disruption affects extracellular matrix components in dermal fibroblasts. -- Abstract: Autophagy is an intracellular degradative system that is believed to be involved in the aging process. The contribution of autophagy to age-related changes in the human skin is unclear. In this study, we examined the relationship between autophagy and skin aging. Transmission electron microscopy and immunofluorescence microscopy analyses of skin tissue and cultured dermal fibroblasts derived from women of different ages revealed an increase in the number of nascent double-membrane autophagosomes with age. Western blot analysis showed that the amount of LC3-II, a form associated with autophagic vacuolar membranes, was significantly increased in aged dermal fibroblasts compared with that in young dermal fibroblasts. Aged dermal fibroblasts were minimally affected by inhibition of autophagic activity. Although lipofuscin autofluorescence was elevated in aged dermal fibroblasts, the expression of Beclin-1 and Atg5—genes essential for autophagosome formation—was similar between young and aged dermal fibroblasts, suggesting that the increase of autophagosomes in aged dermal fibroblasts was due to impaired autophagic flux rather than an increase in autophagosome formation. Treatment of young dermal fibroblasts with lysosomal protease inhibitors, which mimic the condition of aged dermal fibroblasts with reduced autophagic activity, altered the fibroblast content of type I procollagen, hyaluronan and elastin, and caused a breakdown of collagen fibrils. Collectively, these findings suggest that the autophagy pathway is impaired in aged dermal fibroblasts, which leads to deterioration of dermal integrity and skin fragility.

  14. Age-related disruption of autophagy in dermal fibroblasts modulates extracellular matrix components

    International Nuclear Information System (INIS)

    Highlights: •Autophagosomes accumulate in aged dermal fibroblasts. •Autophagic degradation is impaired in aged dermal fibroblasts. •Autophagy disruption affects extracellular matrix components in dermal fibroblasts. -- Abstract: Autophagy is an intracellular degradative system that is believed to be involved in the aging process. The contribution of autophagy to age-related changes in the human skin is unclear. In this study, we examined the relationship between autophagy and skin aging. Transmission electron microscopy and immunofluorescence microscopy analyses of skin tissue and cultured dermal fibroblasts derived from women of different ages revealed an increase in the number of nascent double-membrane autophagosomes with age. Western blot analysis showed that the amount of LC3-II, a form associated with autophagic vacuolar membranes, was significantly increased in aged dermal fibroblasts compared with that in young dermal fibroblasts. Aged dermal fibroblasts were minimally affected by inhibition of autophagic activity. Although lipofuscin autofluorescence was elevated in aged dermal fibroblasts, the expression of Beclin-1 and Atg5—genes essential for autophagosome formation—was similar between young and aged dermal fibroblasts, suggesting that the increase of autophagosomes in aged dermal fibroblasts was due to impaired autophagic flux rather than an increase in autophagosome formation. Treatment of young dermal fibroblasts with lysosomal protease inhibitors, which mimic the condition of aged dermal fibroblasts with reduced autophagic activity, altered the fibroblast content of type I procollagen, hyaluronan and elastin, and caused a breakdown of collagen fibrils. Collectively, these findings suggest that the autophagy pathway is impaired in aged dermal fibroblasts, which leads to deterioration of dermal integrity and skin fragility

  15. Impact of the neutron and nuclear matter equations of state on neutron skin and neutron drip lines in chiral effective field theory

    CERN Document Server

    Sammarruca, Francesca

    2016-01-01

    We present predictions of the binding energy per nucleon and the neutron skin thickness in highly neutron-rich isotopes of Oxygen, Magnesium, and Aluminum. The calculations are carried out at and below the neutron drip line. The nuclear properties are obtained via an energy functional whose input is the equation of state of isospin-asymmetric in?finite matter. The latter is based on a microscopic derivation applying chiral few-nucleon forces. We highlight the impact of the equation of state at diff?erent orders of chiral effective fi?eld theory and discuss the role of three-neutron forces.

  16. Polypeptide structure of a human dermal fibroblast-activating factor (FAF) derived from the U937 cultured line of human monocyte-like cells

    International Nuclear Information System (INIS)

    Six liter batches of 1 x 106 U937 cells/ml of serum-free RPMI medium were incubated with 100 ng/ml of phorbol myristate acetate for 48 hr at 370C in 5% CO2 in air to generate FAFs, as quantified by the stimulation of uptake of [3H]thymidine by quiescent human dermal fibroblasts. Filtration of the supernatants on Sephadex G-75 resolved two FAFs of approximately 40,000 and 10-13,000 daltons. The latter principle was purified to homogeneity by sequential Sephadex G-50 filtration, revealing an apparent m.w. of 7-8000, Mono-Q FPLC anion-exchange chromatography with a linear gradient from 20 mM Tris-HCl (pH 8.3) to 0.5 M NaCl-20 mM Tris-HCl in 30 min, and two cycles of high-performance liquid chromatography (HPLC) on a 300 A pore 10 μm C4 column at 1 ml/min with 0.05% trifluoroacetic acid (TFA) in water to 30:70 (v:v) and then to 60:40 (v:v) acetonitrile: 0.05% TFA linearly in 15 min and 30 min, respectively, The FAF activity eluted from HPLC in a sharp peak of O.D. 215 nm at 45% acetonitrile. Analyses of amino acid composition of the highly purified 7-8000 dalton FAF-U937 revealed 37% hydrophobic, 14% basic, and 21% acidic or amide residues, as well as one tryrosine and one methionine. This U937 cell-derived FAF appears to be a unique acidic polypeptide growth factor

  17. Advances in Skin Substitutes—Potential of Tissue Engineered Skin for Facilitating Anti-Fibrotic Healing

    Directory of Open Access Journals (Sweden)

    Mathew Varkey

    2015-07-01

    Full Text Available Skin protects the body from exogenous substances and functions as a barrier to fluid loss and trauma. The skin comprises of epidermal, dermal and hypodermal layers, which mainly contain keratinocytes, fibroblasts and adipocytes, respectively, typically embedded on extracellular matrix made up of glycosaminoglycans and fibrous proteins. When the integrity of skin is compromised due to injury as in burns the coverage of skin has to be restored to facilitate repair and regeneration. Skin substitutes are preferred for wound coverage when the loss of skin is extensive especially in the case of second or third degree burns. Different kinds of skin substitutes with different features are commercially available; they can be classified into acellular skin substitutes, those with cultured epidermal cells and no dermal components, those with only dermal components, and tissue engineered substitutes that contain both epidermal and dermal components. Typically, adult wounds heal by fibrosis. Most organs are affected by fibrosis, with chronic fibrotic diseases estimated to be a leading cause of morbidity and mortality. In the skin, fibroproliferative disorders such as hypertrophic scars and keloid formation cause cosmetic and functional problems. Dermal fibroblasts are understood to be heterogeneous; this may have implications on post-burn wound healing since studies have shown that superficial and deep dermal fibroblasts are anti-fibrotic and pro-fibrotic, respectively. Selective use of superficial dermal fibroblasts rather than the conventional heterogeneous dermal fibroblasts may prove beneficial for post-burn wound healing.

  18. Skin Diseases: Skin Health and Skin Diseases

    Science.gov (United States)

    Skip Navigation Bar Home Current Issue Past Issues Skin Diseases Skin Health and Skin Diseases Past Issues / Fall 2008 Table of Contents ... acne to wrinkles Did you know that your skin is the largest organ of your body? It ...

  19. Skin toxicity and quality of life in patients with metastatic colorectal cancer during first-line panitumumab plus FOLFIRI treatment in a single-arm phase II study

    International Nuclear Information System (INIS)

    Integument-related toxicities are common during epidermal growth factor receptor (EGFR)-targeted therapy. Panitumumab is a fully human monoclonal antibody targeting the EGFR that significantly improves progression-free survival when added to chemotherapy in patients with metastatic colorectal cancer who have wild-type (WT) KRAS tumours. Primary efficacy and tolerability results from a phase II single-arm study of first-line panitumumab plus FOLFIRI in patients with metastatic colorectal cancer have been reported. Here we report additional descriptive tolerability and quality of life data from this trial. Integument-related toxicities and quality of life were analysed; toxicities were graded using modified National Cancer Institute Common Toxicity Criteria. Kaplan-Meier estimates of time to and duration of first integument-related toxicity were prepared. Quality of life was measured using EuroQoL EQ-5D and EORTC QLQ-C30. Best overall response was analysed by skin toxicity grade and baseline quality of life. Change in quality of life was analysed by skin toxicity severity. 154 patients were enrolled (WT KRAS n = 86; mutant KRAS n = 59); most (98%) experienced integument-related toxicities (most commonly rash [42%], dry skin [40%] and acne [36%]). Median time to first integument-related toxicity was 8 days; median duration was 334 days. Overall, proportionally more patients with grade 2+ skin toxicity responded (56%) compared with those with grade 0/1 (29%). Mean overall EQ-5D health state index scores (0.81 vs. 0.78), health rating scores (72.5 vs. 71.0) and QLQ-C30 global health status scores (65.8 vs. 66.7) were comparable at baseline vs. safety follow-up (8 weeks after completion), respectively and appeared unaffected by skin toxicity severity. First-line panitumumab plus FOLFIRI has acceptable tolerability and appears to have little impact on quality of life, despite the high incidence of integument-related toxicity. ClinicalTrials.gov NCT00508404

  20. Fetal ACL Fibroblasts Exhibit Enhanced Cellular Properties Compared with Adults

    OpenAIRE

    Stalling, Simone S.; Nicoll, Steven B.

    2008-01-01

    Fetal tendons and skin heal regeneratively without scar formation. Cells isolated from these fetal tissues exhibit enhanced cellular migration and collagen production in comparison to cells from adult tissue. We determined whether fetal and adult fibroblasts isolated from the anterior cruciate ligament (ACL), a tissue that does not heal regeneratively, exhibit differences in cell migration rates and collagen elaboration. An in vitro migration assay showed fetal ACL fibroblasts migrated twice ...

  1. Replacement of murine fibroblasts by human fibroblasts irradiated in obtaining feeder layer for the culture of human keratinocytes

    Energy Technology Data Exchange (ETDEWEB)

    Yoshito, Daniele; Sufi, Bianca S.; Santin, Stefany P.; Mathor, Monica B. [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil); Altran, Silvana C.; Isaac, Cesar [Universidade Sao Paulo (USP), Sao Paulo, SP (Brazil). Fac. de Medicina. Lab. de Microcirurgia Plastica; Esteves-Pedro, Natalia M. [Universidade Sao Paulo (USP), Sao Paulo, SP (Brazil). Fac. de Ciencias Farmaceuticas. Lab. de Controle Biologico; Herson, Marisa R. [DonorTissue Bank of Victoria (Australia)

    2011-07-01

    Human autologous epithelia cultivated in vitro, have been used successfully in treating damage to skin integrity. The methodology allowed the cultivation of these epithelia was described by Rheinwald and Green in 1975, this methodology consisted in seeding keratinocytes onto a feeder layer composed of lineage 3T3 murine fibroblasts, the proliferation rate is controlled through the action of ionizing radiation. However, currently there is a growing concern about the possibility of transmitting prions and murine viruses to transplanted patients. Taking into account this concern, in this present work, we replaced the feeder layer originally composed of murine fibroblasts by human fibroblasts. To obtain this new feeder layer was necessary to standardize the enough irradiation dose to inhibit the replication of human fibroblasts and the verification of effectiveness of the development of keratinocytes culture on a feeder layer thus obtained. According to the obtained results we can verify that the human fibroblasts irradiated at various tested doses (60, 70, 100, 200, 250 and 300 Gy) had their mitotic activity inactivated by irradiation, allowing the use of any of these doses to confection of the feeder layer, since these fibroblasts irradiated still showed viable until fourteen days of cultivation. In the test of colony formation efficiency was observed that keratinocytes seeded on irradiated human fibroblasts were able to develop satisfactorily, preserving their clonogenic potential. Therefore it was possible the replacement of murine fibroblasts by human fibroblasts in confection of the feeder layer, in order to eliminate this xenobiotic component of the keratinocytes culture. (author)

  2. Replacement of murine fibroblasts by human fibroblasts irradiated in obtaining feeder layer for the culture of human keratinocytes

    International Nuclear Information System (INIS)

    Human autologous epithelia cultivated in vitro, have been used successfully in treating damage to skin integrity. The methodology allowed the cultivation of these epithelia was described by Rheinwald and Green in 1975, this methodology consisted in seeding keratinocytes onto a feeder layer composed of lineage 3T3 murine fibroblasts, the proliferation rate is controlled through the action of ionizing radiation. However, currently there is a growing concern about the possibility of transmitting prions and murine viruses to transplanted patients. Taking into account this concern, in this present work, we replaced the feeder layer originally composed of murine fibroblasts by human fibroblasts. To obtain this new feeder layer was necessary to standardize the enough irradiation dose to inhibit the replication of human fibroblasts and the verification of effectiveness of the development of keratinocytes culture on a feeder layer thus obtained. According to the obtained results we can verify that the human fibroblasts irradiated at various tested doses (60, 70, 100, 200, 250 and 300 Gy) had their mitotic activity inactivated by irradiation, allowing the use of any of these doses to confection of the feeder layer, since these fibroblasts irradiated still showed viable until fourteen days of cultivation. In the test of colony formation efficiency was observed that keratinocytes seeded on irradiated human fibroblasts were able to develop satisfactorily, preserving their clonogenic potential. Therefore it was possible the replacement of murine fibroblasts by human fibroblasts in confection of the feeder layer, in order to eliminate this xenobiotic component of the keratinocytes culture. (author)

  3. Keratinocyte growth factor mRNA expression in periodontal ligament fibroblasts

    DEFF Research Database (Denmark)

    Dabelsteen, S; Wandall, H H; Grøn, B; Dabelsteen, E

    1997-01-01

    Keratinocyte growth factor (KGF) is a fibroblast growth factor which mediates epithelial growth and differentiation. KGF is expressed in subepithelial fibroblasts, but generally not in fibroblasts of deep connective tissue, such as fascia and ligaments. Here we demonstrate that KGF mRNA is...... expressed in periodontal ligament fibroblasts, and that the expression is increased upon serum stimulation. Fibroblasts from human periodontal ligament, from buccal mucosa, from gingiva, and from skin were established from explants. Alkaline phosphatase activity was used as an indicator of the periodontal...

  4. A comparative evaluation of in vitro skin sensitisation tests: the human cell-line activation test (h-CLAT) versus the local lymph node assay (LLNA).

    Science.gov (United States)

    Ashikaga, Takao; Sakaguchi, Hitoshi; Sono, Sakiko; Kosaka, Nanae; Ishikawa, Makie; Nukada, Yuko; Miyazawa, Masaaki; Ito, Yuichi; Nishiyama, Naohiro; Itagaki, Hiroshi

    2010-08-01

    We previously developed the human cell-line activation test (h-CLAT) in vitro skin sensitisation test, based on our reported finding that a 24-hour exposure of THP-1 cells (a human monocytic leukaemia cell line) to sensitisers is sufficient to induce the augmented expression of CD86 and CD54. The aim of this study is to confirm the predictive value of h-CLAT for skin sensitisation activity by employing a larger number of test chemicals. One hundred chemicals were selected, according to their categorisation in the local lymph node assay (LLNA), as being: extreme, strong, moderate and weak sensitisers, and non-sensitisers. The correlation of the h-CLAT results with the LLNA results was 84%. There were some false negatives (e.g. benzoyl peroxide, hexyl cinnamic aldehyde) and some false positives (e.g. 1-bromobutane, diethylphthalate). Eight out of the 9 false negatives (89%) were water-insoluble chemicals. The h-CLAT could positively predict not only extreme and strong sensitisers, but also moderate and weak sensitisers, though the detection rates of weak sensitisers and non-sensitisers were comparatively low. Some sensitisers enhanced both CD86 and CD54 levels, and some enhanced the level of only one of them. The use of the combination of CD86 and CD54 induction as a positive indicator, improved the accuracy of the test. In conclusion, the h-CLAT is expected to be a useful cell-based in vitro method for predicting skin sensitisation potential. PMID:20822320

  5. Evaluation of in vitro toxicity of Rumalaya liniment using mouse embryonic fibroblasts and human keratinocytes

    Directory of Open Access Journals (Sweden)

    Sandeep Ravi Varma

    2011-01-01

    Full Text Available The skin irritation potential of topical formulations is investigated prior to human exposure to identify the chemicals which might induce adverse skin reactions. Rumalaya liniment (RL a novel formulation using natural oils and plant extracts is used for reducing inflammations associated with musculoskeletal disorders. Preclinical studies on RL are needed prior to skin application. The aim of the study was to evaluate the possible cytotoxic effects of RL and a commercial sample (CS on mouse embryo fibroblasts and human keratinocytes using neutral red uptake, (3-(4,5- Dimethylthiazol -2-yl-2,5-di phenyl tetrazolium bromide (MTT and resazurin assay. The CTC 50 values obtained for RL was significantly higher than that of CS, which revealed that RL is less toxic to CS. RL was less toxic (<17% on both cell lines at 400 ?g/ml and was nontoxic at further lower concentrations, whereas the toxicity of CS was above 59% even at 400 ?g/ml. It was observed from the present study that by using three different assay methods and two different cell lines, the toxicity of RL was significantly lower than that observed with CS. From the study, it could be concluded that RL could be safer to skin due to their low cytotoxicity as compared with CS.

  6. Inducible responses to DNA damage in the mouse embryo fibroblasts cell line C3H/10T1/2 and its transformed counterpart C3H/MCA

    International Nuclear Information System (INIS)

    Early passage mouse embryo fibroblasts cells (C3H/10T1/2) were treated with ultraviolet (UV) radiation of 12-O-tetradecanoylphorbol-13-acetate (TPA) in order to determine whether such treatment induced DNA repair processes, measured as increased survival and mutagenesis of Herpes simplex (HSV-1). No enhanced host cell reactivation of UV-irradiated virus was observed following treatment of cells with UV-irradiation or TPA. Replication of undamaged virus in untreated C3H cells resulted in an increase over the background mutation frequency. When the cells were UV-irradiated and infected with unirradiated virus, a decrease in mutagenesis was observed. Decreased untargeted mutagenesis was shown to be dose- and time-dependent, reaching a minimum at a fluence of 5-7 Jm/sup /minus/2/ for 24 hours between irradiation and infection of cells. There was no change in mutagenesis of UV-irradiated virus grown in UV-irradiated cells compared to untreated cells. The repair capacity of methylcholanthrene-transformed C3H cells (MCA cells) was compared with untransformed C3H cells. The cell lines demonstrated similar cell survival curves following UV-irradiation but differed markedly in their ability to repair damaged HSV-1

  7. Intra- and inter-laboratory reproducibility and accuracy of the LuSens assay: A reporter gene-cell line to detect keratinocyte activation by skin sensitizers.

    Science.gov (United States)

    Ramirez, Tzutzuy; Stein, Nadine; Aumann, Alexandra; Remus, Tina; Edwards, Amber; Norman, Kimberly G; Ryan, Cindy; Bader, Jackie E; Fehr, Markus; Burleson, Florence; Foertsch, Leslie; Wang, Xiaohong; Gerberick, Frank; Beilstein, Paul; Hoffmann, Sebastian; Mehling, Annette; van Ravenzwaay, Bennard; Landsiedel, Robert

    2016-04-01

    Several non-animal methods are now available to address the key events leading to skin sensitization as defined by the adverse outcome pathway. The KeratinoSens™ assay addresses the cellular event of keratinocyte activation and is a method accepted under OECD TG 442D. In this study, the results of an inter-laboratory evaluation of the "me-too" LuSens assay, a bioassay that uses a human keratinocyte cell line harboring a reporter gene construct composed of the rat antioxidant response element (ARE) of the NADPH:quinone oxidoreductase 1 gene and the luciferase gene, are described. Earlier in-house validation with 74 substances showed an accuracy of 82% in comparison to human data. When used in a battery of non-animal methods, even higher predictivity is achieved. To meet European validation criteria, a multicenter study was conducted in 5 laboratories. The study was divided into two phases, to assess 1) transferability of the method, and 2) reproducibility and accuracy. Phase I was performed by testing 8 non-coded test substances; the results showed a good transferability to naïve laboratories even without on-site training. Phase II was performed with 20 coded test substances (performance standards recommended by OECD, 2015). In this phase, the intra- and inter-laboratory reproducibility as well as accuracy of the method was evaluated. The data demonstrate a remarkable reproducibility of 100% and an accuracy of over 80% in identifying skin sensitizers, indicating a good concordance with in vivo data. These results demonstrate good transferability, reliability and accuracy of the method thereby achieving the standards necessary for use in a regulatory setting to detect skin sensitizers. PMID:26796489

  8. Chronic γ-irradiation results in increased cell killing and chromosomal aberration with specific breakpoints in fibroblast cell strains derived from non-Hodgkin's lymphoma patients

    International Nuclear Information System (INIS)

    Cultured skin fibroblast cells from 16 NHL (non-Hodgkin's lymphoma) patients and 2 clinically normal subjects were compared for cell survival and chromosomal aberration after chronic γ-irradiation. Fibroblasts from an ataxia telangiectasia (AT) homozygote and an AT heterozygote were used as positive controls. Following irradiation, fibroblasts from all 16 NHL patients showed an increase in both cell death and chromosomal aberration (breaks and rearrangements) compared to normal subjects. The difference in frequency of chromosomal aberration between normals and NHL-patients remained virtually unchanged over a period of 24-72 h post irradiation incubation of cells. Cell cycle analysis by flow cytometry carried out in 1 normal and 1 NHL fibroblast cell strain showed that more cells representing the NHL patient were in G2/M phase compared to the normal at various times of cytogenetic analysis. While the AT homozygote appeared to be the most radiosensitive, the AT heterozygote showed a slightly higher incidence of cell death and chromosomal aberration than the normals. The cellular and chromosomal radiosensitivity of fibroblast cell lines from NHL-patients differed slightly from that of AT heterozygote but clearly occupied an intermediate position between the AT homozygote and the normal subjects. Cells from 3 of the NHL patients showed radiation-induced specific chromosomal breaks involving chromosomes 1, 2, 6, 8, 10 and 11 which correspond to known fragile sites. Such breakpoints associated with increased radiosensitivity may be indicative of predisposition to malignancy in the patients studied. (author). 30 refs., 2 figs., 4 tabs

  9. Your Skin

    Science.gov (United States)

    ... Skiing, Snowboarding, Skating Crushes What's a Booger? Your Skin KidsHealth > For Kids > Your Skin Print A A ... are really dead skin cells. continue Bye-Bye Skin Cells These old cells are tough and strong, ...

  10. DNA repair in human fibroblasts, as reflected by host-cell reactivation of a transfected UV-irradiated luciferase gene, is not related to donor age

    International Nuclear Information System (INIS)

    The effect of donor age on the ability of mammalian cells to repair ultraviolet (UV)-induced DNA damage has been studied using several approaches, most recently via assays that measure the host-cell reactivation (HCR) of UV-irradiated reporter gene-containing plasmid vectors following their transfection into cells. Plasmid HCR assays indirectly quantify a cell line's ability to perform nucleotide excision repair (NER) by measuring the enzyme activity of the repaired reporter gene, e.g., chloramphenical acetyltransferase (cat) or luciferase (luc), and are useful in studies investigating whether increasing age may be a risk factor for the deficient repair of potentially cancer-causing, sunlight-induced, DNA lesions in skin cells. In our study, we quantified the DNA repair ability of cultured, nontransformed, human skin fibroblast lines through their HCR of a transfected UV-C-irradiated plasmid containing luc. HCR was measured at various times after transfection in five lines from normal donors of ages 21-96 years, and from one donor who had xeroderma pigmentosum (XP). The normal lines displayed increasing HCR at successive post-transfection time points and showed no significant correlation between HCR and donor age. The XP-A line, known to be markedly deficient in NER of UV-induced DNA damage, showed minimal evidence of HCR compared to the normal lines. To further assess potential variation in HCR with donor age, fibroblast lines from five old donors, ages 84-94 years, were compared with lines from five young donors, ages 17-26 years. While significant differences in HCR were found between some lines, no significant difference was found between the young and old age groups (P=0.44). Our study provides no indication that the higher incidence of skin cancer observed with increasing age is due to an age-related decrease in the ability to repair UV-induced DNA damage

  11. Mitochondria-Targeted Vitamin E Protects Skin from UVB-Irradiation.

    Science.gov (United States)

    Kim, Won-Serk; Kim, Ikyon; Kim, Wang-Kyun; Choi, Ju-Yeon; Kim, Doo Yeong; Moon, Sung-Guk; Min, Hyung-Keun; Song, Min-Kyu; Sung, Jong-Hyuk

    2016-05-01

    Mitochondria-targeted vitamin E (MVE) is designed to accumulate within mitochondria and is applied to decrease mitochondrial oxidative damage. However, the protective effects of MVE in skin cells have not been identified. We investigated the protective effect of MVE against UVB in dermal fibroblasts and immortalized human keratinocyte cell line (HaCaT). In addition, we studied the wound-healing effect of MVE in animal models. We found that MVE increased the proliferation and survival of fibroblasts at low concentration (i.e., nM ranges). In addition, MVE increased collagen production and downregulated matrix metalloproteinase1. MVE also increased the proliferation and survival of HaCaT cells. UVB increased reactive oxygen species (ROS) production in fibroblasts and HaCaT cells, while MVE decreased ROS production at low concentration. In an animal experiment, MVE accelerated wound healing from laser-induced skin damage. These results collectively suggest that low dose MVE protects skin from UVB irradiation. Therefore, MVE can be developed as a cosmetic raw material. PMID:26869457

  12. Fibroblast biology in pterygia.

    Science.gov (United States)

    Kim, Kyoung Woo; Park, Soo Hyun; Kim, Jae Chan

    2016-01-01

    Activation of fibroblasts is a vital process during wound healing. However, if prolonged and exaggerated, profibrotic pathways lead to tissue fibrosis or scarring and further organ malfunction. Although the pathogenesis of pterygium is known to be multi-factorial, additional studies are needed to better understand the pathways initiated by fibroblast activation for the purpose of therapeutic translation. Regarding pterygium as a possible systemic disorder, we discuss the different cell types that pterygium fibroblasts originate from. These may include bone marrow-derived progenitor cells, cells undergoing epithelial-mesenchymal transition (EMT), and local resident stromal cells. We also describe how pterygium fibroblasts can be activated and perpetuate profibrotic signaling elicited by various proliferative drivers, immune-inflammation, and novel factors such as stromal cell-derived factor-1 (SDF-1) as well as a known key fibrotic factor, transforming growth factor-beta (TGF-?). Finally, epigenetic modification is discussed to explain inherited susceptibility to pterygium. PMID:26675401

  13. How to Approach Finnish Retail Market when Launching a New Skin Care Line: a Case Study of Crations Couleurs

    OpenAIRE

    Nordenswan, Katarina; Huttunen, Anne

    2012-01-01

    The cosmetics industry is one of the biggest lines of businesses in the world. In Finland people spend thousands of Euros per year on cosmetic and hygiene products. Everything changes constantly and this has reflected to the cosmetics industry as well as consumers. People increasingly desire several options to choose from and want quick results. The topic for this thesis came from a French cosmetic company Crations Couleurs which develops and manufactures raw materials for different cosm...

  14. LINES

    Directory of Open Access Journals (Sweden)

    Minas Bakalchev

    2015-10-01

    Full Text Available The perception of elements in a system often creates their interdependence, interconditionality, and suppression. The lines from a basic geometrical element have become the model of a reductive world based on isolation according to certain criteria such as function, structure, and social organization. Their traces are experienced in the contemporary world as fragments or ruins of a system of domination of an assumed hierarchical unity. How can one release oneself from such dependence or determinism? How can the lines become less systematic and forms more autonomous, and less reductive? How is a form released from modernistic determinism on the new controversial ground? How can these elements or forms of representation become forms of action in the present complex world? In this paper, the meaning of lines through the ideas of Le Corbusier, Leonidov, Picasso, and Hitchcock is presented. Spatial research was made through a series of examples arising from the projects of the architectural studio Residential Transformations, which was a backbone for mapping the possibilities ranging from playfulness to exactness, as tactics of transformation in the different contexts of the contemporary world.

  15. Multiple functions of gingival and mucoperiosteal fibroblasts in oral wound healing and repair.

    Science.gov (United States)

    Chiquet, Matthias; Katsaros, Christos; Kletsas, Dimitris

    2015-06-01

    Fibroblasts are cells of mesenchymal origin. They are responsible for the production of most extracellular matrix in connective tissues and are essential for wound healing and repair. In recent years, it has become clear that fibroblasts from different tissues have various distinct traits. Moreover, wounds in the oral cavity heal under very special environmental conditions compared with skin wounds. Here, we reviewed the current literature on the various interconnected functions of gingival and mucoperiosteal fibroblasts during the repair of oral wounds. The MEDLINE database was searched with the following terms: (gingival OR mucoperiosteal) AND fibroblast AND (wound healing OR repair). The data gathered were used to compare oral fibroblasts with fibroblasts from other tissues in terms of their regulation and function during wound healing. Specifically, we sought answers to the following questions: (i) what is the role of oral fibroblasts in the inflammatory response in acute wounds; (ii) how do growth factors control the function of oral fibroblasts during wound healing; (iii) how do oral fibroblasts produce, remodel and interact with extracellular matrix in healing wounds; (iv) how do oral fibroblasts respond to mechanical stress; and (v) how does aging affect the fetal-like responses and functions of oral fibroblasts? The current state of research indicates that oral fibroblasts possess unique characteristics and tightly controlled specific functions in wound healing and repair. This information is essential for developing new strategies to control the intraoral wound-healing processes of the individual patient. PMID:25867977

  16. Loss of PPAR? expression by fibroblasts enhances dermal wound closure

    Directory of Open Access Journals (Sweden)

    Sha Wei

    2012-04-01

    Full Text Available Abstract Background Peroxisome proliferator-activated receptor (PPAR? may be a key regulator of connective tissue deposition and remodeling in vivo. PPAR? expression is reduced in dermal fibroblasts isolated from fibrotic areas of scleroderma patients; PPAR? agonists suppress the persistent fibrotic phenotype of this cell type. Previously, we showed that loss of PPAR? expression in fibroblasts resulted in enhanced bleomycin-induced skin fibrosis. However, whether loss of PPAR? expression in skin fibroblasts affects cutaneous tissue repair or homeostasis is unknown. Results Mice deleted for PPAR? in skin fibroblasts show an enhanced rate of dermal wound closure, concomitant with elevated phosphorylation of Smad3, Akt and ERK, and increased expression of proliferating cell nuclear antigen (PCNA, collagen, ?-smooth muscle actin (?-SMA and CCN2. Conversely, dermal homeostasis was not appreciably affected by loss of PPAR? expression. Conclusion PPAR? expression by fibroblasts suppresses cutaneous tissue repair. In the future, direct PPAR? antagonists and agonists might be of clinical benefit in controlling chronic wounds or scarring, respectively.

  17. Solar aging of the skin

    International Nuclear Information System (INIS)

    Aging of the skin is a combination of chronological aging and degenerative influences from the environment. Most important is the influence of ultraviolet radiation upon dermal fibroblasts. As a result of changes in collagen and elastic fibres with deposition of ''solar elastosis'', chronically sun-exposed skin looses elasticity and develops wrinkles. Results of recent investigations show that solar degenerative changes of the skin are spontaneously repaired when the radiation is stopped and/or sun filters are used. This repair process is accelerated by the topical use of retinoic acid

  18. The development of a 3D immunocompetent model of human skin

    International Nuclear Information System (INIS)

    As the first line of defence, skin is regularly exposed to a variety of biological, physical and chemical insults. Therefore, determining the skin sensitization potential of new chemicals is of paramount importance from the safety assessment and regulatory point of view. Given the questionable biological relevance of animal models to human as well as ethical and regulatory pressure to limit or stop the use of animal models for safety testing, there is a need for developing simple yet physiologically relevant models of human skin. Herein, we describe the construction of a novel immunocompetent 3D human skin model comprising of dendritic cells co-cultured with keratinocytes and fibroblasts. This model culture system is simple to assemble with readily-available components and importantly, can be separated into its constitutive individual layers to allow further insight into cell–cell interactions and detailed studies of the mechanisms of skin sensitization. In this study, using non-degradable microfibre scaffolds and a cell-laden gel, we have engineered a multilayer 3D immunocompetent model comprised of keratinocytes and fibroblasts that are interspersed with dendritic cells. We have characterized this model using a combination of confocal microscopy, immuno-histochemistry and scanning electron microscopy and have shown differentiation of the epidermal layer and formation of an epidermal barrier. Crucially the immune cells in the model are able to migrate and remain responsive to stimulation with skin sensitizers even at low concentrations. We therefore suggest this new biologically relevant skin model will prove valuable in investigating the mechanisms of allergic contact dermatitis and other skin pathologies in human. Once fully optimized, this model can also be used as a platform for testing the allergenic potential of new chemicals and drug leads. (paper)

  19. Fibroblast differentiation in subcutaneous fibrosis after postmastectomy radiotherapy

    International Nuclear Information System (INIS)

    In order to acquire a better understanding of the mechanism of radiation-induced fibrosis, we studied the differentiation of normal skin fibroblasts cultured from breast cancer radiotherapy patients with different risk of fibrosis. The differentiation state of fibroblasts was characterized in clonal cultures using established cytomorphological criteria. Collagen synthesis was determined by 3H-proline incorporation into pepsin-resistant protein. Radiation-induced inactivation of fibroblasts was paralleled by an increase in terminally differentiated fibrocytes, demonstrating that premature terminal differentiation is an important response to irradiation of fibroblasts from radiotherapy patients. Surviving colony-forming fibroblasts showed a change in differentiation with an increase in the ratio L:E of progenitor fibroblasts in late (L) compared to early (E) differentiation states. Furthermore, increased collagen production was observed after irradiation. The results provide evidence supporting a role of terminal fibroblast differentiation in radiation-induced fibrosis and imply that the progenitor population surviving radiotherapy might be more prone to terminal differentiation than before radiotherapy

  20. Differential heat shock response of primary human cell cultures and established cell lines

    DEFF Research Database (Denmark)

    Richter, W W; Issinger, O G

    1986-01-01

    The influence of hyperthermia on the cellular growth and protein synthesis pattern from primary human brain tumour cells and skin fibroblasts was compared with established and experimentally transformed tumour cell lines. Primary cell cultures did not show any visible morphological changes after 42...... degrees C treatment, whereas in immortalized cell lines usually 90% of the cells were found in suspension. Enhanced expression of the major heat shock protein (hsp 70) was found in all heat-treated cells. In contrast to the primary cell cultures, established and transformed cell lines synthesized a...

  1. Abscisic acid ameliorates the systemic sclerosis fibroblast phenotype in vitro

    International Nuclear Information System (INIS)

    Highlights: ► ABA is an endogenous hormone in humans, regulating different cell responses. ► ABA reverts some of the functions altered in SSc fibroblasts to a normal phenotype. ► UV-B irradiation increases ABA content in SSc cultures. ► SSc fibroblasts could benefit from exposure to ABA and/or to UV-B. -- Abstract: The phytohormone abscisic acid (ABA) has been recently identified as an endogenous hormone in humans, regulating different cell functions, including inflammatory processes, insulin release and glucose uptake. Systemic sclerosis (SSc) is a chronic inflammatory disease resulting in fibrosis of skin and internal organs. In this study, we investigated the effect of exogenous ABA on fibroblasts obtained from healthy subjects and from SSc patients. Migration of control fibroblasts induced by ABA was comparable to that induced by transforming growth factor-β (TGF-β). Conversely, migration toward ABA, but not toward TGF-β, was impaired in SSc fibroblasts. In addition, ABA increased cell proliferation in fibroblasts from SSc patients, but not from healthy subjects. Most importantly, presence of ABA significantly decreased collagen deposition by SSc fibroblasts, at the same time increasing matrix metalloproteinase-1 activity and decreasing the expression level of tissue inhibitor of metalloproteinase (TIMP-1). Thus, exogenously added ABA appeared to revert some of the functions altered in SSc fibroblasts to a normal phenotype. Interestingly, ABA levels in plasma from SSc patients were found to be significantly lower than in healthy subjects. UV-B irradiation induced an almost 3-fold increase in ABA content in SSc cultures. Altogether, these results suggest that the fibrotic skin lesions in SSc patients could benefit from exposure to high(er) ABA levels.

  2. Abscisic acid ameliorates the systemic sclerosis fibroblast phenotype in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Bruzzone, Santina, E-mail: santina.bruzzone@unige.it [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Advanced Biotechnology Center, Largo Rosanna Benzi 10, 16132 Genova (Italy); Battaglia, Florinda [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Mannino, Elena [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Parodi, Alessia [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Fruscione, Floriana [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Advanced Biotechnology Center, Largo Rosanna Benzi 10, 16132 Genova (Italy); Basile, Giovanna [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Salis, Annalisa; Sturla, Laura [Department of Experimental Medicine, Section of Biochemistry, University of Genova, Viale Benedetto XV 1, 16132 Genova (Italy); Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Negrini, Simone; Kalli, Francesca; Stringara, Silvia [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Filaci, Gilberto [Centre of Excellence for Biomedical Research, University of Genova, Viale Benedetto XV 9, 16132 Genova (Italy); Department of Internal Medicine, Viale Benedetto XV 6, 16132 Genova (Italy); and others

    2012-05-25

    Highlights: Black-Right-Pointing-Pointer ABA is an endogenous hormone in humans, regulating different cell responses. Black-Right-Pointing-Pointer ABA reverts some of the functions altered in SSc fibroblasts to a normal phenotype. Black-Right-Pointing-Pointer UV-B irradiation increases ABA content in SSc cultures. Black-Right-Pointing-Pointer SSc fibroblasts could benefit from exposure to ABA and/or to UV-B. -- Abstract: The phytohormone abscisic acid (ABA) has been recently identified as an endogenous hormone in humans, regulating different cell functions, including inflammatory processes, insulin release and glucose uptake. Systemic sclerosis (SSc) is a chronic inflammatory disease resulting in fibrosis of skin and internal organs. In this study, we investigated the effect of exogenous ABA on fibroblasts obtained from healthy subjects and from SSc patients. Migration of control fibroblasts induced by ABA was comparable to that induced by transforming growth factor-{beta} (TGF-{beta}). Conversely, migration toward ABA, but not toward TGF-{beta}, was impaired in SSc fibroblasts. In addition, ABA increased cell proliferation in fibroblasts from SSc patients, but not from healthy subjects. Most importantly, presence of ABA significantly decreased collagen deposition by SSc fibroblasts, at the same time increasing matrix metalloproteinase-1 activity and decreasing the expression level of tissue inhibitor of metalloproteinase (TIMP-1). Thus, exogenously added ABA appeared to revert some of the functions altered in SSc fibroblasts to a normal phenotype. Interestingly, ABA levels in plasma from SSc patients were found to be significantly lower than in healthy subjects. UV-B irradiation induced an almost 3-fold increase in ABA content in SSc cultures. Altogether, these results suggest that the fibrotic skin lesions in SSc patients could benefit from exposure to high(er) ABA levels.

  3. Polycomponent mesotherapy formulations for the treatment of skin aging and improvement of skin quality

    Directory of Open Access Journals (Sweden)

    Prikhnenko S

    2015-04-01

    Full Text Available Sergey Prikhnenko Private Practice, Novosibirsk, Russia Abstract: Skin aging can largely be attributed to dermal fibroblast dysfunction and a decrease in their biosynthetic activity. Regardless of the underlying causes, aging fibroblasts begin to produce elements of the extracellular matrix in amounts that are insufficient to maintain the youthful appearance of skin. The goal of mesopreparations is primarily to slow down and correct changes in skin due to aging. The rationale for developing complex polycomponent mesopreparations is based on the principle that aging skin needs to be supplied with the various substrates that are key to the adequate functioning of the fibroblast. The quintessential example of a polycomponent formulation – NCTF® (New Cellular Treatment Factor – includes vitamins, minerals, amino acids, nucleotides, coenzymes and antioxidants, as well as hyaluronic acid, designed to help fibroblasts function more efficiently by providing a more optimal environment for biochemical processes and energy generation, as well as resisting the effects of oxidative stress. In vitro experiments suggest that there is a significant increase in the synthetic and prophylactic activity of fibroblasts with treated NCTF, and a significant increase in the ability of cells to resist oxidative stress. The current article looks at the rationale behind the development of polycomponent mesopreparations, using NCTF as an example. Keywords: mesotherapy, skin aging, skin quality

  4. The elemental analysis of normal and Menkes' fibroblast cells with the SPMP

    International Nuclear Information System (INIS)

    A scanning proton microprobe has been used to study the elemental distributions, including trace elements, in human skin fibroblast cells. Both normal fibroblasts and fibroblasts cultured from patients with Menkes' disease, an X-linked genetic disorder known to be associated with defective copper metabolism, were examined by the probe. Cells grown in a copper-rich medium exhibited higher levels of intracellular copper but were found to be susceptible to the toxic effect of the elevated copper level. In normal maintenance medium the Menkes' cells recorded an average intracellular copper level six times higher than normal fibroblasts. (orig.)

  5. Dupuytren's Contracture: Fibroblast Contraction?

    Science.gov (United States)

    Gabbiani, Giulio; Majno, Guido

    1972-01-01

    In 6 cases of Dupuytren's disease and 1 of Ledderhose's disease, the nodules of the palmar and plantar aponeurosis were examined by light and electron microscopy. The cells composing these nodules, presumably fibroblasts, showed three significant ultrastructural features: (1) a fibrillar system similar to that of smooth muscle cells; (2) nuclear deformations such as are found in contracted cells, the severest being recognizable by light microscopy (cross-banded nuclei); (3) cell-to-cell and cell-to-stroma attachments. Based on these data and on recent information about the biology of the fibroblasts, it is suggested that these cells are fibroblasts that have modulated into contractile cells (myofibroblasts), and that their contraction plays a role in the pathogenesis of the contracture observed clinically. ImagesFig 10Fig 5Fig 11Fig 6 and 7Fig 8Fig 1Fig 2Fig 9Fig 3Fig 4 PMID:5009249

  6. Chronic actinic damage of facial skin.

    Science.gov (United States)

    Bilaç, Cemal; Şahin, Mustafa Turhan; Öztürkcan, Serap

    2014-01-01

    Chronic actinic damage of the skin manifests itself as extrinsic skin aging (photoaging) and photocarcinogenesis. During the last decade, substantial progress has been made in understanding cellular and molecular mechanisms of photoaging. DNA photodamage and ultraviolet-generated reactive oxygen species are the initial events that lead to most of the typical histologic and clinical manifestations of chronic photodamage of the skin. Chronic actinic damage affects all layers of the skin. Keratinocytes, melanocytes, fibroblasts, and endothelial cells are altered by ultraviolet radiation and can result in numerous changes in human skin, particularly the skin of fair-skinned individuals. These changes include actinic keratosis, thickening and wrinkling, elastosis, telengiectasia, solar comedones, diffuse or mottled hyperpigmentation, and skin cancers. There are many options in the treatment of changes caused by chronic actinic damage. The most effective measure of prevention of the photoaging and photocarcinogenesis is sun protection. PMID:25441468

  7. Immortalization of Werner syndrome and progeria fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Saito, H.; Moses, R.E. (Baylor College of Medicine, Houston, TX (USA))

    1991-02-01

    Human fibroblast cells from two different progeroid syndromes, Werner syndrome (WS) and progeria, were established as immortalized cell lines by transfection with plasmid DNA containing the SV40 early region. The lineage of each immortalized cell line was confirmed by VNTR analysis. Each of the immortalized cell lines maintained its original phenotype of slow growth. DNA repair ability of these cells was also studied by measuring sensitivity to killing by uv or the DNA-damaging drugs methyl methansulfonate, bleomycin, and cis-dichlorodiamine platinum. The results showed that both WS and progeria cells have normal sensitivity to these agents.

  8. Immortalization of Werner syndrome and progeria fibroblasts

    International Nuclear Information System (INIS)

    Human fibroblast cells from two different progeroid syndromes, Werner syndrome (WS) and progeria, were established as immortalized cell lines by transfection with plasmid DNA containing the SV40 early region. The lineage of each immortalized cell line was confirmed by VNTR analysis. Each of the immortalized cell lines maintained its original phenotype of slow growth. DNA repair ability of these cells was also studied by measuring sensitivity to killing by uv or the DNA-damaging drugs methyl methansulfonate, bleomycin, and cis-dichlorodiamine platinum. The results showed that both WS and progeria cells have normal sensitivity to these agents

  9. Skin Conditions

    Science.gov (United States)

    Your skin is your body's largest organ. It covers and protects your body. Your skin Holds body fluids in, preventing dehydration Keeps harmful ... it Anything that irritates, clogs, or inflames your skin can cause symptoms such as redness, swelling, burning, ...

  10. Skin Cancer

    Science.gov (United States)

    ... are specialized skin cells that produce pigment called melanin. The melanin pigment produced by melanocytes gives skin its color. ... absorbing and scattering the energy. People with more melanin have darker skin and better protection from UV ...

  11. Aging Skin

    Science.gov (United States)

    ... email address Submit Home > Healthy Aging > Wellness Healthy Aging Aging skin More information on aging skin When it ... treated early. Return to top More information on Aging skin Read more from womenshealth.gov Varicose Veins ...

  12. Radiation response in vitro of fibroblasts from a Fanconi anemia patient with marked clinical radiosensitivity

    Energy Technology Data Exchange (ETDEWEB)

    Djuzenova, C.; Flentje, M. [Dept. of Radiotherapy, Univ. of Wuerzburg, Wuerzburg (Germany); Plowman, P.N. [Radiotherapy/Clinical Oncology, St. Bartholomew' s Hospital, London (United Kingdom)

    2004-12-01

    Background: fanconi anemia (FA) is an autosomal recessive chromosome instability disorder characterized by progressive pancytopenia and cancer susceptibility. The risks of radiation therapy in FA patients who have cancer remain to be investigated. Recently, Marcou et al. (2001) reported a case of severe clinical radiosensitivity in a female FA patient with a tonsillar squamous cell carcinoma treated by radiotherapy. By contrast, her in vitro irradiated skin fibroblasts revealed nearly normal radiosensitivity as determined by the colony survival assay. Material and methods: in view of this discrepancy, the radiation response of this particular FA fibroblast strain (designated 425BR) was further analyzed in the present study by means of the alkaline single-cell gel electrophoresis (Comet) assay, and also by the cytochalasin-blocked micronuclei (MN) test. In addition, the expression levels of DNA repair proteins, hMre11, Rad50, and Rad51, were investigated using Western blot and foci immunofluorescence staining. Results: the Comet assay revealed that the initial DNA fragmentation in irradiated FA cells was two times higher and the DNA rejoining process was three times slower than that in control (1BR3) fibroblasts. Moreover, although the baseline level of MNs was lower in FA cells than in controls, the FA fibroblasts were more prone (about two times) to MN production than control cells when irradiated with 2-4 Gy. Western blot analysis of the DNA repair proteins (hMre11, Rad50, and Rad51) did not reveal any abnormalities in protein expression levels or their migration patterns in the fibroblasts derived from an FA patient either before or after irradiation. At the same time, in vitro irradiated cells from the FA patient exhibited a significantly reduced number of nuclei with focally concentrated DNA repair Rad51 protein than in control cells. Conclusion: the increased DNA damage and MN induction in irradiated FA fibroblasts, and the reduction of the formation of DNA repair foci containing Rad51 suggest a possible link to the profound clinical radiosensitivity reported earlier for this FA patient. The findings on this particular FA cell strain presented in the study point toward the difficulties involved in the prediction of the radiation response of cell lines and tumors based solely on the colony survival test. (orig.)

  13. Microarray Analysis of Host Cell Gene Transcription in Response to Varicella-Zoster Virus Infection of Human T Cells and Fibroblasts In Vitro and SCIDhu Skin Xenografts In Vivo

    OpenAIRE

    Jones, Jeremy O.; Arvin, Ann M.

    2003-01-01

    During primary infection, varicella-zoster virus (VZV) is spread via lymphocytes to skin, where it induces a rash and establishes latency in sensory ganglia. A live, attenuated varicella vaccine (vOka) was generated by using the VZV Oka strain (pOka), but the molecular basis for vOka attenuation remains unknown. Little is known concerning the effects of wild-type or attenuated VZV on cellular gene regulation in the host cells that are critical for pathogenesis. In this study, transcriptional ...

  14. Skin Photoaging and the Role of Antioxidants in Its Prevention

    OpenAIRE

    Pandel, Ruža; Poljšak, Borut; Godic, Aleksandar; Dahmane, Raja

    2013-01-01

    Photoaging of the skin depends primarily on the degree of ultraviolet radiation (UVR) and on an amount of melanin in the skin (skin phototype). In addition to direct or indirect DNA damage, UVR activates cell surface receptors of keratinocytes and fibroblasts in the skin, which leads to a breakdown of collagen in the extracellular matrix and a shutdown of new collagen synthesis. It is hypothesized that dermal collagen breakdown is followed by imperfect repair that yields a deficit in the stru...

  15. Laser printing of skin cells and human stem cells.

    Science.gov (United States)

    Koch, Lothar; Kuhn, Stefanie; Sorg, Heiko; Gruene, Martin; Schlie, Sabrina; Gaebel, Ralf; Polchow, Bianca; Reimers, Kerstin; Stoelting, Stephanie; Ma, Nan; Vogt, Peter M; Steinhoff, Gustav; Chichkov, Boris

    2010-10-01

    Laser printing based on laser-induced forward transfer (LIFT) is a new biofabrication technique for the arrangement of biological materials or living cells in well-defined patterns. In the current study, skin cell lines (fibroblasts/keratinocytes) and human mesenchymal stem cells (hMSC) were chosen for laser printing experiments due to their high potential in regeneration of human skin and new application possibilities of stem cell therapy. To evaluate the influence of LIFT on the cells, their survival rate, their proliferation and apoptotic activity, and the DNA damages and modifications of their cell surface markers were assessed and statistically evaluated over several days. The cells survived the transfer procedure with a rate of 98%? +/- 1% standard error of the mean (skin cells) and 90%? +/- 10% (hMSC), respectively. All used cell types maintain their ability to proliferate after LIFT. Further, skin cells and hMSC did not show an increase of apoptosis or DNA fragmentation. In addition, the hMSC keep their phenotype as proven by fluorescence activated cell sorting (FACS) analysis. This study demonstrates LIFT as a suitable technique for unharmed computer-controlled positioning of different cell types and a promising tool for future applications in the ex vivo generation of tissue replacements. PMID:19883209

  16. Molecularly Characterized Solvent Extracts and Saponins from Polygonum hydropiper L. Show High Anti-Angiogenic, Anti-Tumor, Brine Shrimp, and Fibroblast NIH/3T3 Cell Line Cytotoxicity

    Science.gov (United States)

    Ayaz, Muhammad; Junaid, Muhammad; Ullah, Farhat; Sadiq, Abdul; Subhan, Fazal; Khan, Mir Azam; Ahmad, Waqar; Ali, Gowhar; Imran, Muhammad; Ahmad, Sajjad

    2016-01-01

    Polygonum hydropiper is used as anti-cancer and anti-rheumatic agent in folk medicine. This study was designed to investigate the anti-angiogenic, anti-tumor, and cytotoxic potentials of different solvent extracts and isolated saponins. Samples were analyzed using GC, Gas Chromatography–Mass Spectrometry (GC–MS) to identify major and bioactive compounds. Quantitation of antiangiogenesis for the plant's samples including methanolic extract (Ph.Cr), its subsequent fractions; n-hexane (Ph.Hex), chloroform (Ph.Chf), ethyl acetate (Ph.EtAc), n-Butanol (Ph.Bt), aqueous (Ph.Aq), saponins (Ph.Sp) were performed using the chick embryo chorioallantoic membrane (CAM) assay. Potato disc anti-tumor assay was performed on Agrobacterium tumefaciens containing tumor inducing plasmid. Cytotoxicity was performed against Artemia salina and mouse embryonic fibroblast NIH/3T3 cell line following contact toxicity and MTT cells viability assays, respectively. The GC–MS analysis of Ph.Cr, Ph.Hex, Ph.Chf, Ph.Bt, and Ph.EtAc identified 126, 124, 153, 131, and 164 compounds, respectively. In anti-angiogenic assay, Ph.Chf, Ph.Sp, Ph.EtAc, and Ph.Cr exhibited highest activity with IC50 of 28.65, 19.21, 88.75, and 461.53 μg/ml, respectively. In anti-tumor assay, Ph.Sp, Ph.Chf, Ph.EtAc, and Ph.Cr were most potent with IC50 of 18.39, 73.81, 217.19, and 342.53 μg/ml, respectively. In MTT cells viability assay, Ph.Chf, Ph.EtAc, Ph.Sp were most active causing 79.00, 72.50, and 71.50% cytotoxicity, respectively, at 1000 μg/ml with the LD50 of 140, 160, and 175 μg/ml, respectively. In overall study, Ph.Chf and Ph.Sp have shown overwhelming results which signifies their potentials as sources of therapeutic agents against cancer. PMID:27065865

  17. Molecularly Characterized Solvent Extracts and Saponins from Polygonum hydropiper L. Show High Anti-Angiogenic, Anti-Tumor, Brine Shrimp, and Fibroblast NIH/3T3 Cell Line Cytotoxicity.

    Science.gov (United States)

    Ayaz, Muhammad; Junaid, Muhammad; Ullah, Farhat; Sadiq, Abdul; Subhan, Fazal; Khan, Mir Azam; Ahmad, Waqar; Ali, Gowhar; Imran, Muhammad; Ahmad, Sajjad

    2016-01-01

    Polygonum hydropiper is used as anti-cancer and anti-rheumatic agent in folk medicine. This study was designed to investigate the anti-angiogenic, anti-tumor, and cytotoxic potentials of different solvent extracts and isolated saponins. Samples were analyzed using GC, Gas Chromatography-Mass Spectrometry (GC-MS) to identify major and bioactive compounds. Quantitation of antiangiogenesis for the plant's samples including methanolic extract (Ph.Cr), its subsequent fractions; n-hexane (Ph.Hex), chloroform (Ph.Chf), ethyl acetate (Ph.EtAc), n-Butanol (Ph.Bt), aqueous (Ph.Aq), saponins (Ph.Sp) were performed using the chick embryo chorioallantoic membrane (CAM) assay. Potato disc anti-tumor assay was performed on Agrobacterium tumefaciens containing tumor inducing plasmid. Cytotoxicity was performed against Artemia salina and mouse embryonic fibroblast NIH/3T3 cell line following contact toxicity and MTT cells viability assays, respectively. The GC-MS analysis of Ph.Cr, Ph.Hex, Ph.Chf, Ph.Bt, and Ph.EtAc identified 126, 124, 153, 131, and 164 compounds, respectively. In anti-angiogenic assay, Ph.Chf, Ph.Sp, Ph.EtAc, and Ph.Cr exhibited highest activity with IC50 of 28.65, 19.21, 88.75, and 461.53 μg/ml, respectively. In anti-tumor assay, Ph.Sp, Ph.Chf, Ph.EtAc, and Ph.Cr were most potent with IC50 of 18.39, 73.81, 217.19, and 342.53 μg/ml, respectively. In MTT cells viability assay, Ph.Chf, Ph.EtAc, Ph.Sp were most active causing 79.00, 72.50, and 71.50% cytotoxicity, respectively, at 1000 μg/ml with the LD50 of 140, 160, and 175 μg/ml, respectively. In overall study, Ph.Chf and Ph.Sp have shown overwhelming results which signifies their potentials as sources of therapeutic agents against cancer. PMID:27065865

  18. Insights in the etiopathology of galactosyltransferase II (GalT-II deficiency from transcriptome-wide expression profiling of skin fibroblasts of two sisters with compound heterozygosity for two novel B3GALT6 mutations

    Directory of Open Access Journals (Sweden)

    Marco Ritelli

    2015-03-01

    Full Text Available Mutations in B3GALT6, encoding the galactosyltransferase II (GalT-II involved in the synthesis of the glycosaminoglycan (GAG linkage region of proteoglycans (PGs, have recently been associated with a spectrum of connective tissue disorders, including spondyloepimetaphyseal dysplasia with joint laxity type 1 (SEMDJL1 and EhlersDanlos-like syndrome. Here, we report on two sisters compound heterozygous for two novel B3GALT6 mutations that presented with severe short stature and progressive kyphoscoliosis, joint hypermobility and laxity, hyperextensible skin, platyspondyly, short ilia, and elbow malalignment. Microarray-based transcriptome analysis revealed the differential expression of several genes encoding extracellular matrix (ECM structural components, including COMP, SPP1, COL5A1, and COL15A1, enzymes involved in GAG synthesis and in ECM remodeling, such as CSGALNACT1, CHPF, LOXL3, and STEAP4, signaling transduction molecules of the TGF?/BMP pathway, i.e., GDF6, GDF15, and BMPER, and transcription factors of the HOX and LIM families implicated in skeletal and limb development. Immunofluorescence analyses confirmed the down-regulated expression of some of these genes, in particular of the cartilage oligomeric matrix protein and osteopontin, encoded by COMP and SPP1, respectively, and showed the predominant reduction and disassembly of the heparan sulfate specific GAGs, as well as of the PG perlecan and type III and V collagens. The key role of GalT-II in GAG synthesis and the crucial biological functions of PGs are consistent with the perturbation of many physiological functions that are critical for the correct architecture and homeostasis of various connective tissues, including skin, bone, cartilage, tendons, and ligaments, and generates the wide phenotypic spectrum of GalT-II-deficient patients.

  19. Comparison of the irritation potentials of Boswellia serrata gum resin and of acetyl-11-keto-beta-boswellic acid by in vitro cytotoxicity tests on human skin-derived cell lines.

    Science.gov (United States)

    Burlando, Bruno; Parodi, Alessandro; Volante, Andrea; Bassi, Anna Maria

    2008-03-15

    Indian frankincense is a gum resin from Boswellia serrata of Burseraceae used in Ayurveda and Western medicine for the antinflammatory effects of boswellic acids, particularly 3-O-acetyl-11-keto-beta-boswellic acid (AKBA). We evaluated in vitro cytotoxicities of B. serrata extract and AKBA on differentiated and undifferentiated keratinocytes (HaCaT and NCTC 2544), and foetal dermal fibroblasts (HFFF2), using neutral red uptake (NRU), MTT, and DNA assays. Comparison between NRU and MTT, and between the extract and AKBA, suggested a relatively higher toxicity of both substances on lysosomes respect to mitochondria. Extract cytotoxicity on lysosomes was higher in NCTC and HFFF2 than on the more differentiated HaCaT. DNA assay showed low extract inhibition on HFFF2 proliferation, possibly due to lower growth rate, and a stronger effect on NCTC than on HaCaT, possibly related to higher proapoptotic effect on the less differentiated NCTC, as also suggested by higher AKBA toxicity on NCTC than on HaCaT. In general, gum resin and AKBA toxicities were slightly lower or higher than that of the reference compound SDS. Our in vitro model allowed to compare the sensitivities of different human skin cells to B. serrata, and indicated that the gum resin and AKBA exert moderate to low toxicity on the skin. PMID:18304763

  20. PED/PEA-15 Controls Fibroblast Motility and Wound Closure by ERK1/2-Dependent Mechanisms

    OpenAIRE

    Caserta, Sergio; Perruolo, Giuseppe; Vasaturo, Angela; Miele, Claudia; Buonomo, Roberta; Pagliara, Valentina; Guido, Stefano; Mansueto, Gelsomina; Beguinot, Francesco; Garbi, Corrado; Cassese, Angela; Oriente, Francesco; Formisano, Pietro; Giacco, Ferdinando

    2012-01-01

    Cell migration is dependent on the control of signaling events that play significant roles in creating contractile force and in contributing to wound closure. We evaluated wound closure in fibroblasts from mice overexpressing (TgPED) or lacking ped/pea-15 (KO), a gene overexpressed in patients with type 2 diabetes. Cultured skin fibroblasts isolated from TgPED mice showed a significant reduction in the ability to recolonize wounded area during scratch assay, compared to control fibroblasts. T...

  1. Purification of the migration stimulating factor produced by fetal and breast cancer patient fibroblasts

    International Nuclear Information System (INIS)

    The authors have previously shown that (i) human skin fibroblasts of fetal and adult origin display distinctive migratory phenotypes, (ii) this difference in cell behavior results from the production of a soluble migration stimulating factor (MSF) by fetal cells, and (iii) skin fibroblasts from breast cancer patients commonly resemble fetal fibroblasts both in migratory phenotype and in production of MSF. Data are now presented indicating that MSF present in the conditioned medium of fetal and cancer patient fibroblasts is precipitated at 10% saturation ammonium sulfate and binds to heparin and cation-exchange resins. Based on this information, they have devised a scheme for the purification of MSF involving the sequential application of ammonium sulfate precipitation, heparin affinity, gel filtration, and reverse-phase chromatography. Purified MSF has an estimated molecular mass of 70 kDa; amino acid analysis reveals a relatively high level of proline (13.34 residues per 100). The results further suggest that skin fibroblasts from breast cancer patients produce an additional factor with migration stimulating activity; this factor is precipitated at higher concentrations of ammonium sulfate and binds to anion-exchange resins. They have previously discussed the possible direct involvement of fetal-like fibroblasts in cancer pathogenesis. The availability of MSF obtained from cancer patient fibroblasts provides a potential means with which to examine the complex cellular interactions contributing to this process as well as develop a screening regime for identifying individuals at elevated risk of developing cancer

  2. GHK Peptide as a Natural Modulator of Multiple Cellular Pathways in Skin Regeneration.

    Science.gov (United States)

    Pickart, Loren; Vasquez-Soltero, Jessica Michelle; Margolina, Anna

    2015-01-01

    GHK (glycyl-L-histidyl-L-lysine) is present in human plasma, saliva, and urine but declines with age. It is proposed that GHK functions as a complex with copper 2+ which accelerates wound healing and skin repair. GHK stimulates both synthesis and breakdown of collagen and glycosaminoglycans and modulates the activity of both metalloproteinases and their inhibitors. It stimulates collagen, dermatan sulfate, chondroitin sulfate, and the small proteoglycan, decorin. It also restores replicative vitality to fibroblasts after radiation therapy. The molecule attracts immune and endothelial cells to the site of an injury. It accelerates wound-healing of the skin, hair follicles, gastrointestinal tract, boney tissue, and foot pads of dogs. It also induces systemic wound healing in rats, mice, and pigs. In cosmetic products, it has been found to tighten loose skin and improve elasticity, skin density, and firmness, reduce fine lines and wrinkles, reduce photodamage, and hyperpigmentation, and increase keratinocyte proliferation. GHK has been proposed as a therapeutic agent for skin inflammation, chronic obstructive pulmonary disease, and metastatic colon cancer. It is capable of up- and downregulating at least 4,000 human genes, essentially resetting DNA to a healthier state. The present review revisits GHK's role in skin regeneration in the light of recent discoveries. PMID:26236730

  3. Basal Cell Carcinoma in Gorlin's Patients: a Matter of Fibroblasts-Led Protumoral Microenvironment?

    Science.gov (United States)

    Gache, Yannick; Brellier, Florence; Rouanet, Sophie; Al-Qaraghuli, Sahar; Goncalves-Maia, Maria; Burty-Valin, Elodie; Barnay, Stéphanie; Scarzello, Sabine; Ruat, Martial; Sevenet, Nicolas; Avril, Marie-Françoise; Magnaldo, Thierry

    2015-01-01

    Basal cell carcinoma (BCC) is the commonest tumor in human. About 70% sporadic BCCs bear somatic mutations in the PATCHED1 tumor suppressor gene which encodes the receptor for the Sonic Hedgehog morphogen (SHH). PATCHED1 germinal mutations are associated with the dominant Nevoid Basal Cell Carcinoma Syndrome (NBCCS), a major hallmark of which is a high susceptibility to BCCs. Although the vast majority of sporadic BCCs arises exclusively in sun exposed skin areas, 40 to 50% BCCs from NBCCS patients develop in non photo-exposed skin. Since overwhelming evidences indicate that microenvironment may both be modified by- and influence the- epithelial tumor, we hypothesized that NBCCS fibroblasts could contribute to BCCs in NBCCS patients, notably those developing in non photo-exposed skin areas. The functional impact of NBCCS fibroblasts was then assessed in organotypic skin cultures with control keratinocytes. Onset of epidermal differentiation was delayed in the presence of primary NBCCS fibroblasts. Unexpectedly, keratinocyte proliferation was severely reduced and showed high levels of nuclear P53 in both organotypic skin cultures and in fibroblast-led conditioning experiments. However, in spite of increased levels of senescence associated β-galactosidase activity in keratinocytes cultured in the presence of medium conditioned by NBCCS fibroblasts, we failed to observe activation of P16 and P21 and then of bona fide features of senescence. Constitutive extinction of P53 in WT keratinocytes resulted in an invasive phenotype in the presence of NBCCS fibroblasts. Finally, we found that expression of SHH was limited to fibroblasts but was dependent on the presence of keratinocytes. Inhibition of SHH binding resulted in improved epidermal morphogenesis. Altogether, these data suggest that the repertoire of diffusible factors (including SHH) expressed by primary NBCCS fibroblasts generate a stress affecting keratinocytes behavior and epidermal homeostasis. Our findings suggest that defects in dermo/epidermal interactions could contribute to BCC susceptibility in NBCCS patients. PMID:26694869

  4. Basal Cell Carcinoma in Gorlin’s Patients: a Matter of Fibroblasts-Led Protumoral Microenvironment?

    Science.gov (United States)

    Gache, Yannick; Brellier, Florence; Rouanet, Sophie; Al-Qaraghuli, Sahar; Goncalves-Maia, Maria; Burty-Valin, Elodie; Barnay, Stéphanie; Scarzello, Sabine; Ruat, Martial; Sevenet, Nicolas; Avril, Marie-Françoise; Magnaldo, Thierry

    2015-01-01

    Basal cell carcinoma (BCC) is the commonest tumor in human. About 70% sporadic BCCs bear somatic mutations in the PATCHED1 tumor suppressor gene which encodes the receptor for the Sonic Hedgehog morphogen (SHH). PATCHED1 germinal mutations are associated with the dominant Nevoid Basal Cell Carcinoma Syndrome (NBCCS), a major hallmark of which is a high susceptibility to BCCs. Although the vast majority of sporadic BCCs arises exclusively in sun exposed skin areas, 40 to 50% BCCs from NBCCS patients develop in non photo-exposed skin. Since overwhelming evidences indicate that microenvironment may both be modified by- and influence the- epithelial tumor, we hypothesized that NBCCS fibroblasts could contribute to BCCs in NBCCS patients, notably those developing in non photo-exposed skin areas. The functional impact of NBCCS fibroblasts was then assessed in organotypic skin cultures with control keratinocytes. Onset of epidermal differentiation was delayed in the presence of primary NBCCS fibroblasts. Unexpectedly, keratinocyte proliferation was severely reduced and showed high levels of nuclear P53 in both organotypic skin cultures and in fibroblast-led conditioning experiments. However, in spite of increased levels of senescence associated β-galactosidase activity in keratinocytes cultured in the presence of medium conditioned by NBCCS fibroblasts, we failed to observe activation of P16 and P21 and then of bona fide features of senescence. Constitutive extinction of P53 in WT keratinocytes resulted in an invasive phenotype in the presence of NBCCS fibroblasts. Finally, we found that expression of SHH was limited to fibroblasts but was dependent on the presence of keratinocytes. Inhibition of SHH binding resulted in improved epidermal morphogenesis. Altogether, these data suggest that the repertoire of diffusible factors (including SHH) expressed by primary NBCCS fibroblasts generate a stress affecting keratinocytes behavior and epidermal homeostasis. Our findings suggest that defects in dermo/epidermal interactions could contribute to BCC susceptibility in NBCCS patients. PMID:26694869

  5. Biosynthesis of collagen by fibroblasts kept in culture

    International Nuclear Information System (INIS)

    The sinthesis of collagen is studied in fibroblasts of different origins with the purpose of obtaining an appropriate system for the study of its biosynthesis and processing. The percentage of collagen synthesis vary according to the fibroblast origin. Experiences are performed with fibroblasts kept in culture from: chicken - and guinea pig embryos, carragheenin - induced granulomas in adult guinea pig and from human skin. The collagen pattern synthesized after acetic acid - or saline extractions in the presence of inhibitors is also determined. This pattern is then assayed by poliacrilamide - 5% - SDS gel electrophoresis accompanied by fluorography. The importance of the cell culture system in the elucidation of collagen biosynthesis is pointed out. (M.A.)

  6. Salmonella enterica Serovar Typhimurium Invades Fibroblasts by Multiple Routes Differing from the Entry into Epithelial Cells▿

    OpenAIRE

    Aiastui, Ana; Pucciarelli, M. Graciela; García-del Portillo, Francisco

    2010-01-01

    Fibroblasts are ubiquitous cells essential to tissue homeostasis. Despite their nonphagocytic nature, fibroblasts restrain replication of intracellular bacterial pathogens such as Salmonella enterica serovar Typhimurium. The extent to which the entry route of the pathogen determines this intracellular response is unknown. Here, we analyzed S. Typhimurium invasion in fibroblasts obtained from diverse origins, including primary cultures and stable nontransformed cell lines derived from normal t...

  7. Cadherin-11 regulates fibroblast inflammation

    OpenAIRE

    Chang, Sook Kyung; Noss, Erika H.; Chen, Mei; Gu, Zhizhan; Townsend, Kirk; Grenha, Rosa; de Leon, Luis; LEE, SOO YOUNG; Lee, David M.; Brenner, Michael B

    2011-01-01

    Fibroblasts are important participants in inflammation. Although not leukocytes, their capacity to produce cytokines, chemokines, and other inflammatory factors locally in tissues suggests that they can contribute to inflammatory diseases. For example, fibroblasts in a rheumatoid arthritis (RA) joint are a dominant source of IL-6 and RANKL in the synovium, both of which are therapeutic targets for inflammation and bone erosion. Previously, we found that fibroblasts can be targeted by mAb dire...

  8. Sagging Skin

    Science.gov (United States)

    ... Dermatologic Surgery Expertise for the life of your skin (TM) Public Resources Dermatologic Surgery Conditions Treatments and ... learn-more"> 3 million Americans are diagnosed with skin cancer every year Any suspicious new growths or ...

  9. Skin Complications

    Science.gov (United States)

    ... Text Size: A A A Listen En Español Skin Complications Diabetes can affect every part of the ... lipoidica diabeticorum, diabetic blisters, and eruptive xanthomatosis. General Skin Conditions Bacterial Infections Several kinds of bacterial infections ...

  10. Skin Cancer

    Science.gov (United States)

    Skin cancer is the most common form of cancer in the United States. The two most common types ... face, neck, hands, and arms. Another type of skin cancer, melanoma, is more dangerous but less common. Anyone ...

  11. Skin Aging

    Science.gov (United States)

    ... too. Sunlight is a major cause of skin aging. You can protect yourself by staying out of ... person has smoked. Many products claim to revitalize aging skin or reduce wrinkles, but the Food and ...

  12. Skin turgor

    Science.gov (United States)

    ... up during a check. This can indicate severe dehydration that needs quick treatment. You have reduced skin turgor and are unable ... Urinalysis Intravenous fluids may be needed for severe ... treat other conditions that affect skin turgor and elasticity.

  13. Repair of DNA strand breaks in progeric fibroblasts and aging human diploid cells

    International Nuclear Information System (INIS)

    The rate of rejoining of DNA strand breaks induced by 10 krad of γ-irradiation has been studied in normal human diploid skin fibroblasts and skin fibroblasts from six patients with symptoms of progeria. Although slightly more rapid in very early passage, the repair rate in normal cells was similar throughout most of their life span in vitro. The appearance of cells with reduced repair capacity was evident as the cultures became senescent. The progeric fibroblasts varied greatly in their response to irradiation. The rate of repair was greatly reduced in two strains, whereas in two others extensive DNA degradation was consistently observed in unirradiated cells. Degradation was apparently related to the radiation received from the incorporated radiolabel. Normal repair was seen in progeric fibroblasts transformed by SV40 virus

  14. Induction of pluripotent stem cells from fetal and adult cynomolgus monkey fibroblasts using four human transcription factors.

    Science.gov (United States)

    Okahara-Narita, Junko; Umeda, Rieko; Nakamura, Shinichiro; Mori, Takahide; Noce, Toshiaki; Torii, Ryuzo

    2012-04-01

    Induced pluripotent stem (iPS) cells have the potential to become a universal resource for cell-based therapies in regenerative medicine; however, prior to the use of such iPS cell-based therapies, preclinical assessment of their safety and efficacy is essential. Non-human primates serve as valuable animal models for human diseases or biomedical research; therefore, in this study, we generated cynomolgus monkey iPS cells from adult skin and fetal fibroblast cells by the retrovirally mediated introduction of four human transcription factors: c-Myc, Klf4, Oct3/4, and Sox2 (the so-called "Yamanaka factors"). Twenty to 30 days after the introduction of these factors, several cynomolgus monkey embryonic stem (ES) cell-like colonies appeared on SNL and mouse embryonic fibroblast (MEF) feeder layers. These colonies were picked and cultivated in primate ES medium. Seven iPS cell lines were established, and we detected the expression of pluripotent markers that are also expressed in ES cells. Reverse transcription polymerase chain reaction (PCR) showed that these iPS cells expressed endogenous c-Myc, Klf4, Oct3/4, and Sox2 genes, whereas several transgenes were silenced. Embryoid body and teratoma formation showed that the cynomolgus iPS cells had the developmental potential to differentiate into cells of all three primary germ layers. In summary, we generated cynomolgus monkey iPS cells by retrovirus-mediated transduction of the human transcription factors, c-Myc, Klf4, Oct3/4, and Sox2 into adult cynomolgus monkey skin cells and fetal fibroblasts. The cynomolgus monkey is the most relevant primate model for human disease, and the highly efficient generation of monkey iPS cells would allow investigation of the treatments of various diseases in this model via therapeutic cloning. PMID:22075965

  15. Age-related skin changes

    Directory of Open Access Journals (Sweden)

    Božanić Snežana

    2012-01-01

    Full Text Available Age-related skin changes can be induced by chronological ageing, manifested in subcutaneous fat reduction, and photo-ageing eliciting increased elastotic substance in the upper dermis, destruction of its fibrilar structure, augmented intercellular substance and moderate inflammatory infiltrate. Forty-five biopsy skin samples of the sun-exposed and sun-protected skin were analyzed. The patients were both males and females, aged from 17 to 81 years. The thickness of the epidermal layers and the number of cellular living layers is greater in younger skin. The amount of keratohyaline granules is enlarged in older skin. Dermoepidermal junction is flattened and the presence of elastotic material in the dermis is pronounced with age. The amount of inflammatory infiltrate is increased, the fibrous trabeculae are thickened in older skin and the atrophy of the hypodermis is observed. Chronological ageing alters the fibroblasts metabolism by reducing their life span, capacity to divide and produce collagen. During ageing, the enlargement of collagen fibrils diminishes the skin elasticity.

  16. Dermal Wound Fibroblasts and Matrix Metaloproteinases (MMPs): Their Possible Role in Allergic Contact Dermatitis

    OpenAIRE

    Mohammad Reza Khorramizadeh; Reza Falak; Mohammad Pezeshki; Farnaz Safavifar; Parvin Mansouri; Aziz Ghahary; Farshid Saadat; Kamran Varshokar

    2004-01-01

    This study was conducted to examine if allergic contact dermatitis (ACD) alters the expression of MMPs in human dermal fibroblasts. Fibroblasts are the primary source for MMP and matrix production in skin. MMPs are known to involve in a number of physiological and pathological processes. Some published data indicated a gelatinase-like activity in acute and chronic phases of allergic contact dermatitis. However, no exact source of gelatinase activity was demonstrated. Moreover, little is known...

  17. Streptococcus pneumoniae cocultured with fibroblasts enhances both interferon production and cytotoxic activity by lymphocytes.

    OpenAIRE

    Weigent, D A; Baron, S; Stanton, G. J.

    1985-01-01

    Cell-mediated cytotoxicity against normal human fibroblasts was dependent on treatment of the fibroblasts with Streptococcus pneumoniae. Both spontaneous and interferon (IFN)-enhanced lymphocytes killed human foreskin (HFS) or skin muscle cells cocultured with S. pneumoniae five- to eightfold more than control nontreated cells. Based on Percoll gradient centrifugation, the cytotoxic effector cell migrated like a large granular lymphocyte. The human IFN produced from mixtures of HFS cells, lym...

  18. Radioprotection by glutathione ester: transport of glutathione ester into human lymphoid cells and fibroblasts.

    OpenAIRE

    Wellner, V P; Anderson, M E; Puri, R N; JENSEN, G.L.; Meister, A.

    1984-01-01

    Glutathione is not effectively transported into human lymphoid cells, normal human skin fibroblasts, and fibroblasts from patients with genetic deficiencies of gamma-glutamylcysteine synthetase or glutathione synthetase. On the other hand, the monoethyl ester of glutathione, in which the carboxyl group of the glycine residue is esterified, is readily transported into these cells and is hydrolyzed intracellularly. This leads to greatly increased cellular levels of glutathione, which often exce...

  19. The Role of the Skin Barrier in Occupational Skin Diseases.

    Science.gov (United States)

    Kasemsarn, Pranee; Bosco, Joanna; Nixon, Rosemary L

    2016-01-01

    Occupational skin diseases (OSDs) are the second most common occupational diseases worldwide. Occupational contact dermatitis (OCD) is the most frequent OSD, and comprises irritant contact dermatitis (ICD), allergic contact dermatitis (ACD), contact urticaria and protein contact dermatitis. There are many endogenous and exogenous factors which affect the development of OCD, including age, sex, ethnicity, atopic skin diathesis, certain occupations and environmental factors. One of the most important contributing causes is skin barrier dysfunction. The skin provides a first-line defense from environmental assaults and incorporates physical, chemical and biological protection. Skin barrier disturbance plays a crucial role in various skin diseases such as atopic dermatitis (AD), ichthyosis, ICD and ACD. Genetic factors, such as filaggrin gene (FLG) mutations, and external factors, such as skin irritants interfering with stratum corneum structure and composition, may lead to abnormalities in skin barrier function and increased vulnerability to skin diseases. FLG encodes the cornified envelope protein, filaggrin, which is involved in skin barrier function. FLG mutation is associated with the development of OCD. High-risk occupations for OCD include health care workers, hairdressers and construction workers. There are often multiple contributing causes to OCD, as workers are exposed to both irritants and allergens. AD is also associated with skin barrier disruption and plays an important role in OCD. ICD often precedes and facilitates the development of ACD, with impairment of the skin barrier contributing to the concurrence of ICD and ACD in many workers with OCD. PMID:26844905

  20. Skin decontamination

    International Nuclear Information System (INIS)

    The problem matter is briefly characterized of skin damage by irradiation and of the binding of radioactive materials to the skin. The results are summed up of some experiments aimed at skin resorption of various radioactive materials. Main attention is devoted to methods of skin decontamination. The principles are described in detail of the operation of the so-called hygiene loops where complete decontamination of personnel proceeds. The organizational chart of such a unit is presented in figure. Information is also presented on the composition of solutions for skin decontamination. (Z.M.)

  1. Sensitive skin.

    Science.gov (United States)

    Misery, L; Loser, K; Ständer, S

    2016-02-01

    Sensitive skin is a clinical condition defined by the self-reported facial presence of different sensory perceptions, including tightness, stinging, burning, tingling, pain and pruritus. Sensitive skin may occur in individuals with normal skin, with skin barrier disturbance, or as a part of the symptoms associated with facial dermatoses such as rosacea, atopic dermatitis and psoriasis. Although experimental studies are still pending, the symptoms of sensitive skin suggest the involvement of cutaneous nerve fibres and neuronal, as well as epidermal, thermochannels. Many individuals with sensitive skin report worsening symptoms due to environmental factors. It is thought that this might be attributed to the thermochannel TRPV1, as it typically responds to exogenous, endogenous, physical and chemical stimuli. Barrier disruptions and immune mechanisms may also be involved. This review summarizes current knowledge on the epidemiology, potential mechanisms, clinics and therapy of sensitive skin. PMID:26805416

  2. Accelerated telomere shortening and replicative senescence in human fibroblasts overexpressing mutant and wild-type lamin A

    International Nuclear Information System (INIS)

    LMNA mutations are responsible for a variety of genetic disorders, including muscular dystrophy, lipodystrophy, and certain progeroid syndromes, notably Hutchinson-Gilford Progeria. Although a number of clinical features of these disorders are suggestive of accelerated aging, it is not known whether cells derived from these patients exhibit cellular phenotypes associated with accelerated aging. We examined a series of isogenic skin fibroblast lines transfected with LMNA constructs bearing known pathogenic point mutations or deletion mutations found in progeroid syndromes. Fibroblasts overexpressing mutant lamin A exhibited accelerated rates of loss of telomeres and shortened replicative lifespans, in addition to abnormal nuclear morphology. To our surprise, these abnormalities were also observed in lines overexpressing wild-type lamin A. Copy number variants are common in human populations; those involving LMNA, whether arising meiotically or mitotically, might lead to progeroid phenotypes. In an initial pilot study of 23 progeroid cases without detectable WRN or LMNA mutations, however, no cases of altered LMNA copy number were detected. Nevertheless, our findings raise a hypothesis that changes in lamina organization may cause accelerated telomere attrition, with different kinetics for overexpession of wild-type and mutant lamin A, which leads to rapid replicative senescence and progroid phenotypes

  3. Human skin equivalent as an alternative to animal testing

    OpenAIRE

    Brunner, Herwig; Kersen, Silke; Weimer, Michaela; Mertsching, Heike

    2008-01-01

    The 3-D skin equivalent can be viewed as physiologically comparable to the natural skin and therefore is a suitable alternative for animal testing. This highly differentiated in vitro human skin equivalent is used to assess the efficacy and mode of action of novel agents. This model is generated from primary human keratinocytes on a collagen substrate containing human dermal fibroblasts. It is grown at the air-liquid interface which allows full epidermal stratification and epidermal-dermal in...

  4. Skin manifestations in a case of trisomy 16 mosaicism

    DEFF Research Database (Denmark)

    Ousager, Lilian Bomme; Brandrup, Flemming; Andersen, Charlotte Brasch; Erlandsson, Anne

    2006-01-01

    We present a 48-year-old man with unilateral dermatological manifestations including hypertrichosis, telangiectasia, hyperkeratosis and hyperpigmentation. Additional findings included skeletal abnormalities and left-sided hearing loss. Skin biopsies showed changes characteristic of porokeratosis....... Fibroblast karyotyping from affected skin demonstrated trisomy 16 mosaicism, in contrast to the normal karyotype in unaffected skin and blood lymphocytes. The possible role of trisomy 16 in porokeratosis is discussed....

  5. Gene targeting in adult rhesus macaque fibroblasts

    Directory of Open Access Journals (Sweden)

    Wolf Don P

    2008-03-01

    Full Text Available Abstract Background Gene targeting in nonhuman primates has the potential to produce critical animal models for translational studies related to human diseases. Successful gene targeting in fibroblasts followed by somatic cell nuclear transfer (SCNT has been achieved in several species of large mammals but not yet in primates. Our goal was to establish the protocols necessary to achieve gene targeting in primary culture of adult rhesus macaque fibroblasts as a first step in creating nonhuman primate models of genetic disease using nuclear transfer technology. Results A primary culture of adult male fibroblasts was transfected with hTERT to overcome senescence and allow long term in vitro manipulations. Successful gene targeting of the HPRT locus in rhesus macaques was achieved by electroporating S-phase synchronized cells with a construct containing a SV40 enhancer. Conclusion The cell lines reported here could be used for the production of null mutant rhesus macaque models of human genetic disease using SCNT technology. In addition, given the close evolutionary relationship and biological similarity between rhesus macaques and humans, the protocols described here may prove useful in the genetic engineering of human somatic cells.

  6. SKIN AND HAIR CHANGES AFTER FORTY

    Directory of Open Access Journals (Sweden)

    Manisha

    2014-04-01

    Full Text Available Aging is a continuous, dynamic, and an irreversible process. Direct exposure to ultra-violet radiations, skin is particularly prone to early aging, known as photo aging. Skin aging is particularly important because of its visibility and social impact. As women age we will notice changes to our skin and hair during the menopause. Dry, thinning, fragile, less tolerant and sagging skin are common complaints. The main reasons for the change in skin is the loss of estrogen, testosterone and dehydroepiandrosterone (DHEA etc, 1, 2, 3 from the age of 35 onwards up to menopause, the more we have had long-term exposure to the elements, such as sun and wind the more this becomes evident. Estrogen is very involved in the normal function of the skin. It directly affects the function of key cells in the skin, like the fibroblast (produces collagen and elastin, keratinocyte (closely involved in skin protection and melanocytes (involved in evenness of skin color, etc.. It also helps regulate hair follicle function (hair production as well as sebaceous gland activity (producing skin oils. After the age of forty most of women enters menopause, during which estrogens levels decreases, which leads to different types of hair and skin changes which has been described in this article.

  7. Excision of gamma-ray induced thymine lesions by preparations from ataxia telangiectasia fibroblasts

    International Nuclear Information System (INIS)

    The capacity of whole cell sonicates of skin fibroblasts of normal individuals and patients with the autosomal recessive disease Ataxia telangiectasia (AT) to remove aerobic gamma-ray products of the 5,6-dihydroxydihydrothymine type (tsub(O2)sup(?)) from exogenous DNA substrates was investigated. All four AT strains (AT CRL 1312, AT CRL 1343, AT GM 367, AT 4BI) possessed normal capabilities to excise tsub(O2)sup(?) from irradiated bacteriophage DNA and irradiated chromatin isolated from normal and AT-skin fibroblasts

  8. Naïve adult stem cells isolation from primary human fibroblast cultures.

    Science.gov (United States)

    Wenzel, Vera; Roedl, Daniela; Ring, Johannes; Djabali, Karima

    2013-01-01

    Over the last decade, several adult stem cell populations have been identified in human skin (1-4). The isolation of multipotent adult dermal precursors was first reported by Miller F. D laboratory (5, 6). These early studies described a multipotent precursor cell population from adult mammalian dermis (5). These cells--termed SKPs, for skin-derived precursors-- were isolated and expanded from rodent and human skin and differentiated into both neural and mesodermal progeny, including cell types never found in skin, such as neurons (5). Immunocytochemical studies on cultured SKPs revealed that cells expressed vimentin and nestin, an intermediate filament protein expressed in neural and skeletal muscle precursors, in addition to fibronectin and multipotent stem cell markers (6). Until now, the adult stem cells population SKPs have been isolated from freshly collected mammalian skin biopsies. Recently, we have established and reported that a population of skin derived precursor cells could remain present in primary fibroblast cultures established from skin biopsies (7). The assumption that a few somatic stem cells might reside in primary fibroblast cultures at early population doublings was based upon the following observations: (1) SKPs and primary fibroblast cultures are derived from the dermis, and therefore a small number of SKP cells could remain present in primary dermal fibroblast cultures and (2) primary fibroblast cultures grown from frozen aliquots that have been subjected to unfavorable temperature during storage or transfer contained a small number of cells that remained viable (7). These rare cells were able to expand and could be passaged several times. This observation suggested that a small number of cells with high proliferation potency and resistance to stress were present in human fibroblast cultures (7). We took advantage of these findings to establish a protocol for rapid isolation of adult stem cells from primary fibroblast cultures that are readily available from tissue banks around the world (Figure 1). This method has important significance as it allows the isolation of precursor cells when skin samples are not accessible while fibroblast cultures may be available from tissue banks, thus, opening new opportunities to dissect the molecular mechanisms underlying rare genetic diseases as well as modeling diseases in a dish. PMID:23685623

  9. Fibroblastic rheumatism: an addition to fibromatosis.

    Science.gov (United States)

    Ji, Lanlan; Geng, Yan; Hao, Yanjie; Zhang, Zhuoli

    2011-10-01

    Fibroblastic rheumatism (FR) is a rare rheumatologic entity of unknown etiology. It is characterized by symmetrical polyarthritis associated with multiple cutaneous nodules. Bone erosion can occur as the disease progresses and destructive arthropathy is not rare. We report on an 18-year-old man with FR who presented a 6-year history of cutaneous nodules localized at para-articular sites with only minimal oligoarthralgia on exertion. There was no visceral involvement, and all the routine and immunological tests were normal. The diagnosis of FR was confirmed by histological examination of a nodule, which composed of myofibroblastic proliferation and thickened collagen fibers. Most skin lesions resolved after treated with IFN-α, however there was sequelae of permanent disability due to the progressive bone erosion despite weekly methotrexate treatment. PMID:21549631

  10. The redox status of cystinotic fibroblasts.

    Science.gov (United States)

    Vitvitsky, Victor; Witcher, Marc; Banerjee, Ruma; Thoene, Jess

    2010-04-01

    A key unresolved question in the pathogenesis of phenotype development in nephropathic cystinosis is whether intralysosomal cystine, the hallmark of this lethal inborn error of metabolism, alters cytoplasmic redox potential. Variable findings on this issue have been reported. This study of fetal and non-fetal skin and lung-derived cystinotic fibroblasts compared to origin and age-matched normal control fibroblasts reveals that cystinotic cells do not exhibit redox perturbations. We find that the steady-state redox status as assessed by the [GSH]/[GSSG] ratio, an indicator of the intracellular redox poise, is unchanged in cystinotic cells. Furthermore, the dependence of the intracellular GSH and cysteine pool sizes and the [GSH]/[GSSG] ratio are similarly dependent on the two major sources of cysteine, i.e. the transsulfuration pathway and the plasma membrane cystine transporter, xc(-), in both cystinotic and control cells, and the presence of lysosomal cystine has no measurable effect on the redox status of these cells. Hence, mechanisms other than cytosolic redox perturbations are involved in the etiology of nephropathic cystinosis. PMID:20061170

  11. Effects of lunar and mars dust simulants on HaCaT keratinocytes and CHO-K1 fibroblasts

    Science.gov (United States)

    Rehders, Maren; Grosshuser, Bianka B.; Smarandache, Anita; Sadhukhan, Annapurna; Mirastschijski, Ursula; Kempf, Jrgen; Dnne, Matthias; Slenzka, Klaus; Brix, Klaudia

    2011-04-01

    Exposure to lunar dust during Apollo missions resulted in occasional reports of ocular, respiratory and dermal irritations which showed that lunar dust has a risk potential for human health. This is caused by its high reactivity as well as its small size, leading to a wide distribution also inside habitats. Hence, detailed information regarding effects of extraterrestrial lunar dusts on human health is required to best support future missions to moon, mars or other destinations. In this study, we used several methods to assess the specific effects of extraterrestrial dusts onto mammalian skin by exposing HaCaT keratinocytes and CHO-K1 fibroblasts to dusts simulating lunar or mars soils. These particular cell types were chosen because the skin protects the human body from potentially harmful substances and because a well orchestrated program ensures proper wound healing. Keratinocytes and fibroblasts were exposed to the dusts for different durations of time and their effects on morphology and viability of the cells were determined. Cytotoxicity was measured using the MTT assay and by monitoring culture impedance, while phalloidin staining of the actin cytoskeleton was performed to address structural integrity of the cells which was also investigated by propidium iodide intake. It was found that the effects of the two types of dust simulants on the different features of both cell lines varied to a considerable extent. Moreover, proliferation of HaCaT keratinocytes, as analyzed by Ki67 labeling, was suppressed in sub-confluent cultures exposed to lunar dust simulant. Furthermore, experimental evidence is provided for a delay in regeneration of keratinocyte monolayers from scratch-wounding when exposed to lunar dust simulant. The obtained results will facilitate further investigations of dust exposure during wound healing and will ease risk assessment studies e.g., for lunar lander approaches. The investigations will help to determine safety measures to be taken during extraterrestrial expeditions in order to minimize risks to human health associated with exposure of human skin to dust contaminants.

  12. Development of nanofibrous scaffolds containing gum tragacanth/poly (ε-caprolactone) for application as skin scaffolds

    International Nuclear Information System (INIS)

    Outstanding wound healing activity of gum tragacanth (GT) and higher mechanical strength of poly (ε-caprolactone) (PCL) may produce an excellent nanofibrous patch for either skin tissue engineering or wound dressing application. PCL/GT scaffold containing different concentrations of PCL with different blend ratios of GT/PCL was produced using 90% acetic acid as solvent. The results demonstrated that the PCL/GT (3:1.5) with PCL concentration of 20% (w/v) produced nanofibers with proper morphology. Scanning electron microscopy (SEM) and differential scanning calorimetry (DSC) were utilized to characterize the nanofibers. Surface wettability, functional groups analysis, porosity and tensile properties of nanofibers were evaluated. Morphological characterization showed that the addition of GT to PCL solution results in decreasing the average diameter of the PCL/GT nanofibers. However, the hydrophilicity increased in the PCL/GT nanofibers. Slight increase in melting peaks was observed due to the blending of PCL with GT nanofibers. PCL/GT nanofibers were used for in vitro cell culture of human fibroblast cell lines AGO and NIH 3T3 fibroblast cells. MTT assay and SEM results showed that the biocomposite PCL/GT mats enhanced the fibroblast adhesion and proliferation compared to PCL scaffolds. The antibacterial activity of PCL/GT and GT nanofibers against Staphylococcus aureus and Pseudomonas aeruginosa was also examined. - Highlights: • A new skin tissue engineering scaffold from poly (ε-caprolactone) (PCL) and gum tragacanth (GT) has been developed. • These scaffolds might be an effectual simulator of the structure and composition of native skin. • Very slight increase in melting peaks was observed due to the blending of PCL with GT nanofibers. • Biodegradation, water uptake and hydrophilicity properties of these scaffolds showed that produced scaffolds were adherent. • The electrospun PCL/GT scaffold can promote the skin regeneration of full-thickness-injured tissues

  13. Development of nanofibrous scaffolds containing gum tragacanth/poly (ε-caprolactone) for application as skin scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Ranjbar-Mohammadi, Marziyeh [Textile Engineering Department, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Bahrami, S. Hajir, E-mail: hajirb@aut.ac.ir [Textile Engineering Department, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Center for excellence Modern Textile Characterization, Tehran (Iran, Islamic Republic of)

    2015-03-01

    Outstanding wound healing activity of gum tragacanth (GT) and higher mechanical strength of poly (ε-caprolactone) (PCL) may produce an excellent nanofibrous patch for either skin tissue engineering or wound dressing application. PCL/GT scaffold containing different concentrations of PCL with different blend ratios of GT/PCL was produced using 90% acetic acid as solvent. The results demonstrated that the PCL/GT (3:1.5) with PCL concentration of 20% (w/v) produced nanofibers with proper morphology. Scanning electron microscopy (SEM) and differential scanning calorimetry (DSC) were utilized to characterize the nanofibers. Surface wettability, functional groups analysis, porosity and tensile properties of nanofibers were evaluated. Morphological characterization showed that the addition of GT to PCL solution results in decreasing the average diameter of the PCL/GT nanofibers. However, the hydrophilicity increased in the PCL/GT nanofibers. Slight increase in melting peaks was observed due to the blending of PCL with GT nanofibers. PCL/GT nanofibers were used for in vitro cell culture of human fibroblast cell lines AGO and NIH 3T3 fibroblast cells. MTT assay and SEM results showed that the biocomposite PCL/GT mats enhanced the fibroblast adhesion and proliferation compared to PCL scaffolds. The antibacterial activity of PCL/GT and GT nanofibers against Staphylococcus aureus and Pseudomonas aeruginosa was also examined. - Highlights: • A new skin tissue engineering scaffold from poly (ε-caprolactone) (PCL) and gum tragacanth (GT) has been developed. • These scaffolds might be an effectual simulator of the structure and composition of native skin. • Very slight increase in melting peaks was observed due to the blending of PCL with GT nanofibers. • Biodegradation, water uptake and hydrophilicity properties of these scaffolds showed that produced scaffolds were adherent. • The electrospun PCL/GT scaffold can promote the skin regeneration of full-thickness-injured tissues.

  14. Curious Skin

    OpenAIRE

    Angel, G

    2010-01-01

    Some of Henry Wellcome’s collection of tattoos on human skin will be on display in our forthcoming Skin exhibition. But how did the Parisian doctor from whom they were acquired come by his macabre collection of tattoos in the first place, and what did they mean to those whose skin they were on? It’s Gemma Angel‘s job to find out…

  15. The effect of pantothenic acid deficiency on keratinocyte proliferation and the synthesis of keratinocyte growth factor and collagen in fibroblasts.

    Science.gov (United States)

    Kobayashi, Daisaku; Kusama, Miho; Onda, Masaaki; Nakahata, Norimichi

    2011-01-01

    It has been reported that pantothenic acid (vitamin B5) and panthenol, an alcohol derivative of pantothenic acid, have beneficial moisturizing effects on the skin. However, few studies have investigated the mechanism of action of pantothenic acid on skin tissues. We tried to clarify the role of pantothenic acid on skin function by using keratinocytes and fibroblasts. The depletion of pantothenic acid from the culture medium suppressed keratinocyte proliferation and promoted differentiation. Moreover, pantothenic acid depletion decreased the synthesis of keratinocyte growth factor and procollagen 4a2 in fibroblasts. These results suggest that pantothenic acid is essential for maintaining keratinocyte proliferation and differentiation. PMID:21258175

  16. Peripheral arterial line (image)

    Science.gov (United States)

    A peripheral arterial line is a small, short plastic catheter placed through the skin into an artery of the arm or leg. The purpose of a peripheral arterial line is to allow continuous monitoring of blood pressure ...

  17. Irradiated fibroblasts promote epithelial–mesenchymal transition and HDGF expression of esophageal squamous cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Bao, Ci-Hang; Wang, Xin-Tong [Department of Radiation Oncology, Qilu Hospital of Shandong University, Jinan 250012 (China); Ma, Wei [Department of Radiation Oncology, Cancer Hospital, Genaral Hospital of Ningxia Medical University, Yinchuan 750000 (China); Wang, Na-Na; Nesa, Effat un; Wang, Jian-Bo; Wang, Cong; Jia, Yi-Bin; Wang, Kai [Department of Radiation Oncology, Qilu Hospital of Shandong University, Jinan 250012 (China); Tian, Hui [Department of Thoracic Surgery, Qilu Hospital of Shandong University, Jinan 250012 (China); Cheng, Yu-Feng, E-mail: qlcyf1965@126.com [Department of Radiation Oncology, Qilu Hospital of Shandong University, Jinan 250012 (China)

    2015-03-06

    Recent evidence suggested that nonirradiated cancer-associated fibroblasts (CAFs) promoted aggressive phenotypes of cancer cells through epithelial–mesenchymal transition (EMT). Hepatoma-derived growth factor (HDGF) is a radiosensitive gene of esophageal squamous cell carcinoma (ESCC). This study aimed to investigate the effect of irradiated fibroblasts on EMT and HDGF expression of ESCC. Our study demonstrated that coculture with nonirradiated fibroblasts significantly increased the invasive ability of ESCC cells and the increased invasiveness was further accelerated when they were cocultured with irradiated fibroblasts. Scattering of ESCC cells was also accelerated by the supernatant from irradiated fibroblasts. Exposure of ESCC cells to supernatant from irradiated fibroblasts resulted in decreased E-cadherin, increased vimentin in vitro and β-catenin was demonstrated to localize to the nucleus in tumor cells with irradiated fibroblasts in vivo models. The expression of HDGF and β-catenin were increased in both fibroblasts and ESCC cells of irradiated group in vitro and in vivo models. Interestingly, the tumor cells adjoining the stromal fibroblasts displayed strong nuclear HDGF immunoreactivity, which suggested the occurrence of a paracrine effect of fibroblasts on HDGF expression. These data suggested that irradiated fibroblasts promoted invasion, growth, EMT and HDGF expression of ESCC. - Highlights: • Irradiated CAFs accelerated invasiveness and scattering of ESCC cell lines. • Irradiated CAFs promoted EMT of ESCC cells. • Irradiated fibroblasts induced nuclear β-catenin relocalization in ESCC cells. • Irradiated fibroblasts increased HDGF expression in vitro and in vivo.

  18. Feasibility of skin surface elastography by tracking skin surface topography

    Science.gov (United States)

    Coutts, Louise V.; Miller, Naomi R.; Harland, Christopher C.; Bamber, Jeffrey C.

    2013-12-01

    Recent advances have led to a multitude of image modalities being used for visualization of tissue stiffness. High-resolution images of tissue stiffness are desirable, as they have the potential to provide useful diagnostic information. A noncontact optical imaging method has the attractions of low cost, simplicity, and utility when skin contact is undesirable. However, previous optical techniques have required the application of paint or ink to the surface of the skin and so have required contact. Therefore, the present study assessed the feasibility of tracking skin surface topography to produce elastograms. The study showed, by analyzing a variety of silicone skin surface replicas from various body sites of subjects of different ages, that skin surface elastography by tracking surface topography would be feasible. The study further showed that the quality of the strain images can be optimized by measuring skin line pattern frequency. Skin samples with high skin line frequency will achieve best spatial resolution, in the order of 1 mm, comparable to contact techniques reported previously. A mechanically inhomogeneous silicone replica was then imaged, illustrating the technique's ability to detect strain contrast. Finally, the feasibility of implementing the technique in vivo was illustrated using a single pigmented skin lesion.

  19. Sjögren-Larsson syndrome. Impaired fatty alcohol oxidation in cultured fibroblasts due to deficient fatty alcohol:nicotinamide adenine dinucleotide oxidoreductase activity.

    OpenAIRE

    Rizzo, W B; Dammann, A L; Craft, D A

    1988-01-01

    Lipid metabolism was studied in cultured skin fibroblasts from patients with the inherited disorder, Sjögren-Larsson syndrome (SLS). Intact SLS fibroblasts incubated in the presence of [1-14C]palmitate accumulated more radioactive hexadecanol than did normal cells, whereas incorporation of radioactivity into other cellular lipids was unaltered. The hexadecanol content of SLS fibroblasts was abnormally elevated. Hexadecanol accumulation was not due to increased fatty alcohol synthesis nor its ...

  20. Skin cancer

    International Nuclear Information System (INIS)

    This chapter reviews the development of skin cancer associated with radiation, focusing on the knowledge of A-bomb radiation-induced skin cancer. Since the discovery of X radiation in 1895, acute and chronic radiation dermatitis has been the first matter of concern. Then, in 1902, skin cancer found among radiological personnel has posed a social problem. In earlier study determining the relationship between skin cancer and A-bomb radiation, there is no increase in the incidence of either skin cancer or precancerous condition during the first 20 years after A-bombing. More recent studies have showed that there is a significant correlation between the incidence of skin cancer and distance from the hypocenter; and the incidence of skin cancer is found to be remarkably increased since 1975 in the group exposed at ≤2,000 m. Excess relative risk is 2.2 at one Gy dose. The incidence of skin cancer is also found to be extremely increased with aging. Relative risk is high in younger A-bomb survivors at the time of exposure. Histologically, basal cell carcinoma is more senstitive to ionizing radiation than squamous cell carcinoma. (N.K.)

  1. Tattoo ink nanoparticles in skin tissue and fibroblasts

    OpenAIRE

    Grant, Colin A; Peter C. Twigg; Richard Baker; Tobin, Desmond J.

    2015-01-01

    Tattooing has long been practised in various societies all around the world and is becoming increasingly common and widespread in the West. Tattoo ink suspensions unquestionably contain pigments composed of nanoparticles, i.e., particles of sub-100 nm dimensions. It is widely acknowledged that nanoparticles have higher levels of chemical activity than their larger particle equivalents. However, assessment of the toxicity of tattoo inks has been the subject of little research and ink manufactu...

  2. Altered Gene Expression in Schizophrenia: Findings from Transcriptional Signatures in Fibroblasts and Blood

    OpenAIRE

    Cattane, Nadia; Minelli, Alessandra; Milanesi, Elena; Maj, Carlo; Bignotti, Stefano; Bortolomasi, Marco; Chiavetto, Luisella Bocchio; Gennarelli, Massimo

    2015-01-01

    Background Whole-genome expression studies in the peripheral tissues of patients affected by schizophrenia (SCZ) can provide new insight into the molecular basis of the disorder and innovative biomarkers that may be of great utility in clinical practice. Recent evidence suggests that skin fibroblasts could represent a non-neural peripheral model useful for investigating molecular alterations in psychiatric disorders. Methods A microarray expression study was conducted comparing skin fibroblas...

  3. Antioxidant and potential anti-inflammatory activity of extracts and formulations of white tea, rose, and witch hazel on primary human dermal fibroblast cells

    OpenAIRE

    Hili Pauline; Thring Tamsyn SA; Naughton Declan P

    2011-01-01

    Abstract Background Numerous reports have identified therapeutic roles for plants and their extracts and constituents. The aim of this study was to assess the efficacies of three plant extracts for their potential antioxidant and anti-inflammatory activity in primary human skin fibroblasts. Methods Aqueous extracts and formulations of white tea, witch hazel and rose were subjected to assays to measure anti-collagenase, anti-elastase, trolox equivalent and catalase activities. Skin fibroblast ...

  4. The time-pattern of rises and falls in proliferation fades with senescence of mortal lines and is perpetuated in immortal rat hepatoma Fao cell line.

    Science.gov (United States)

    Maigné, J; Deschatrette, J; Sarrazin, S; Hecquet, B; Guerroui, S; Wolfrom, C

    1998-02-01

    Immortal cells perpetuate the rises and falls of proliferation that are progressively damped in mortal long-term cultured cells. For immortal rat hepatoma Fao cells, similar waves of proliferation occurred about every 3-4 wk. Under the same conditions, embryonic human fibroblasts and transformed but not immortalized embryonic fibroblasts display similarly recurring proliferation waves that progressively decrease in amplitude until senescence of the lines. In addition, strains of diploid normal human skin fibroblasts cultured under different culture conditions display a similar time-pattern of proliferation. Although the amplitude and baseline of these fluctuations are characteristic for each cell line, a common point was marked slow down in proliferation after every sequence of about 25 population doublings for all cells. Renewed proliferation waves of Fao cells allow about 22-23 additional population doublings each. Normal embryonic fibroblast culture and its transformed counterpart accumulate about 30 and 60 population doublings, respectively, before senescence. Normal fibroblast strains accumulate about 25 population doublings over their entire life spans. This halt in proliferation after every stretch of about 25 population doublings may correspond to a structural or functional stop following attrition of telomeric DNA. This putative stop may be bypassed once in transformed embryonic cells and repetitively in immortal cells. In support of this hypothesis, we observed rapid telomere shortening, in two steps, during divisions of mortal embryonic cells, and maintenance of long telomeres in immortal Fao cells, which may indicate episodic repair of telomeres. Alternatively, such maintenance of long telomeres may reflect survival and successive clonal growth of rare cells with long telomeres. We suggest that the balance between telomere attrition and repair processes regulates the waves of proliferation. PMID:9542655

  5. Menkes disease: a biochemical abnormality in cultured human fibroblasts.

    Science.gov (United States)

    Goka, T J; Stevenson, R E; Hefferan, P M; Howell, R R

    1976-01-01

    Cultured skin fibroblasts from patients with Menkes disease, an X-linked disorder involving a defect in copper metabolism, were analyzed for copper concentration by means of atomic absorption spectrophotometry. These cultures consistently exhibited elevated copper concentrations (mean = 335.5 ng of copper per mg of protein) when compared to control fibroblast cultures (mean = 59.2 ng of copper per mg of protein). External factors that could influence the copper content of cultures were found not to affect the differences in copper concentration between control and Menkes cells. Furthermore, Menkes cells could be differentiated from cultured fibroblasts of controls, of presumed heterozygotes, and of Wilson's disease patients by copper concentration. These observations led to the conclusion that the increased copper content of cultured Menkes cells was characteristic of Menkes disease, resulting from the expression of the genetic abnormality. This provides a genetic marker, a defect in metal metabolism demonstrated in human fibroblasts, that should prove valuable in both the diagnosis of Menkes disease and in the study of the fundamental defect of this genetic disorder. PMID:1061160

  6. Transfer of fibroblast sheets cultured on thermoresponsive dishes with membranes.

    Science.gov (United States)

    Kawecki, Marek; Kraut, Małgorzata; Klama-Baryła, Agnieszka; Łabuś, Wojciech; Kitala, Diana; Nowak, Mariusz; Glik, Justyna; Sieroń, Aleksander L; Utrata-Wesołek, Alicja; Trzebicka, Barbara; Dworak, Andrzej; Szweda, Dawid

    2016-06-01

    In cell or tissue engineering, it is essential to develop a support for cell-to-cell adhesion, which leads to the generation of cell sheets connected by extracellular matrix. Such supports must be hydrophobic and should result in a detachable cell sheet. A thermoresponsive support that enables the cultured cell sheet to detach using only a change in temperature could be an interesting alternative in regenerative medicine. The aim of this study was to evaluate plates covered with thermoresponsive polymers as supports for the formation of fibroblast sheets and to develop a damage-free procedure for cell sheet transfer with the use of membranes as transfer tools. Human skin fibroblasts were seeded on supports coated with a thermoresponsive polymer: commercial UpCell™ dishes (NUNC™) coated with thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) and dishes coated with thermoresponsive poly(tri(ethylene glycol) monoethyl ether methacrylate) (P(TEGMA-EE)). Confluent fibroblast sheets were effectively cultured and harvested from both commercial PNIPAM-coated dishes and laboratory P(TEGMA-EE)-coated dishes. To transfer a detached cell sheet, two membranes, Immobilon-P(®) and SUPRATHEL(®), were examined. The use of SUPRATHEL for relocating the cell sheets opens a new possibility for the clinical treatment of wounds. This study established the background for implementing thermoresponsive supports for transplanting in vitro cultured fibroblasts. PMID:27153827

  7. Skin Cancer in Skin of Color

    OpenAIRE

    Bradford, Porcia T.

    2009-01-01

    Skin cancers in skin of color often present atypically or with advanced stage in comparison to Caucasian patients. Health care providers must maintain a high index of suspicion when examining skin lesions in skin of color.

  8. Role of Age-Associated Alterations of the Dermal Extracellular Matrix Microenvironment in Human Skin Aging

    OpenAIRE

    Quan, TaiHao; Gary J. Fisher

    2015-01-01

    Human skin is largely composed of a collagen-rich connective tissue, which provides structural and functional support. The collagen-rich connective tissue is produced, organized, and maintained by dermal fibroblasts. During aging, dermal collagen fibrils undergo progressive loss and fragmentation, leading to thin and structurally weakened skin. Age-related alterations of collagen fibrils impairs skin structure and function and creates a tissue microenvironment that promotes age-related skin d...

  9. Role of DNA lesions and DNA repair in mutagenesis by carcinogens in diploid human fibroblasts

    International Nuclear Information System (INIS)

    The authors investigated the cytotoxicity, mutagenicity, and transforming activity of carcinogens and radiation in diploid human fibroblasts, using cells which differ in their DNA repair capacity. The results indicate that cell killing and induction of mutations are correlated with the number of specific lesions remaining unrepaired in the cells at a particular time posttreatment. DNA excision repair acts to eliminate potentially cytotoxic and mutagenic (and transforming) damage from DNA before these can be converted into permanent cellular effects. Normal human fibroblasts were derived from skin biopsies or circumcision material. Skin fibroblasts from xeroderma pigmentosum (XP) patients provided cells deficient in nucleotide excision repair of pyrimidine dimers or DNA adducts formed by bulky ring structures. Cytotoxicity was determined from loss of ability to form a colony. The genetic marker used was resistance to 6-thioguanine (TG). Transformation was measured by determining the frequency of anchorage-independent cells

  10. Fibroblasts and myofibroblasts in wound healing

    Directory of Open Access Journals (Sweden)

    Darby IA

    2014-11-01

    Full Text Available Ian A Darby,1 Betty Laverdet,2 Frédéric Bonté3, Alexis Desmoulière2 1School of Medical Sciences, RMIT University, Melbourne, VIC, Australia; 2Department of Physiology and EA 6309, FR 3503, Faculties of Medicine and Pharmacy, University of Limoges, Limoges, France; 3LVMH Recherche, Saint Jean de Braye, France Abstract: (Myofibroblasts are key players for maintaining skin homeostasis and for orchestrating physiological tissue repair. (Myofibroblasts are embedded in a sophisticated extracellular matrix (ECM that they secrete, and a complex and interactive dialogue exists between (myofibroblasts and their microenvironment. In addition to the secretion of the ECM, (myofibroblasts, by secreting matrix metalloproteinases and tissue inhibitors of metalloproteinases, are able to remodel this ECM. (Myofibroblasts and their microenvironment form an evolving network during tissue repair, with reciprocal actions leading to cell differentiation, proliferation, quiescence, or apoptosis, and actions on growth factor bioavailability by binding, sequestration, and activation. In addition, the (myofibroblast phenotype is regulated by mechanical stresses to which they are subjected and thus by mechanical signaling. In pathological situations (excessive scarring or fibrosis, or during aging, this dialogue between the (myofibroblasts and their microenvironment may be altered or disrupted, leading to repair defects or to injuries with damaged and/or cosmetic skin alterations such as wrinkle development. The intimate dialogue between the (myofibroblasts and their microenvironment therefore represents a fascinating domain that must be better understood in order not only to characterize new therapeutic targets and drugs able to prevent or treat pathological developments but also to interfere with skin alterations observed during normal aging or premature aging induced by a deleterious environment. Keywords: myofibroblast, fibroblast, α-smooth muscle actin, mechanical signaling, fibrosis, scarring 

  11. Skin Pigment

    Science.gov (United States)

    ... Disorders Overview of Skin Pigment Albinism Vitiligo Hyperpigmentation Melasma Melanin is the brown pigment that produces the ... such spots include freckles, age spots ( lentigines ), and melasma . Melanin is produced by specialized cells (melanocytes) that ...

  12. Evaluation of X-Inactivation Status and Cytogenetic Stability of Human Dermal Fibroblasts after Long-Term Culture

    OpenAIRE

    Zhi-Gang Xue; Zhan-Ping Shi; Juan Dong; Ting-Ting Liao; Yan-Peng Wang; Xue-Ping Sun; Zheng-Jie Yan; Xiao-Qiao Qian; Yu-Gui Cui; Juan Chen; Jia-Yin Liu; Guoping Fan

    2010-01-01

    Human primary fibroblasts are a popular type of somatic cells for the production of induced pluripotent stem (iPS) cells. Here we characterized biological properties of primary fibroblasts in terms of cell-growth rate, cytogenetic stability, and the number of inactive X chromosomes during long-term passaging. We produced eight lines of female human dermal fibroblasts (HDFs) and found normal karyotype and expected pattern of X chromosome inactivation (XCI) at low passages (Passage P1-5). Howev...

  13. Nucleolin enhances the proliferation and migration of heat-denatured human dermal fibroblasts.

    Science.gov (United States)

    Jiang, Bimei; Li, Yuanbin; Liang, Pengfei; Liu, Yanjuan; Huang, Xu; Tong, Zhongyi; Zhang, Pihong; Huang, Xiaoyuan; Liu, Ying; Liu, Zhenguo

    2015-11-12

    Denatured dermis, a part of dermis in burned skin, has the ability to restore its normal morphology and functions after their surrounding microenvironment is improved. However, the cellular and molecular mechanisms by which the denatured dermis could improve wound healing are still unclear. This study aimed to investigate the role of nucleolin during the recovery of heat-denatured human dermal fibroblasts. Nucleolin mRNA and protein expression were significantly increased time-dependently during the recovery of heat-denatured human dermal fibroblasts (52?C, 30 seconds). Heat-denaturation promoted a time-dependent cell proliferation, migration, chemotaxis, and scratched wound healing during the recovery of human dermal fibroblasts. These effects were prevented by knockdown of nucleolin expression with small interference RNA (siRNA), whereas overexpression of nucleolin enhanced cell proliferation, migration, and chemotaxis of human dermal fibroblasts with heat-denaturation. In addition, the expression of transforming growth factor-beta 1(TGF-?1) was significantly increased during the recovery of heat-denatured dermis and human dermal fibroblasts. TGF-?1 expression was up-regulated by nucleolin in human dermal fibroblasts. The results suggest that nucleolin expression is up-regulated, and play an important role in promoting cell proliferation, migration, and chemotaxis of human dermal fibroblasts during the recovery of heat-denatured dermis with a mechanism probably related to TGF-?1. PMID:26148015

  14. Sarcoptes scabiei Mites Modulate Gene Expression in Human Skin Equivalents

    OpenAIRE

    MORGAN, MARJORIE S.; Arlian, Larry G.; Markey, Michael P.

    2013-01-01

    The ectoparasitic mite, Sarcoptes scabiei that burrows in the epidermis of mammalian skin has a long co-evolution with its hosts. Phenotypic studies show that the mites have the ability to modulate cytokine secretion and expression of cell adhesion molecules in cells of the skin and other cells of the innate and adaptive immune systems that may assist the mites to survive in the skin. The purpose of this study was to identify genes in keratinocytes and fibroblasts in human skin equivalents (H...

  15. Suppressive effects of induced pluripotent stem cell-conditioned medium on in vitro hypertrophic scarring fibroblast activation.

    Science.gov (United States)

    Ren, Ye; Deng, Chen-Liang; Wan, Wei-Dong; Zheng, Jiang-Hong; Mao, Guang-Yu; Yang, Song-Lin

    2015-04-01

    Hypertrophic scarring (HS) is a type of fibrosis that occurs in the skin, and is characterized by fibroblast activation and excessive collagen production. However, at present, therapeutic strategies for this condition are ineffective. Previous studies have identified that the mutual regulation of chronic inflammation, mechanical force and fibroblast activation leads to the formation of HS. Induced pluripotent stem cells (iPSCs) are novel bioengineered embryonic?like stem cells, initially created from mouse adult fibroblasts. The current study demonstrated that iPSC?conditioned medium (iPSC?CM) may significantly suppress hypertrophic scar fibroblast activation. It was observed that in the presence of iPSC?CM, the level of collagen I was markedly reduced and ??smooth muscle actin, a marker for myofibroblasts (activated fibroblasts that mediate mechanical force?induced HS formation), exhibited a significantly lower level of expression in human dermal fibroblasts (HDFs) activated with transforming growth factor??1. Additionally, iPSC?CM attenuated the local inflammatory cell response by blocking the adhesion of human acute monocytic leukemia cell monocytes and fibroblasts in vitro. In addition, the contractile ability of HDFs may be reduced by iPSC?CM. These observations suggest that iPSC?CM may protect against processes leading to hypertrophic scarring by attenuating fibroblast activation, blocking inflammatory cell recruitment and adhesion and reducing the contractile ability of fibroblasts. PMID:25524174

  16. Lysine hydroxylation of collagen in a fibroblast cell culture system

    Science.gov (United States)

    Uzawa, Katsuhiro; Yeowell, Heather N.; Yamamoto, Kazushi; Mochida, Yoshiyuki; Tanzawa, Hideki; Yamauchi, Mitsuo

    2003-01-01

    The lysine (Lys) hydroxylation pattern of type I collagen produced by human fibroblasts in culture was analyzed and compared. Fibroblasts were cultured from normal human skin (NSF), keloid (KDF), fetal skin (FDF), and skin tissues of Ehlers-Danlos syndrome type VIA and VIB patients (EDS-VIA and -VIB). The type I collagen alpha chains with or without non-helical telopeptides were purified from the insoluble matrix and analyzed. In comparison with NSFs, KDF and FDF showed significantly higher Lys hydroxylation, particularly in the telopeptide domains of both alpha chains. Both EDS-VIA and -VIB showed markedly lower Lys hydroxylation in the helical domains of both alpha chains whereas that in the telopeptides was comparable with those of NSFs. A similar profile was observed in the tissue sample of the EDS-VIB patient. These results demonstrate that the Lys hydroxylation pattern is domain-specific within the collagen molecule and that this method is useful to characterize the cell phenotypes in normal/pathological connective tissues.

  17. Skin Cancer Screening

    Science.gov (United States)

    ... Genetics of Skin Cancer Skin Cancer Screening Research Skin Cancer Screening (PDQ®)–Patient Version What is screening? ... These are called diagnostic tests . General Information About Skin Cancer Key Points Skin cancer is a disease ...

  18. Skin Cancer Prevention

    Science.gov (United States)

    ... Genetics of Skin Cancer Skin Cancer Screening Research Skin Cancer Prevention (PDQ®)–Patient Version What is prevention? ... to keep cancer from starting. General Information About Skin Cancer Key Points Skin cancer is a disease ...

  19. Dry Skin (Xerosis)

    Science.gov (United States)

    ... Kids’ zone Video library Find a dermatologist Dry skin Overview Dry, ashy skin: People who had atopic ... often have very dry skin as adults. Dry skin: Overview Also called xerosis Dry skin is common. ...

  20. Skin color - patchy

    Science.gov (United States)

    Patchy skin color is areas where the skin color is irregular. Mottling or mottled skin refers to blood vessel changes in ... in the skin cells that gives skin its color Growth of bacteria or other organisms on the ...

  1. Glutathione metabolism in the HaCaT cell line as a model for the detoxification of the model sensitisers 2,4-dinitrohalobenzenes in human skin.

    Science.gov (United States)

    Jacquoilleot, Sandrine; Sheffield, David; Olayanju, Adedamola; Sison-Young, Rowena; Kitteringham, Neil R; Naisbitt, Dean J; Aleksic, Maja

    2015-08-19

    Glutathione (GSH) is the most prominent antioxidant in cells and the co-factor of an important set of enzymes involved in the skin metabolic clearance system, glutathione S-transferases (GST). Here, we describe an LC-MS (liquid chromatography-mass spectroscopy) method to measure GSH and its disulfide form (GSSG) in HaCaT cells and a 3D Reconstructed Human Epidermis (RHE) model. In our assay, the basal level of GSH in both systems was in the low nmol/mg soluble protein range, while the level of GSSG was systematically below our limit of quantification (0.1 μM). We found that 2,4-dinitrohalobenzenes deplete the GSH present in HaCaT cells within the first hour of exposure, in a dose dependent manner. The level of GSH in HaCaT cells treated with a single non-toxic dose of 10 μM of dinitrohalobenzene was also shown to increase after two hours. While cells treated with 1-chloro-2,4-dinitrobenzene (DNCB) and 1-fluoro-2,4-dinitrobenzene (DNFB) repleted GSH to levels similar to untreated control cells within 24h, 1-bromo-2,4-dinitrobenzene (DNBB) seemed to prevent such a repletion and appeared to be the most toxic compound in all assays. A mathematical modelling of experimental results was performed to further rationalise the differences observed between test chemicals. For this purpose the biological phenomena observed were simplified into two sequential events: the initial depletion of the GSH stock after chemical treatment followed by the repletion of the GSH once the chemical was cleared. Activation of the nuclear factor E2-related factor 2 (Nrf2) pathway was observed with all compounds within two hours, and at concentrations less than 10 μM. These data show that GSH depletion and repletion occur rapidly in skin cells and emphasize the importance of conducting kinetic studies when performing in vitro experiments exploring skin sensitization. PMID:26022718

  2. Demonstration of a specific mitochondrial isovaleryl-CoA dehydrogenase deficiency in fibroblasts from patients with isovaleric acidemia.

    OpenAIRE

    Rhead, W J; Tanaka, K

    1980-01-01

    To study the enzymatic basis of isovaleric acidemia, we have developed assay methods for isovaleryl-CoA and butyryl-CoA dehydrogenases that measure the amount of tritium released from the respective [2,3-3H]acyl CoAs. Because assay of these enzymes in human fibroblast homogenates was subject to interference by nonspecific reactions, we have isolated mitochondria from cultured skin fibroblasts by protease treatment, homogenization, and differential centrifugation. By using this assay method wi...

  3. Annexin A1 N-Terminal Derived Peptide Ac2-26 Stimulates Fibroblast Migration in High Glucose Conditions

    OpenAIRE

    Bizzarro, Valentina; Fontanella, Bianca; Carratù, Anna; Belvedere, Raffaella; Marfella, Raffaele; Parente, Luca; Petrella, Antonello

    2012-01-01

    Deficient wound healing in diabetic patients is very frequent, but the cellular and molecular causes are poorly defined. In this study, we have evaluated whether Annexin A1 derived peptide Ac2-26 stimulates fibroblast migration in high glucose conditions. Using normal human skin fibroblasts WS1 in low glucose (LG) or high glucose (HG) we observed the enrichment of Annexin A1 protein at cell movement structures like lamellipodial extrusions and interestingly, a significant decrease in levels o...

  4. Activation of vitamin D receptor (VDR)- and peroxisome proliferator-activated receptor (PPAR)-signaling pathways through 1,25(OH)2D3 in melanoma cell lines and other skin-derived cell lines

    OpenAIRE

    Sertznig, Pit; Seifert, Markus; Tilgen, Wolfgang; Reichrath, Jörg

    2009-01-01

    We have investigated expression of vitamin D receptor (VDR) and peroxisome proliferator-activated receptors (PPAR)α, δ, γ in primary cultured normal melanocytes (NHM), melanoma cell lines (MeWo, SK-Mel-5, SK-Mel-25, SK-Mel-28), a cutaneous squamous cell carcinoma cell line (SCL-1) and an immortalized sebocyte cell line (SZ95). LNCaP prostate cancer cells, MCF-7 breast cancer cells and embryonic kidney cells (HEK-293) were used as controls. VDR and PPAR mRNA were detected, quantitated and comp...

  5. Low dose radiation effects on the transcription of consensus radiation response genes in primary and immortalized human fibroblast cells

    International Nuclear Information System (INIS)

    Complete text of publication follows. OBJECTIVE: The linear non-threshold model suggests that tumors might be induced even by low radiation doses. Still, most of the conventional methods are unable to detect damages below 100 mGy. We have studied whether transcriptional responses of consensus radiation response genes can be detected after low dose radiation exposure in directly exposed or bystander primary human fibroblast cells. The short term proliferation capacity of primary fibroblast cells in culture limits their long term application. Therefore we tried to immortalize the cells by the introduction of the human telomerase gene using retroviral vectors. METHODS: Primary human fibroblast cell lines were established from skin biopsies of cancer patients and foreskin samples of young children. To create immortalized cell lines the human telomerase gene was cloned into a retroviral vector. Primary fibroblast cells were transduced and their proliferation capacity studied. To investigate radiation induced transcriptional alterations, cells were irradiated with 60Co γ-rays (0; 0.01; 0,04; 0,1; 2 and 8 Gy) and 2 hours later total cellular RNA was isolated both from directly exposed and bystander cells. Transcriptional alterations were followed in consensus radiation response genes (CDKN1, GADD45, GDF15, IER5, PLK3, TP53INP1) with quantitative real time PCR (Corbett/ SybrGreen). RESULTS: There is an elevated expression of CDKN1, GADD45, GDF15, PLK3, TP53INP1 in the exposed cells. We see only for the PLK3 a dose-dependent increase which manifested also at low doses. It seems this gene is the most sensitive to radiation at low doses. The hTERT-immortalized cells were morphologically identical to the primary cells. the radiation-induced transcriptional profile of immortalized cells were very similar to the primary ones. CONCLUSIONS: hTERT immortalized cells can be used to mimic alterations in primary cells. Low dose irradiation doesn't influence the expression of most of the studied genes. PLK3 might be an efficient marker to estimate individually low dose effects.

  6. Skin decontamination

    International Nuclear Information System (INIS)

    A general survey of skin decontamination is given. The success of every decontamination treatments depends mainly on the speed, but also on the care, with which the action is taken. The best way to remove the skin contaminants is thorough washing under lukewarm running water with mild soap and a soft brush. This washing is to be repeated several times for a period of several minutes. If results are not satisfactory, light duty detergents and wetting agents available commercially may also be used. Some solutions which have proved useful are mentioned. The decontamination solutions are best used in the order given. When one has no satisfactory decontamination effect, the next one is to be used. If necessary, these agents must be used several times in the stated order as long as this does not involve too much strain for the skin. All the decontamination measures mentioned refer, of course, to intact healthy skin. After decontamination has been completed, the skin should be treated with a protective cream

  7. STAT6-Dependent Collagen Synthesis in Human Fibroblasts Is Induced by Bovine Milk

    OpenAIRE

    Kippenberger, Stefan; Zller, Nadja; Kleemann, Johannes; Mller, Jutta; Kaufmann, Roland; Hofmann, Matthias; Bernd, August; Meissner, Markus; Valesky, Eva

    2015-01-01

    Since the domestication of the urus, 10.000 years ago, mankind utilizes bovine milk for different purposes. Besides usage as a nutrient also the external application of milk on skin has a long tradition going back to at least the ancient Aegypt with Cleopatra VII as a great exponent. In order to test whether milk has impact on skin physiology, cultures of human skin fibroblasts were exposed to commercial bovine milk. Our data show significant induction of proliferation by milk (max. 2,3-fold,...

  8. Aging and senescence of skin cells in culture

    DEFF Research Database (Denmark)

    Rattan, Suresh

    Studying age-related changes in the physiology, biochemistry, and molecular biology of isolated skin cell populations in culture has greatly expanded the understanding of the fundamental aspects of skin aging. The three main cell types that have been studied extensively with respect to cellular...... aging in vitro are dermal fibroblasts, epidermal keratinocytes, and melanocytes. Serial subcultivation of normal diploid skin cells can be performed only a limited number of times, and the emerging senescent phenotype can be categorized into structural, physiological, biochemical, and molecular...... phenotypes, which can be used as biomarkers of cellular aging in vitro. The rate and phenotype of aging are different in different cell types. There are both common features and specific features of aging of skin fibroblasts, keratinocytes, melanocytes, and other cell types. A progressive accumulation of...

  9. A quantitative in vitro study of fibroblast and endothelial cell migration in response to serum and wound fluid

    International Nuclear Information System (INIS)

    Chemoattractant activity for irradiated and nonirradiated rabbit skin fibroblast and bovine aortic arch endothelial cells was assayed in rabbit wound fluid and sera using a modification of the agarose well method originally described for polymorphonuclear leukocytes. Both serum and wound fluid contained chemoattractants for fibroblasts and endothelial cells. Fibroblast migration was decreased by 70 to 80% when the serum or wound fluid was heated to 56 degrees C for 30 min while endothelial cell migration was reduced by 50 to 60%. Platelet-poor plasma-derived serum had no directive effect on the migration of either cell type

  10. Simultaneous Reprogramming and Gene Correction of Patient Fibroblasts

    Directory of Open Access Journals (Sweden)

    Sara E. Howden

    2015-12-01

    Full Text Available The derivation of genetically modified induced pluripotent stem (iPS cells typically involves multiple steps, requiring lengthy cell culture periods, drug selection, and several clonal events. We report the generation of gene-targeted iPS cell lines following a single electroporation of patient-specific fibroblasts using episomal-based reprogramming vectors and the Cas9/CRISPR system. Simultaneous reprogramming and gene targeting was tested and achieved in two independent fibroblast lines with targeting efficiencies of up to 8% of the total iPS cell population. We have successfully targeted the DNMT3B and OCT4 genes with a fluorescent reporter and corrected the disease-causing mutation in both patient fibroblast lines: one derived from an adult with retinitis pigmentosa, the other from an infant with severe combined immunodeficiency. This procedure allows the generation of gene-targeted iPS cell lines with only a single clonal event in as little as 2 weeks and without the need for drug selection, thereby facilitating “seamless” single base-pair changes.

  11. Study of mast cell count in skin tags

    Directory of Open Access Journals (Sweden)

    Zaher Hesham

    2007-01-01

    Full Text Available Background: Skin tags or acrochordons are common tumors of middle-aged and elderly subjects. They consist of loose fibrous tissue and occur mainly on the neck and major flexures as small, soft, pedunculated protrusions. Objectives: The aim was to compare the mast cells count in skin tags to adjacent normal skin in diabetic and nondiabetic participants in an attempt to elucidate the possible role of mast cells in the pathogenesis of skin tags. Participants and Methods: Thirty participants with skin tags were divided into group I (15 nondiabetic participants and group II (15 diabetic participants. Three biopsies were obtained from each participant: a large skin tag, a small skin tag and adjacent normal skin. Mast cell count from all the obtained sections was carried out, and the mast cell density was expressed as the average mast cell count/high power field (HPF. Results: A statistically significant increase in mast cells count in skin tags in comparison to normal skin was detected in group I and group II. There was no statistically significant difference between mast cell counts in skin tags of both the groups. Conclusion: Both the mast cell mediators and hyperinsulinemia are capable of inducing fibroblast proliferation and epidermal hyperplasia that are the main pathologic abnormalities seen in all types of skin tags. However, the presence of mast cells in all examined skin tags regardless of diabetes and obesity may point to the possible crucial role of mast cells in the etiogenesis of skin tags through its interaction with fibroblasts and keratinocytes.

  12. Effect of radiation on reconstitution of skin equivalent (dermal alterations)

    International Nuclear Information System (INIS)

    Dermal equivalents have been treated by single doses of gamma irradiation of 10, 20, 30 and 50 Gray. Numerations at different times show a dose and time dependant diminution of cellular population. This diminution is histologically observed in dermal part of reconstituted skin, in association with cellular and functional alterations of fibroblast cells. Modifications of epidermal epithelia are also noted in some reconstituted skin. This model would be useful to apprehend the effect of a dermal irradiation lesion on the later epidermization. (author)

  13. Response of Human Skin Equivalents to Sarcoptes scabiei

    OpenAIRE

    MORGAN, MARJORIE S.; Arlian, Larry G.

    2010-01-01

    Studies have shown that molecules in an extract made from bodies of the ectoparasitic mite, Sarcoptes scabiei De Geer, modulate cytokine secretion from cultured human keratinocytes and fibroblasts. In vivo, in the parasitized skin, these cells interact with each other by contact and cytokine mediators and with the matrix in which they reside. Therefore, these cell types may function differently together than they do separately. In this study, we used a human skin equivalent (HSE) model to inv...

  14. A Luciferase-Expressing Leishmania braziliensis Line That Leads to Sustained Skin Lesions in BALB/c Mice and Allows Monitoring of Miltefosine Treatment Outcome

    Science.gov (United States)

    Coelho, Adriano C.; Oliveira, Jordana C.; Espada, Caroline R.; Reimão, Juliana Q.; Trinconi, Cristiana T.; Uliana, Silvia R. B.

    2016-01-01

    Background Leishmania braziliensis is the most prevalent species isolated from patients displaying cutaneous and muco-cutaneous leishmaniasis in South America. However, there are difficulties for studying L. braziliensis pathogenesis or response to chemotherapy in vivo due to the natural resistance of most mouse strains to infection with these parasites. The aim of this work was to develop an experimental set up that could be used to assess drug efficacy against L. braziliensis. The model was tested using miltefosine. Methodology/Principal Findings A L. braziliensis line, originally isolated from a cutaneous leishmaniasis patient, was passaged repeatedly in laboratory rodents and further genetically manipulated to express luciferase. Once collected from a culture of parasites freshly transformed from amastigotes, 106 wild type or luciferase-expressing stationary phase promastigotes were inoculated subcutaneously in young BALB/c mice or golden hamsters. In both groups, sustained cutaneous lesions developed at the site of inoculation, no spontaneous self- healing being observed 4 months post-inoculation, if left untreated. Compared to the wild type line features, no difference was noted for the luciferase-transgenic line. Infected animals were treated with 5 or 15 mg/kg/day miltefosine orally for 15 days. At the end of treatment, lesions had regressed and parasites were not detected. However, relapses were observed in animals treated with both doses of miltefosine. Conclusions/Significance Here we described experimental settings for a late-healing model of cutaneous leishmaniasis upon inoculation of a luciferase-expressing L. braziliensis line that can be applied to drug development projects. These settings allowed the monitoring of the transient efficacy of a short-term miltefosine administration. PMID:27144739

  15. Wnt signaling in skin homeostasis and pathology.

    Science.gov (United States)

    Augustin, Iris

    2015-04-01

    The mammalian skin mediates the primary interphase between the body and the external environment and provides the first line of defense against pathogens, mechanical trauma, sunlight injuries, and chemical stress. Proper physical, biochemical, and immunological composition of the skin is necessary to maintain its barrier function. Therefore, the skin reflects a complex dynamic organ with high cellular turnover during normal tissue replacement and wound repair. Stem cell reservoirs ensure constant skin renewal. Wnt signaling controls stem cell maintenance and fate decisions in various tissues and also reflects a key pathway in controlling skin development and homeostasis. Disruption of Wnt signaling in the skin causes disorders such as alopecia, chronic inflammatory skin diseases or cancer. This review summarizes the role of Wnt signaling during skin development, homeostasis, and disease. PMID:25819237

  16. Abnormal iron metabolism in fibroblasts from a patient with the neurodegenerative disease hereditary ferritinopathy

    Directory of Open Access Journals (Sweden)

    Ghetti Bernardino

    2010-11-01

    Full Text Available Abstract Background Nucleotide duplications in exon 4 of the ferritin light polypeptide (FTL gene cause the autosomal dominant neurodegenerative disease neuroferritinopathy or hereditary ferritinopathy (HF. Pathologic examination of patients with HF has shown abnormal ferritin and iron accumulation in neurons and glia in the central nervous system (CNS as well as in cells of other organ systems, including skin fibroblasts. To gain some understanding on the molecular basis of HF, we characterized iron metabolism in primary cultures of human skin fibroblasts from an individual with the FTL c.497_498dupTC mutation. Results Compared to normal controls, HF fibroblasts showed abnormal iron metabolism consisting of increased levels of ferritin polypeptides, divalent metal transporter 1, basal iron content and reactive oxygen species, and decreased levels of transferrin receptor-1 and IRE-IRP binding activity. Conclusions Our data indicates that HF fibroblasts replicate the abnormal iron metabolism observed in the CNS of patients with HF. We propose that HF fibroblasts are a unique cellular model in which to study the role of abnormal iron metabolism in the pathogenesis of HF without artifacts derived from over-expression or lack of endogenous translational regulatory elements.

  17. Increased biosynthesis and processing of fibronectin in fibroblasts from diabetic mice

    International Nuclear Information System (INIS)

    Diabetic connective tissues exhibit a deranged regulation of extracellular matrix biosynthesis. Fibronectin is shown to be increased in human dermal connective tissue by immunofluorescence, mainly at the dermoepidermal and capillary basement membranes. The rate of fibronectin biosynthesis, excretion, and incorporation in a pericellular polymeric form was investigated using genetically diabetic KK mouse skin and fibroblasts as compared to swiss and C57BL mouse skin and fibroblasts. The rate of incorporation of [35S]methionine into proteins recovered in the culture medium or in deoxycholate and NaDodSO4 or urea extracts was investigated. The rate of incorporation in the medium and deoxycholate extracts was comparable. However, the relative rate of incorporation of the tracer in the NaDodSO4-extractable, pericellular polymeric form was increased in the diabetic KK fibroblasts both for total proteins and for fibronectin. In pulse-chase experiments, the deoxycholate-soluble and NaDodSO4-soluble fractions exhibited a precursor-product relationship. The rate of passage of fibronectin from the deoxycholate-soluble (cellular compartment) form to the NaDodSO4-soluble (pericellular polymeric) form was strongly accelerated in the diabetic fibroblast cultures. These results confirm the increased rate of synthesis of fibronectin in diabetic fibroblasts as well as its processing from the cellular compartment to the polymeric pericellular form

  18. Skin Cancer

    Science.gov (United States)

    ... the sun too often for too long can lead to skin cancer, even if you don't burn. A tan ... re wearing sunscreen. Sunscreen cannot give you 100% protection against the sun's harmful UV radiation. 3. Wear a wide-brimmed hat, protective clothing ...

  19. RAGE and TGF-β1 Cross-Talk Regulate Extracellular Matrix Turnover and Cytokine Synthesis in AGEs Exposed Fibroblast Cells

    Science.gov (United States)

    Serban, Andreea Iren; Stanca, Loredana; Geicu, Ovidiu Ionut; Munteanu, Maria Cristina; Dinischiotu, Anca

    2016-01-01

    AGEs accumulation in the skin affects extracellular matrix (ECM) turnover and triggers diabetes associated skin conditions and accelerated skin aging. The receptor of AGEs (RAGE) has an essential contribution to cellular dysfunction driven by chronic inflammatory responses while TGF-β1 is critical in both dermal homeostasis and inflammation. We investigated the contribution of RAGE and TGF-β1 to the modulation of inflammatory response and ECM turnover in AGEs milieu, using a normal fibroblast cell line. RAGE, TGF-β1, collagen I and III gene and protein expression were upregulated after exposure to AGEs-BSA, and MMP-2 was activated. AGEs-RAGE was pivotal in NF-κB dependent collagen I expression and joined with TGF-β1 to stimulate collagen III expression, probably via ERK1/2 signaling. AGEs-RAGE axis induced upregulation of TGF-β1, TNF-α and IL-8 cytokines. TNF-α and IL-8 were subjected to TGF-β1 negative regulation. RAGE’s proinflammatory signaling also antagonized AGEs-TGF-β1 induced fibroblast contraction, suggesting the existence of an inhibitory cross-talk mechanism between TGF-β1 and RAGE signaling. RAGE and TGF-β1 stimulated anti-inflammatory cytokines IL-2 and IL-4 expression. GM-CSF and IL-6 expression appeared to be dependent only on TGF-β1 signaling. Our data also indicated that IFN-γ upregulated in AGEs-BSA milieu in a RAGE and TGF-β1 independent mechanism. Our findings raise the possibility that RAGE and TGF-β1 are both involved in fibrosis development in a complex cross-talk mechanism, while also acting on their own individual targets. This study contributes to the understanding of impaired wound healing associated with diabetes complications. PMID:27015414

  20. A simple technique for preparation of chicken-embryo-skin cell cultures.

    Science.gov (United States)

    Silim, A; El Azhary, M A; Roy, R S

    1982-01-01

    A simple, rapid technique was developed for preparing chicken-embryo-skin cell cultures utilizing trypsinization of the skin of intact 12-day-old chicken embryos. When cell cultures were inoculated with fowl pox virus, those that consisted of at least 80% epithelial cells yielded a higher virus titer than fibroblast cell cultures. PMID:6284112

  1. Protective Effects of Triphala on Dermal Fibroblasts and Human Keratinocytes.

    Science.gov (United States)

    Varma, Sandeep R; Sivaprakasam, Thiyagarajan O; Mishra, Abheepsa; Kumar, L M Sharath; Prakash, N S; Prabhu, Sunil; Ramakrishnan, Shyam

    2016-01-01

    Human skin is body's vital organ constantly exposed to abiotic oxidative stress. This can have deleterious effects on skin such as darkening, skin damage, and aging. Plant-derived products having skin-protective effects are well-known traditionally. Triphala, a formulation of three fruit products, is one of the most important rasayana drugs used in Ayurveda. Several skin care products based on Triphala are available that claim its protective effects on facial skin. However, the skin protective effects of Triphala extract (TE) and its mechanistic action on skin cells have not been elucidated in vitro. Gallic acid, ellagic acid, and chebulinic acid were deduced by LC-MS as the major constituents of TE. The identified key compounds were docked with skin-related proteins to predict their binding affinity. The IC50 values for TE on human dermal fibroblasts (HDF) and human keratinocytes (HaCaT) were 204.90 ± 7.6 and 239.13 ± 4.3 μg/mL respectively. The antioxidant capacity of TE was 481.33 ± 1.5 mM Trolox equivalents in HaCaT cells. Triphala extract inhibited hydrogen peroxide (H2O2) induced RBC haemolysis (IC50 64.95 μg/mL), nitric oxide production by 48.62 ± 2.2%, and showed high reducing power activity. TE also rescued HDF from H2O2-induced damage; inhibited H2O2 induced cellular senescence and protected HDF from DNA damage. TE increased collagen-I, involucrin and filaggrin synthesis by 70.72 ± 2.3%, 67.61 ± 2.1% and 51.91 ± 3.5% in HDF or HaCaT cells respectively. TE also exhibited anti-tyrosinase and melanin inhibition properties in a dose-dependent manner. TE increased the mRNA expression of collagen-I, elastin, superoxide dismutase (SOD-2), aquaporin-3 (AQP-3), filaggrin, involucrin, transglutaminase in HDF or HaCaT cells, and decreased the mRNA levels of tyrosinase in B16F10 cells. Thus, Triphala exhibits protective benefits on skin cells in vitro and can be used as a potential ingredient in skin care formulations. PMID:26731545

  2. Protective Effects of Triphala on Dermal Fibroblasts and Human Keratinocytes

    Science.gov (United States)

    Varma, Sandeep R.; Sivaprakasam, Thiyagarajan O.; Mishra, Abheepsa; Kumar, L. M. Sharath; Prakash, N. S.; Prabhu, Sunil; Ramakrishnan, Shyam

    2016-01-01

    Human skin is bodys vital organ constantly exposed to abiotic oxidative stress. This can have deleterious effects on skin such as darkening, skin damage, and aging. Plant-derived products having skin-protective effects are well-known traditionally. Triphala, a formulation of three fruit products, is one of the most important rasayana drugs used in Ayurveda. Several skin care products based on Triphala are available that claim its protective effects on facial skin. However, the skin protective effects of Triphala extract (TE) and its mechanistic action on skin cells have not been elucidated in vitro. Gallic acid, ellagic acid, and chebulinic acid were deduced by LC-MS as the major constituents of TE. The identified key compounds were docked with skin-related proteins to predict their binding affinity. The IC50 values for TE on human dermal fibroblasts (HDF) and human keratinocytes (HaCaT) were 204.90 7.6 and 239.13 4.3 ?g/mL respectively. The antioxidant capacity of TE was 481.33 1.5 mM Trolox equivalents in HaCaT cells. Triphala extract inhibited hydrogen peroxide (H2O2) induced RBC haemolysis (IC50 64.95 ?g/mL), nitric oxide production by 48.62 2.2%, and showed high reducing power activity. TE also rescued HDF from H2O2-induced damage; inhibited H2O2 induced cellular senescence and protected HDF from DNA damage. TE increased collagen-I, involucrin and filaggrin synthesis by 70.72 2.3%, 67.61 2.1% and 51.91 3.5% in HDF or HaCaT cells respectively. TE also exhibited anti-tyrosinase and melanin inhibition properties in a dose-dependent manner. TE increased the mRNA expression of collagen-I, elastin, superoxide dismutase (SOD-2), aquaporin-3 (AQP-3), filaggrin, involucrin, transglutaminase in HDF or HaCaT cells, and decreased the mRNA levels of tyrosinase in B16F10 cells. Thus, Triphala exhibits protective benefits on skin cells in vitro and can be used as a potential ingredient in skin care formulations. PMID:26731545

  3. Effects of the basic fibroblast growth factor and its anti-factor in the healing and collagen maturation of infected skin wound Efeitos do fator de crescimento de fibroblastos básico e do seu anti-fator na cicatrização e maturação do colágeno de feridas infectadas de pele

    Directory of Open Access Journals (Sweden)

    Antonio Medeiros Dantas Filho

    2007-01-01

    Full Text Available PURPOSE: The infection is one of the main factors that affect the physiological evolution of the surgical wounds. The aim of this work is to evaluate the effects of fibroblast growth factor (FGFâ and anti-FGFâ in the healing, synthesis and maturation of collagen when topically used on infected skin wounds of rats. METHODS: An experimental study was perfomed in 60 male Wistar rats. All animals were divided in two groups (A and B. Each group was divided in three subgroups A1, B1; A2, B2 and A3, B3. After anesthesia with pentobarbital, two open squared wounds (1cm², 4cm distant to each other, were done in the dorsal skin of all the rats. In group A (n=30 the wounds were contaminated with multibacterial standard solution, and in group B(n=30 the wounds were maintained sterile. These wounds were named F1 (for inflammation analysis and F2 (for collagen study. The open wounds of A1 and B1 rats were topically treated with saline solution, A2 and B2 were treated with FGFâ and subgroups A3 and B3 were treated with FGFâ and anti-FGFâ. The rats were observed until complete epitelization of F2 wounds for determination of healing time and the expression of types I and III collagen, using Picro Sirius Red staining. Inflammatory reaction in F1 wounds was studied using hematoxilineosin staining. The three variable was measured by the Image Pro-Plus Média Cybernetics software. The statistical analysis was performed by ANOVA and Tukey test, considering pOBJETIVO: Avaliar os efeitos do fator de crescimento de fibroblastos básico (FCFâ e do anti-FCFâ na cicatrização e maturação do colágeno em feridas infectadas na pele de ratos. MÉTODOS: Um estudo experimental foi realizado em 60 ratos Wistar, divididos em dois grupos (A e B. Cada grupo foi divididos em 03 subgrupos A1,B1; A2,B2 e A3,B3. Após anestesia com pentobarbital sódico intraperitoneal, foram feitas duas feridas abertas de 1cm² na pele no dorso distando 4cm uma da outra. Essas feridas foram denominadas feridas F1 (para análise inflamatória e F2 (para estudo do colágeno. No grupo A(n=30, as feridas foram contaminadas com solução multibateriana e no grupo B (n=30 as feridas não foram contaminadas. As feridas receberam tratamento tópico com aplicação única. Nos subgrupos A1 e B1 foram tratadas com solução salina tópica, as dos subgrupos A2 e B2 foram tratadas com o FCFâ e nos subgrupos A3 e B3 foram tratadas com FCFâ e com o anti-FCFâ. Os dados formam analisados pelos testes ANOVA de Tukey, considerando p<0,05 como significante. RESULTADOS: A infecção retardou de modo significante o tempo de cicatrização e a aplicação do FCFâ foi capaz de reverter a inibição da cicatrização provocada pela infecção(p<0.05. A resposta inflamatória foi maior nos grupos tratados com o FCFâ, e a aplicação do anti-FCFâ inibiu a reação inflamatória(p<0.05. Houve aumento significante dos colágenos tipo I e III em todos os subgrupos tratados com FCFâ, comparando com os não tratados, sendo a expressão do tipo I mais intensa do que do tipo III (p<0.05. A aplicação do anti-FCFâ inibiu a expressão das moléculas do colágeno. CONCLUSÕES: O FCFâ foi capaz de acelerar a cicatrização de feridas abertas infectadas e contribui para a maturação do colágeno, ao aumentar a expressão do colágeno tipo I, fenômeno que foi atenuado pela ação do anti-FCFâ.

  4. Effect of periodontal dressings on human gingiva fibroblasts in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Eber, R.M.; Shuler, C.F.; Buchanan, W.; Beck, F.M.; Horton, J.E. (Ohio State Univ. College of Dentistry, Columbus (USA))

    1989-08-01

    In vitro cytotoxicity studies of periodontal dressings have not generally produced a result consistent with in vivo observations. These prior in vitro studies have not used human intraoral cell lines. We tested the effects of two eugenol containing and two non-eugenol periodontal dressings on cultured human gingival fibroblasts (HGF) (ATCC No. 1292). Replicate HGF cultures grown in microtiter plates were exposed to stock, 1:4 and 1:16 dilutions of extracts made from each of the four periodontal dressings. The HGF cultures were pulse labelled with tritiated thymidine (3HTdR) after 24, 48, and 72 hours. Incorporations of the labelled thymidine were measured using liquid scintillation counting and expressed as counts per minute. The results showed that undiluted extracts from all four periodontal dressings totally inhibited 3HTdR uptake (P less than 0.05). The 1:4 dilution of eugenol dressings inhibited 3HTdR uptake significantly more than non-eugenol dressings (P less than 0.05). Interestingly, at 72 hours the 1:16 dilution of the non-eugenol dressings caused significantly increased 3HTdR uptake which was not observed with the eugenol dressings. The present results suggest that the use of a human fibroblastic cell line for testing the effects of periodontal dressings may provide information about the relative biological effects of these dressings. Using this cell line, we have found that eugenol dressings inhibit fibroblast proliferation to a greater extent than non-eugenol dressings.

  5. Effect of periodontal dressings on human gingiva fibroblasts in vitro

    International Nuclear Information System (INIS)

    In vitro cytotoxicity studies of periodontal dressings have not generally produced a result consistent with in vivo observations. These prior in vitro studies have not used human intraoral cell lines. We tested the effects of two eugenol containing and two non-eugenol periodontal dressings on cultured human gingival fibroblasts (HGF) (ATCC No. 1292). Replicate HGF cultures grown in microtiter plates were exposed to stock, 1:4 and 1:16 dilutions of extracts made from each of the four periodontal dressings. The HGF cultures were pulse labelled with tritiated thymidine (3HTdR) after 24, 48, and 72 hours. Incorporations of the labelled thymidine were measured using liquid scintillation counting and expressed as counts per minute. The results showed that undiluted extracts from all four periodontal dressings totally inhibited 3HTdR uptake (P less than 0.05). The 1:4 dilution of eugenol dressings inhibited 3HTdR uptake significantly more than non-eugenol dressings (P less than 0.05). Interestingly, at 72 hours the 1:16 dilution of the non-eugenol dressings caused significantly increased 3HTdR uptake which was not observed with the eugenol dressings. The present results suggest that the use of a human fibroblastic cell line for testing the effects of periodontal dressings may provide information about the relative biological effects of these dressings. Using this cell line, we have found that eugenol dressings inhibit fibroblast proliferation to a greater extent than non-eugenol dressings

  6. Modulation of mitochondrial morphology by bioenergetics defects in primary human fibroblasts

    DEFF Research Database (Denmark)

    Guillery, O.; Malka, F.; Frachon, P.; Milea, D.; Rojo, M.; Lombes, A.

    2008-01-01

    of mitochondrial morphology in five control cultured primary skin fibroblasts and seven with genetic alterations of oxidative phosphorylation. Under basal conditions, control fibroblasts had essentially filamentous mitochondria. Oxidative phosphorylation inhibition with drugs targeting complex I, III...... mitochondria under basal conditions, although when challenged some of them presented with mild alteration of fission or fusion efficacy. Severely defective cells disclosed complete mitochondrial fragmentation under glycolysis inhibition. In conclusion, mitochondrial morphology is modulated by D Psi m but...... loosely linked to mitochondrial oxidative phosphorylation. Its alteration by glycolysis, inhibition points to a severe oxidative phosphorylation defect. (C) 2008 Elsevier B.V. All rights reserved Udgivelsesdato: 2008/4...

  7. Capparis spinosa protects against oxidative stress in systemic sclerosis dermal fibroblasts.

    Science.gov (United States)

    Cao, Yue-Lan; Li, Xin; Zheng, Min

    2010-07-01

    High reactive oxygen species (ROS), Ha-Ras, and active ERK1/2 in fibroblasts play an essential role in the pathogenesis of systemic sclerosis (SSc). The present study was carried out to evaluate the effects of the ethanol extract from fruits of Capparis Spinosa L. (ECS) on oxidative stress and ROS-ERK1/2-Ha-Ras signal loop in SSc dermal fibroblasts in vitro. Cultured dermal fibroblasts from three SSc patients and three normal controls were treated with ECS by different concentration (10, 50, 100 microg/ml). ECS significantly reduced the production of O2(-), H2O2, and ROS in SSc fibroblasts in a dose-dependent manner. ECS effectively minimized the loss of cell viability and apoptosis induced by H2O2 in normal and SSc fibroblasts. Furthermore, the protective effect of ECS on SSc fibroblasts was more significant than on normal ones. ECS decreased the expression of P-ERK1/2 and Ha-Ras in a dose-dependent manner. In conclusion, ECS exhibits a notable activity in protecting against oxidative stress and interrupting of ROS-ERK1/2-Ha-Ras signal loop in SSc, suggesting its potential protective effects against skin sclerosis. PMID:19851777

  8. Lentiviral Engineered Fibroblasts Expressing Codon-Optimized COL7A1 Restore Anchoring Fibrils in RDEB

    Science.gov (United States)

    Georgiadis, Christos; Syed, Farhatullah; Petrova, Anastasia; Abdul-Wahab, Alya; Lwin, Su M.; Farzaneh, Farzin; Chan, Lucas; Ghani, Sumera; Fleck, Roland A.; Glover, Leanne; McMillan, James R.; Chen, Mei; Thrasher, Adrian J.; McGrath, John A.; Di, Wei-Li; Qasim, Waseem

    2016-01-01

    Cells therapies, engineered to secrete replacement proteins, are being developed to ameliorate otherwise debilitating diseases. Recessive dystrophic epidermolysis bullosa (RDEB) is caused by defects of type VII collagen, a protein essential for anchoring fibril formation at the dermal-epidermal junction. Whereas allogeneic fibroblasts injected directly into the dermis can mediate transient disease modulation, autologous gene-modified fibroblasts should evade immunological rejection and support sustained delivery of type VII collagen at the dermal-epidermal junction. We demonstrate the feasibility of such an approach using a therapeutic grade, self-inactivating-lentiviral vector, encoding codon-optimized COL7A1, to transduce RDEB fibroblasts under conditions suitable for clinical application. Expression and secretion of type VII collagen was confirmed with transduced cells exhibiting supranormal levels of protein expression, and ex vivo migration of fibroblasts was restored in functional assays. Gene-modified RDEB fibroblasts also deposited type VII collagen at the dermal-epidermal junction of human RDEB skin xenografts placed on NOD-scid IL2Rgammanull recipients, with reconstruction of human epidermal structure and regeneration of anchoring fibrils at the dermal-epidermal junction. Fibroblast-mediated restoration of protein and structural defects in this RDEB model strongly supports proposed therapeutic applications in man. PMID:26763448

  9. Pentadecapeptide BPC 157 Enhances the Growth Hormone Receptor Expression in Tendon Fibroblasts

    Directory of Open Access Journals (Sweden)

    Chung-Hsun Chang

    2014-11-01

    Full Text Available BPC 157, a pentadecapeptide derived from human gastric juice, has been demonstrated to promote the healing of different tissues, including skin, muscle, bone, ligament and tendon in many animal studies. However, the underlying mechanism has not been fully clarified. The present study aimed to explore the effect of BPC 157 on tendon fibroblasts isolated from Achilles tendon of male Sprague-Dawley rat. From the result of cDNA microarray analysis, growth hormone receptor was revealed as one of the most abundantly up-regulated genes in tendon fibroblasts by BPC 157. BPC 157 dose- and time-dependently increased the expression of growth hormone receptor in tendon fibroblasts at both the mRNA and protein levels as measured by RT/real-time PCR and Western blot, respectively. The addition of growth hormone to BPC 157-treated tendon fibroblasts dose- and time-dependently increased the cell proliferation as determined by MTT assay and PCNA expression by RT/real-time PCR. Janus kinase 2, the downstream signal pathway of growth hormone receptor, was activated time-dependently by stimulating the BPC 157-treated tendon fibroblasts with growth hormone. In conclusion, the BPC 157-induced increase of growth hormone receptor in tendon fibroblasts may potentiate the proliferation-promoting effect of growth hormone and contribute to the healing of tendon.

  10. Recombinant interferon regulation of lung fibroblast growth

    International Nuclear Information System (INIS)

    The authors characterized the effect on human lung fibroblast proliferation of recombinant interferons [gamma (γ), alpha/sub A/ (α/sub A/), alpha/sub D/ (α/sub D/) and beta (β)]. Quiescent and log phase fibroblasts were used. Proliferation was assessed using 3HTdr uptake and cell counts. γ stimulated the 3HTdr uptake of quiescent cells. Maximal stimulation was seen between 100 and 500 IU/ml. Fibroblasts incubated in 100 IU/ml incorporated 160 +/- 9% as much 3HTdr as quiescent controls (p 3HTdr uptake of log phase cells. Fibroblasts incubated in 1000 IU/ml incorporated 41 +/- 9% as much 3HTdr as log phase controls (p 3HTdr as log phase controls (p < .03 for all). γ, α/sub A/, α/sub D/ and β did not stimulate fibroblast prostaglandin E (PGE) production and incubating fibroblasts with indomethacin did not reverse the inhibition of fibroblast proliferation caused by the interferons. Thus, (1) γ can stimulate or inhibit fibroblast growth, (2) α/sub A/, α/sub D/ and β only inhibit fibroblast growth, (3) the inhibition of fibroblast growth caused by γ, α/sub A/, α/sub D/ and β is largely independent of fibroblast PGE production

  11. Study of apoptosis inducing activity of calprotectin on fibroblast cell

    Directory of Open Access Journals (Sweden)

    Amin Rostami

    2010-01-01

    Full Text Available One of the prominent types of connective tissue cells is fibroblast that synthesizes and maintains the extracellular matrix of many animal tissues. Previous studies illustrated that calprotectin protein has different cytotoxicity effects on fibroblast cells. Calprotectin is abundant in the neutrophil cytosol; it has growth-inhibitory and apoptosis-inducing activities against various cell types such as tumor cells. The present study tries to introduce mechanism of growth inhibitory effect of calprotectin on human foreskin fibroblast cells (HFFF and compare to etoposide (chemotherapy agent as control. Calprotectin was purified from human neutrophil by chromatography methods. HFFF cell lines were used, maintained in RPMI 1640 medium supplemented with 10% FCS in a humidified incubator (37 C & 5% CO2. The HFFF cells were exposed to the different concentrations of calprotectin and etoposide for 24, 48 and 72 hours. Cell proliferation was assessed by using dimethylthiazol diphenyl tetrazolium bromide assay. Flow cytometric analysis was performed to evaluate the cytotoxic mechanism of calprotectin on HFFF cells. Our results revealed that calprotectin and etoposide induce growth inhibition of HFFF in dose- and time-dependent manners. Sensitivity of HFFF cells to cytotoxic effect of human calprotectin was highly remarkable. In addition, growth inhibitory effect of this cytotoxic agent mostly was governed through induction of apoptosis in the HFFF cells. Taken together, calprotectin not only has more potent anticancer activity in comparison with the etoposide, but it also is an apoptosis inducer that acts on the proliferation of normal cells like fibroblasts.

  12. Skin Pigmentation Disorders

    Science.gov (United States)

    Pigmentation means coloring. Skin pigmentation disorders affect the color of your skin. Your skin gets its color from a pigment called melanin. Special cells in the skin make melanin. When these cells become damaged or ...

  13. Skin, Hair, and Nails

    Science.gov (United States)

    ... Caring for Your Child All About Food Allergies Skin, Hair, and Nails KidsHealth > For Parents > Skin, Hair, ... La piel, el cabello y las uñas About Skin, Hair and Nails Skin is our largest organ. ...

  14. Skin Care and Aging

    Science.gov (United States)

    ... Home » Skin Care and Aging Heath and Aging Skin Care and Aging Dry Skin and Itching Bruises Wrinkles Age Spots ... doctor. For More Information About Skin Care and Aging American Academy of Dermatology 1-866-503-7546 ( ...

  15. Single Exposure of Human Oral Mucosa Fibroblasts to Ultraviolet B Radiation Reduces Proliferation and Induces COX-2 Expression and Activation

    Directory of Open Access Journals (Sweden)

    Y Boza

    2010-12-01

    Full Text Available The lip vermillion constitutes a transition tissue, between oral mucosa and skin, where oral mucosal cells from epithelial and connective tissue compartments are exposed to ultraviolet (UV sunlight. Fibroblasts are abundant resident cells of the connective tissue which are key regulators of extracellular matrix composition, as well as, epithelial and endothelial cell function. UVB light, an inherent component of sunlight, causes several alterations in skin fibroblasts, including premature senescence and increased cyclooxygenase (COX-2 expression. To assess if UVB irradiation had similar effects on fibroblasts derived from human oral mucosa (HOM, primary cultures of HOM fibroblasts were irradiated with a single dose of 30 or 60 mJ/cm²of UVB light or sham-irradiated. Fibroblast proliferation was assessed from 3 to 48 hrs after UVB-irradiation utilizing [³H]-thymidine incorporation and MTT assays. In addition, COX-2 mRNA expression was detected by RT-PCR, and PGE2 production was assessed using enzyme immunoassay from 0.5 to 24 hrs after UVB-irradiation. The results showed a significant decrease in proliferation of UVB-irradiated HOM fibroblasts as compared to controls as measured by both [³H]-thymidine incorporation and MTT assays (p<0.001. HOM fibroblasts had increased COX-2 mRNA expression at 0.5 and 12 hrs after irradiation, and PGE2 production was elevated at 12 and 24 hrs post-irradiation as compared to controls (p<0.05. The results showed an inhibitory effect of a single dose of UVB irradiation on HOM fibroblast proliferation with an increase in COX-2 expression and activation. Therefore, photodamaged fibroblasts may play and important role in the pathogenesis of UV-induced lesions of the lip.

  16. Skin aging:

    OpenAIRE

    Puizina-Ivić, Neira

    2008-01-01

    There are two main processes that induce skin aging: intrinsic and extrinsic. A stochastic process that implies random cell damage as a result of mutations during metabolic processes due to the production of free radicals is also implicated. Extrinsic aging is caused by environmental factors such as sun exposure, air pollution, smoking, alcohol abuse, and poor nutrition. Intrinsicaging reflects the genetic background and depends on time. Various expressions of intrinsic aging include smooth, ...

  17. Rapid creation of skin substitutes from human skin cells and biomimetic nanofibers for acute full-thickness wound repair.

    Science.gov (United States)

    Mahjour, Seyed Babak; Fu, Xiaoling; Yang, Xiaochuan; Fong, Jason; Sefat, Farshid; Wang, Hongjun

    2015-12-01

    Creation of functional skin substitutes within a clinically acceptable time window is essential for timely repair and management of large wounds such as extensive burns. The aim of this study was to investigate the possibility of fabricating skin substitutes via a bottom-up nanofiber-enabled cell assembly approach and using such substitutes for full-thickness wound repair in nude mice. Following a layer-by-layer (L-b-L) manner, human primary skin cells (fibroblasts and keratinocytes) were rapidly assembled together with electrospun polycaprolactone (PCL)/collagen (3:1, w/w; 8%, w/v) nanofibers into 3D constructs, in which fibroblasts and keratinocytes were located in the bottom and upper portion respectively. Following culture, the constructs developed into a skin-like structure with expression of basal keratinocyte markers and deposition of new matrix while exhibiting good mechanical strength (as high as 4.0MPa by 14 days). Treatment of the full-thickness wounds created on the back of nude mice with various grafts (acellular nanofiber meshes, dermal substitutes, skin substitutes and autografts) revealed that 14-day-cultured skin substitutes facilitated a rapid wound closure with complete epithelialization comparable to autografts. Taken together, skin-like substitutes can be formed by L-b-L assembling human skin cells and biomimetic nanofibers and they are effective to heal acute full-thickness wounds in nude mice. PMID:26187057

  18. Ellagic acid plays a protective role against UV-B-induced oxidative stress by up-regulating antioxidant components in human dermal fibroblasts

    Science.gov (United States)

    Baek, Beomyeol; Lee, Su Hee; Lim, Hye-Won

    2016-01-01

    Ellagic acid (EA), an antioxidant polyphenolic constituent of plant origin, has been reported to possess diverse pharmacological properties, including anti-inflammatory, anti-tumor and immunomodulatory activities. This work aimed to clarify the skin anti-photoaging properties of EA in human dermal fibroblasts. The skin anti-photoaging activity was evaluated by analyzing the reactive oxygen species (ROS), matrix metalloproteinase-2 (MMP-2), total glutathione (GSH) and superoxide dismutase (SOD) activity levels as well as cell viability in dermal fibroblasts under UV-B irradiation. When fibroblasts were exposed to EA prior to UV-B irradiation, EA suppressed UV-B-induced ROS and proMMP-2 elevation. However, EA restored total GSH and SOD activity levels diminished in fibroblasts under UV-B irradiation. EA had an up-regulating activity on the UV-B-reduced Nrf2 levels in fibroblasts. EA, at the concentrations used, was unable to interfere with cell viabilities in both non-irradiated and irradiated fibroblasts. In human dermal fibroblasts, EA plays a defensive role against UV-B-induced oxidative stress possibly through an Nrf2-dependent pathway, indicating that this compound has potential skin antiphotoaging properties. PMID:27162481

  19. Stress responses of human dermal fibroblasts exposed to zinc pyrithione.

    Science.gov (United States)

    Rudolf, Emil; Cervinka, Miroslav

    2011-07-28

    Zinc pyrithione is used as a topical agent in a range of medicinal and cosmetic applications. Despite its extensive use and reported beneficial effects in treatment of various dermal problems, its potential toxicity towards skin cells remains relatively underexplored. In this work we investigated effects of nM zinc pyrithione on cell stress response pathways of primary human skin fibroblasts during 24h of exposure. We demonstrate that zinc pyrithione-induced cytotoxity in dermal fibroblasts is dose-dependent and it associates with increased intracellular zinc concentrations and activated stress response pathways including p53 and stress kinase p38. Higher zinc pyrithione concentrations (500nM and above) stimulate oxidative stress and moderate DNA damage which occur in the presence of activated p38 kinase. Cells further upregulate the expression of p53 which increases its transcriptional activity while mitogenic signaling exemplified by mTOR (mammalian target of rapamycin) expression is suppressed and these steps lead to mitochondrial, caspase-dependent apoptosis. Conversely, lower zinc concentrations (125nM) fail to induce oxidative stress and significant DNA damage; however, treated cells still activate p38 and upregulate the expression and transcriptional activity of p53 and its target gene p21 as well as the expression of p16 in the presence of active mTOR pathway and a changed DNA methylation pattern. The end result is premature senescence phenotype. Specific pharmacological inhibitors as well as gene knockdown technology prove that an interaction between p38, p53 and mTOR might be responsible for these observed endpoints. Taken together, exposure of dermal fibroblasts to varying concentrations of zinc pyrithione may result in either cell death-apoptosis or cellular premature senescence which attests to the ability of this compound to affect this type of cells in an in vitro model system. PMID:21557991

  20. Variable radiosensitivity in fibroblasts from patients with tuberous sclerosis

    International Nuclear Information System (INIS)

    It has been reported that some of the cultured cell strains derived from patients with tuberous sclerosis (TS) showed hypersensitivity to gamma-rays or a radiomimetic chemical. Thirteen fibroblast cell strains from 11 patients with TS were examined for their sensitivity to x-rays as determined from their colony-forming ability. All strains derived from normal-appearing skin of patients, either sporadic or familial cases, showed sensitivity within the normal control range. Five cell strains originating from tumorous skin of 3 patients did not show hypersensitivity. It was concluded that the sensitivity to x-rays of cultured cells of TS is essentially normal. However, the mean D0 or D10 values of the strains from tumorous skin tended to be lower compared to those for normal skin of patients. In addition, the hypersensitivity to x-rays was confirmed in the cell strains of TS which had been shown to be hypersensitive to gamma-rays. These results appear to indicate that at least some of the cells of TS are liable to change to exhibit a hypersensitive trait in unknown acquired conditions

  1. Transplantation of Cultivated Fibroblasts on a Backing of Xenogenic Tissue in the Treatment of Wounds

    Directory of Open Access Journals (Sweden)

    Kabylbek Abugaliyev

    2014-12-01

    Full Text Available Introduction. Trophic ulcers are a common health problem, and there are numerous treatment methods. Irreversible damage in the skin, subcutaneous tissue, and fascia with long-term ulcer existence make standard autotransplantation inneffective. Skin grafts are often complicated by partial or complete rejection of skin flaps. The aim of this study was to examine the feasibility of using transplanted cultivated allogenic fibroblasts on the backing of a cellularless xenogenic fabric for wound healing.Methods. Transplantation of cultured embryonic fibroblasts on a backing of xenogenic tissue was used in the complex treatment of trophic ulcers for stimulation of regenerative processes. Decellularization xenogenic film was previously held. Then allogenic fibroblasts were cultivated on the surface of collagen-elastin matrix. Since 2013, we treated 12 patients with giant ulcers caused by the following: lymphedema (2 patients, vascular disease (3 patients, diabetes (2 patients, after injury (4 patients, and radiation ulcer (1 patient. Dimensions of ulcers were from 150 to 600 cm2. Duration of the lower limb ulcers ranged from 8 months to 10 years. For a number of years, all patients were on a complex therapy, which had not resulted in healing wounds. During the operation when excision of granulation tissue was performed, plastic wounds perforated with the ratio 1:2 autoskin. Xenogenic fabric with cultured fibroblasts was applied on top. In this case, xenogenic film protected the skin from drying, created optimal microclimate, and cultured fibroblasts stimulating regeneration and improving engraftment.Results. The first redress was held on the fifth day. In all cases, the results of engraftment skin grafts achieved maximum possible (100% and optimal (90%. Complete epithelialization of the cell perforation was seen in five patients on the fifth day and three on seventh day after skin plastics. Average period of inpatient treatment was 20.7 days. All patients were discharged with healed wounds.Conclusion. Thus, the treatment of trophic ulcers can be successfully solved using advances in biotechnology. Transplantation of cultivated allogenic fibroblasts on a backing of cellularless xenogenic fabric shows good clinical results due to the stimulation of regenerative processes and creates the optimum environment for autotransplants. 

  2. Cutaneous skin tag

    Science.gov (United States)

    Skin tag; Acrochordon; Fibroepithelial polyp ... have diabetes. They are thought to occur from skin rubbing against skin. ... The tag sticks out of the skin and may have a short, narrow stalk connecting it to the surface of the skin. Some skin tags are as long as ...

  3. Differentiation of human umbilical cord mesenchymal stem cells into dermal fibroblasts in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Han, Yanfu [Department of Burn and Plastic Surgery, Burns Institute, First Hospital Affiliated to General Hospital of PLA, Beijing (China); Chai, Jiake, E-mail: cjk304@126.com [Department of Burn and Plastic Surgery, Burns Institute, First Hospital Affiliated to General Hospital of PLA, Beijing (China); Sun, Tianjun; Li, Dongjie; Tao, Ran [Department of Burn and Plastic Surgery, Burns Institute, First Hospital Affiliated to General Hospital of PLA, Beijing (China)

    2011-10-07

    Highlights: {yields} Mesenchymal stem cells (MSCs) are potential seed cells for tissue-engineered skin. {yields} Tissue-derived umbilical cord MSCs (UCMSCs) can readily be isolated in vitro. {yields} We induce UCMSCs to differentiate into dermal fibroblasts via conditioned medium. {yields} Collagen type I and collagen type III mRNA level was higher in differentiated cells. {yields} UCMSCs-derived fibroblast-like cells strongly express fibroblast-specific protein. -- Abstract: Tissue-derived umbilical cord mesenchymal stem cells (UCMSCs) can be readily obtained, avoid ethical or moral constraints, and show excellent pluripotency and proliferation potential. UCMSCs are considered to be a promising source of stem cells in regenerative medicine. In this study, we collected newborn umbilical cord tissue under sterile conditions and isolated UCMSCs through a tissue attachment method. UCMSC cell surface markers were examined using flow cytometry. On the third passage, UCMSCs were induced to differentiate into dermal fibroblasts in conditioned induction media. The induction results were detected using immunofluorescence with a fibroblast-specific monoclonal antibody and real time PCR for type I and type III collagen. UCMSCs exhibited a fibroblast-like morphology and reached 90% confluency 14 to 18 days after primary culture. Cultured UCMSCs showed strong positive staining for CD73, CD29, CD44, CD105, and HLA-I, but not CD34, CD45, CD31, or HLA-DR. After differentiation, immunostaining for collagen type I, type III, fibroblast-specific protein, vimentin, and desmin were all strongly positive in induced cells, and staining was weak or negative in non-induced cells; total transcript production of collagen type I and collagen type III mRNA was higher in induced cells than in non-induced cells. These results demonstrate that UCMSCs can be induced to differentiate into fibroblasts with conditioned induction media and, in turn, could be used as seed cells for tissue-engineered dermis.

  4. Differentiation of human umbilical cord mesenchymal stem cells into dermal fibroblasts in vitro

    International Nuclear Information System (INIS)

    Highlights: → Mesenchymal stem cells (MSCs) are potential seed cells for tissue-engineered skin. → Tissue-derived umbilical cord MSCs (UCMSCs) can readily be isolated in vitro. → We induce UCMSCs to differentiate into dermal fibroblasts via conditioned medium. → Collagen type I and collagen type III mRNA level was higher in differentiated cells. → UCMSCs-derived fibroblast-like cells strongly express fibroblast-specific protein. -- Abstract: Tissue-derived umbilical cord mesenchymal stem cells (UCMSCs) can be readily obtained, avoid ethical or moral constraints, and show excellent pluripotency and proliferation potential. UCMSCs are considered to be a promising source of stem cells in regenerative medicine. In this study, we collected newborn umbilical cord tissue under sterile conditions and isolated UCMSCs through a tissue attachment method. UCMSC cell surface markers were examined using flow cytometry. On the third passage, UCMSCs were induced to differentiate into dermal fibroblasts in conditioned induction media. The induction results were detected using immunofluorescence with a fibroblast-specific monoclonal antibody and real time PCR for type I and type III collagen. UCMSCs exhibited a fibroblast-like morphology and reached 90% confluency 14 to 18 days after primary culture. Cultured UCMSCs showed strong positive staining for CD73, CD29, CD44, CD105, and HLA-I, but not CD34, CD45, CD31, or HLA-DR. After differentiation, immunostaining for collagen type I, type III, fibroblast-specific protein, vimentin, and desmin were all strongly positive in induced cells, and staining was weak or negative in non-induced cells; total transcript production of collagen type I and collagen type III mRNA was higher in induced cells than in non-induced cells. These results demonstrate that UCMSCs can be induced to differentiate into fibroblasts with conditioned induction media and, in turn, could be used as seed cells for tissue-engineered dermis.

  5. Ultrastructure of elastosis in facial rhytidectomy skin

    International Nuclear Information System (INIS)

    Skin from 19 facial rhytidectomies performed in patients with chronic solar damage was compared with postauricular skin from patients of similar age. Light microscopy demonstrated large areas of amorphous material that stained PAS positive in all 19 face-lift specimens, while none of the controls had such material. Electron microscopy of the ''elastotic'' material revealed large amorphous masses of granular material, with loss of the microfilament component of normal elastin. Current theories suggest that the elastotic material in solar-damaged skin is a product of radiation-damaged fibroblasts, rather than being either collagen or degenerated elastin. Such knowledge may help the plastic surgeons encourage rhytidectomy patients to protect themselves from solar radiation

  6. Shear instability in skin tissue

    CERN Document Server

    Ciarletta, Pasquale; Gower, Artur L

    2013-01-01

    We propose two toy-models to describe, predict, and interpret the wrinkles appearing on the surface of skin when it is sheared. With the first model, we account for the lines of greatest tension present in human skin by subjecting a layer of soft tissue to a pre-stretch, and for the epidermis by endowing one of the layer's faces with a surface tension. For the second model, we consider an anisotropic model for the skin, to reflect the presence of stiff collagen fibres in a softer elastic matrix. In both cases, we find an explicit bifurcation criterion, linking geometrical and material parameters to a critical shear deformation accompanied by small static wrinkles, with decaying amplitudes normal to the free surface of skin.

  7. Effect of estrogens on skin aging and the potential role of SERMs

    OpenAIRE

    Susan Stevenson; Julie Thornton

    2007-01-01

    Susan Stevenson1, Julie Thornton21Burns & Plastic Surgery Research Unit, 2Cutaneous Research, Medical Biosciences, School of Life Sciences, University of Bradford, Bradford, UKAbstract: In humans, structural and functional changes attributable to aging are more visibly evident in the skin than in any other organ. Estrogens have significant effects on skin physiology and modulate epidermal keratinocytes, dermal fibroblasts and melanocytes, in addition to skin appendages including the h...

  8. Dermal fibroblasts from red Duroc and Yorkshire pigs exhibit intrinsic differences in the contraction of collagen gels.

    Science.gov (United States)

    de Hemptinne, Isabelle; Gallant-Behm, Corrie L; Noack, Cassandra L; Parreno, Justin; Hart, David A

    2008-01-01

    Previous studies have shown that the Yorkshire (Y) pig is a model for normal skin wound healing, while red Duroc (RD) pigs form hypercontracted scars similar to human hypertrophic scars. In order to determine potential intrinsic differences in fibroblast phenotypes, the ability of normal dorsal and ventral dermal fibroblasts from Y and RD pigs to contract collagen gels was assessed. Cells plated in gels were cultured in media supplemented with 2% or 10% FBS +/- 1 or 10 ng/mL transforming growth factor beta1. The degree of contraction of the gels was quantified at defined time-points postrelease. Final contraction levels were dependent on cell density and serum concentration for all cell types. The rates of contraction of RD dorsal fibroblasts were significantly greater than those for Y dorsal fibroblasts. Immunocytochemical analysis revealed the presence of alpha-smooth muscle actin in contracted cells. Furthermore, mRNA levels for matrix metalloproteinase-2 and decorin showed specific increases for the RD cells during contraction. These findings have revealed intrinsically different, location-specific in vitro responses with normal dermal fibroblasts from the two breeds of pig, suggesting that the abnormal skin healing phenotype of RD pigs may be attributable in part to intrinsic genetic differences in fibroblasts between the breeds. PMID:18211585

  9. Correlation between the radiation responses of fibroblasts cultured from individual patients and the risk of late reaction after breast radiotherapy.

    Science.gov (United States)

    Nuta, Otilia; Somaiah, Navita; Boyle, Sue; Chua, Melvin Lee Kiang; Gothard, Lone; Yarnold, John; Rothkamm, Kai; Herskind, Carsten

    2016-05-01

    Late normal tissue toxicity varies widely between patients and limits breast radiotherapy dose. Here we aimed to determine its relationship to DNA damage responses of fibroblast cultures from individual patients. Thirty-five breast cancer patients, with minimal or marked breast changes after breast-conserving therapy consented to receive a 4 Gy test irradiation to a small skin field of the left buttock and have punch biopsies taken from irradiated and unirradiated skin. Early-passage fibroblast cultures were established by outgrowth and irradiated in vitro with 0 or 4 Gy. 53BP1 foci, p53 and p21/CDKN1A were detected by immunofluorescence microscopy. Residual 53BP1 foci counts 24 h after in vitro irradiation were significantly higher in fibroblasts from RT-sensitive versus RT-resistant patients. Furthermore, significantly larger fractions of p53- but not p21/CDKN1A-positive fibroblasts were found in cultures from RT-sensitive patients without in vitro irradiation, and 2 h and 6 d post-irradiation. Exploratory analysis showed a stronger p53 response 2 h after irradiation of fibroblasts established from patients with severe reaction. These results associate the radiation response of fibroblasts with late reaction of the breast after RT and suggest a correlation with severity. PMID:26944319

  10. The IGF-1/IGF-1R signaling axis in the skin: a new role for the dermis in aging-associated skin cancer

    OpenAIRE

    Lewis, Davina A.; Travers, Jeffrey B.; Somani, Ally-Khan; Spandau, Dan F.

    2009-01-01

    The appropriate response of human keratinocytes to UVB is dependent on the activation status of the IGF-1 receptor. Keratinocytes grown in conditions where the IGF-1 receptor is inactive, inappropriately replicate in the presence of UVB-induced DNA damage. In human skin epidermal keratinocytes do not express IGF-1, so the IGF-1 receptor on keratinocytes is activated by IGF-1 secreted from dermal fibroblasts. We now demonstrate that the IGF-1 produced by human fibroblasts is essential for the ...

  11. Inhibition of histone deacetylase activity attenuates renal fibroblast activation and interstitial fibrosis in obstructive nephropathy.

    Science.gov (United States)

    Pang, Maoyin; Kothapally, Jagan; Mao, Haiping; Tolbert, Evelyn; Ponnusamy, Murugavel; Chin, Y Eugene; Zhuang, Shougang

    2009-10-01

    Activation of renal interstitial fibroblasts is critically involved in the development of tubulointerstitial fibrosis in chronic kidney diseases. In this study, we investigated the effect of trichostatin A (TSA), a specific histone deacetylase (HDAC) inhibitor, on the activation of renal interstitial fibroblasts in a rat renal interstitial fibroblast line (NRK-49F) and the development of renal fibrosis in a murine model of unilateral ureteral obstruction (UUO). alpha-Smooth muscle actin (alpha-SMA) and fibronectin, two hallmarks of fibroblast activation, were highly expressed in cultured NRK-49F cells, and their expression was inhibited in the presence of TSA. Similarly, administration of TSA suppressed the expression of alpha-SMA and fibronectin and attenuated the accumulation of renal interstitial fibroblasts in the kidney after the obstructive injury. Activation of renal interstitial fibroblasts was accompanied by phosphorylation of signal transducer and activator of transcription 3 (STAT3), and TSA treatment also abolished these responses. Furthermore, inhibition of the STAT3 pathway with AG490 inhibited expression of alpha-SMA and fibronectin in NRK-49F cells. Finally, TSA treatment inhibited tubular cell apoptosis and caspase-3 activation in the obstructive kidney. Collectively, we suggest that pharmacological HDAC inhibition may induce antifibrotic activity by inactivation of renal interstitial fibroblasts and inhibition of renal tubular cell death. STAT3 may mediate those actions of HDACs. PMID:19640900

  12. Dermal Wound Fibroblasts and Matrix Metaloproteinases (MMPs): Their Possible Role in Allergic Contact Dermatitis.

    Science.gov (United States)

    Khorramizadeh, Mohammad Reza; Falak, Reza; Pezeshki, Mohammad; Safavifar, Farnaz; Mansouri, Parvin; Ghahary, Aziz; Saadat, Farshid; Varshokar, Kamran

    2004-03-01

    This study was conducted to examine if allergic contact dermatitis (ACD) alters the expression of MMPs in human dermal fibroblasts. Fibroblasts are the primary source for MMP and matrix production in skin. MMPs are known to involve in a number of physiological and pathological processes. Some published data indicated a gelatinase-like activity in acute and chronic phases of allergic contact dermatitis. However, no exact source of gelatinase activity was demonstrated. Moreover, little is known about the role of MMPs in immune responses.To study and predict the pathophysiological effects of (MMP-2) in allergic contact dermatitic (ACD) patients, we established an in vitro tissue culture survey based on fibroblast explanted from ACD wounds and normal tissues respectively. We also employed a precise proliferation assay [i.e. MTT; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] to analyze and compare three ACD vs. three normal cell strains. Parallel to MTT assay, we assessed the activity as well as the kinetics of gelatinase (MMP-2) in conditioned media using a zymogeraphy analysis. There was a significant difference in proliferation capacity between mean ACD fibroblast strains vs. mean normal cells, particularly in days 6 to 8 post explantation, 492.5+/-6.6 vs. 361.75+/-8.25 respectively. Zymoanalyses indicated significant differences between ACD cells and normal fibroblasts both in time-course and MMP-2 activity per cell fashions, 163.7+/-16.21 for mean ACD fibroblasts vs. 130+/-9.09 for normal cells respectively. These data suggest that fibroblasts overproliferated in the process of ACD. Moreover, simultaneous overexpression of MMPs observed in ACD fibroblasts vs. normal strains, is indicative of altered fibroblast functionality in the process of allergic contactdermatitis. The activity per cell analysis showed that MMP-2 expression in ACD fibroblasts is independent of cell number, suggesting that either intra- or inter-cellular control signals are also altered and that ACD fibroblasts exhibit hyper-responsiveness to mitogenic or fibrogenic stimulants. Altogether, these data address the chronocity and non-healing tendency of ACD wounds. However, more studies are required to examine possible MMPs inhibition and differential expression of mytogenic, fibrogenic and antifibrogenic cytokines in ACD wound beds. In particular, MMP-2 is postulated to be an aim for further gene therapy protocols. PMID:17301385

  13. Dermal Wound Fibroblasts and Matrix Metaloproteinases (MMPs: Their Possible Role in Allergic Contact Dermatitis

    Directory of Open Access Journals (Sweden)

    Mohammad Reza Khorramizadeh

    2004-03-01

    Full Text Available This study was conducted to examine if allergic contact dermatitis (ACD alters the expression of MMPs in human dermal fibroblasts. Fibroblasts are the primary source for MMP and matrix production in skin. MMPs are known to involve in a number of physiological and pathological processes. Some published data indicated a gelatinase-like activity in acute and chronic phases of allergic contact dermatitis. However, no exact source of gelatinase activity was demonstrated. Moreover, little is known about the role of MMPs in immune responses.To study and predict the pathophysiological effects of (MMP-2 in allergic contact dermatitic (ACD patients, we established an in vitro tissue culture survey based on fibroblast explanted from ACD wounds and normal tissues respectively. We also employed a precise proliferation assay [i.e. MTT; 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide] to analyze and compare three ACD vs. three normal cell strains. Parallel to MTT assay, we assessed the activity as well as the kinetics of gelatinase (MMP-2 in conditioned media using a zymogeraphy analysis. There was a significant difference in proliferation capacity between mean ACD fibroblast strains vs. mean normal cells, particularly in days 6 to 8 post explantation, 492.5±6.6 vs. 361.75±8.25 respectively. Zymoanalyses indicated significant differences between ACD cells and normal fibroblasts both in time-course and MMP-2 activity per cell fashions, 163.7±16.21 for mean ACD fibroblasts vs. 130±9.09 for normal cells respectively. These data suggest that fibroblasts overproliferated in the process of ACD.  Moreover, simultaneous overexpression of MMPs observed in ACD fibroblasts vs. normal strains, is indicative of altered fibroblast functionality in the process of allergic contactdermatitis. The activity per cell analysis showed that MMP-2 expression in ACD fibroblasts is independent of cell number, suggesting that either intra- or inter-cellular control signals are also altered and that ACD fibroblasts exhibit hyper-responsiveness to mitogenic or fibrogenic stimulants. Altogether, these data address the chronocity and non-healing tendency of ACD wounds. However, more studies are required to examine possible MMPs inhibition and differential expression of mytogenic, fibrogenic and antifibrogenic cytokines in ACD wound beds. In particular, MMP-2 is postulated to be an aim for further gene therapy protocols.

  14. Question of bone marrow stromal fibroblast traffic

    International Nuclear Information System (INIS)

    Bone marrow stromal fibroblasts (CFU-F) normally do not exchange bone marrow sites in vivo. Restitution of the CFU-F after radiation damage is primarily recovery by the local fibroblasts from potentially lethal damage. Migration of stromal fibroblasts from shielded sites to an irradiated site makes a minimal contribution, if any, to CFU-F recovery. Determination of the relative contribution of donor stromal cells in bone marrow transplants by karyotyping the proliferating bone marrow stromal cells in vitro may not reflect the relative distribution of fibroblasts in the marrow. If there is residual damage to the host stromal fibroblasts from treatment before transplantation, these cells may not be able to proliferate in vitro. Therefore, an occasional transplanted fibroblast may contribute most of the metaphase figures scored for karyotype

  15. Radiation sensitivity of Merkell cell carcinoma cell lines

    International Nuclear Information System (INIS)

    Purpose: Merkel cell carcinoma (MCC), being a small cell carcinoma, would be expected to be sensitive to radiation. Clinical analysis of patients at our center, especially those with macroscopic disease, would suggest the response is quite variable. We have recently established a number of MCC cell lines from patients prior to radiotherapy, and for the first time are in a position to determine their sensitivity under controlled conditions. Methods and Materials: Some of the MCC lines grew as suspension cultures and could not be single cell cloned; therefore, it was not possible to use clonogenic survival for all cell lines. A tetrazolium based (MTT) assay was used for these lines, to estimate cell growth after γ irradiation. Control experiments were conducted on lymphoblastoid cell lines (LCL) and the adherent MCC line, MCC13, to demonstrate that the two assays were comparable under the conditions used. Results: We have examined cell lines from MCC, small cell lung cancer (SCLC), malignant melanomas, Epstein Barr virus (EBV) transformed lymphocytes (LCL), and skin fibroblasts for their sensitivity to γ irradiation using both clonogenic cell survival and MTT assays. The results show that the tumor cell lines have a range of sensitivities, with melanoma being more resistant (surviving fraction at 2 Gy (SF2) 0.57 and 0.56) than the small cell carcinoma lines, MCC (SF2 range 0.21-0.45, mean SF2 0.30, n = 8) and SCLC (SF2 0.31). Fibroblasts were the most sensitive (SF2 0.13-0.20, mean 0.16, n = 5). The MTT assay, when compared to clonogenic assay for the MCC13 adherent line and the LCL, gave comparable results under the conditions used. Conclusion: Both assays gave a range of SF2 values for the MCC cell lines, suggesting that these cancers would give a heterogeneous response in vivo. The results with the two derivative clones of MCC14 (SF2 for MCC14/1 0.38, MCC14/2 0.45) would further suggest that some of them may develop resistance during clonogenic evolution

  16. Radiation sensitivity of Merkel cell carcinoma cell lines

    International Nuclear Information System (INIS)

    Merkel cell carcinoma (MCC), being a small cell carcinoma, would be expected to be sensitive to radiation. Clinical analysis of patients at our center, especially those with macroscopic disease, would suggest the response is quite variable. We have recently established a number of MCC cell lines from patients prior to radiotherapy, and for the first time are in a position to determine their sensitivity under controlled conditions. Some of the MCC lines grew as suspension cultures and could not be single cell cloned; therefore, it was not possible to use clonogenic survival for all cell lines. A tetrazolium based (MTT) assay was used for these lines, to estimate cell growth after ? irradiation. Control experiments were conducted on lymphoblastoid cell lines (LCL) and the adherent MCC line, MCC13, to demonstrate that the two assays were comparable under the conditions used. We have examined cell lines from MCC, small cell lung cancer (SCLC), malignant melanomas, Epstein Barr virus (EBV) transformed lymphocytes (LCL), and skin fibroblasts for their sensitivity to ? irradiation using both clonogenic cell survival and MTT assays. The results show that the tumor cell lines have a range of sensitivities, with melanoma being more resistant (surviving fraction at 2 Gy (SF2) 0.57 and 0.56) than the small cell carcinoma lines, MCC (SF2 range 0.21-0.45, mean SF2 0.30, n = 8) and SCLC (SF2 0.31). Fibroblasts were the most sensitive (SF2 0.13-0.20, mean 0.16, n = 5). The MTT assay, when compared to clonogenic assay for the MCC13 adherent line and the LCL, gave comparable results under the conditions used. Both assays gave a range of SF2 values for the MCC cell lines, suggesting that these cancers would give a heterogeneous response in vivo. The results with the two derivative clones of MCC14 (SF2 for MCC14/1 0.38, MCC14/2 0.45) would further suggest that some of them may develop resistance during clonogenic evolution. 25 refs., 3 figs., 1 tab

  17. Cytotoxic and genotoxic characterization of titanium dioxide, gadolinium oxide, and poly(lactic-co-glycolic acid) nanoparticles in human fibroblasts.

    Science.gov (United States)

    Setyawati, Magdiel Inggrid; Khoo, Pheng Kian Stella; Eng, Bao Hui; Xiong, Sijing; Zhao, Xinxin; Das, Gautom Kumar; Tan, Timothy Thatt-Yang; Loo, Joachim Say Chye; Leong, David Tai; Ng, Kee Woei

    2013-03-01

    Engineered nanomaterials have become prevalent in our everyday life. While the popularity of using nanomaterials in consumer products continues to rise, increasing awareness of nanotoxicology has also fuelled efforts to accelerate our understanding of the ill effects that different nanomaterials can bring to biological systems. In this study, we investigated the potential cytotoxicity and genotoxicity of three nanoparticles: titanium dioxide (TiO(2)), terbium-doped gadolinium oxide (Tb-Gd(2)O(3)), and poly(lactic-co-glycolic acid) (PLGA). To evaluate nanoparticle-induced genotoxicity more realistically, a human skin fibroblast cell line (BJ) with less mutated genotype compared with cancer cell line was used. The nanoparticles were first characterized by size, morphology, and surface charge. Cytotoxicity effects of the nanoparticles were then evaluated by monitoring the proliferation of treated BJ cells. Genotoxic influence was ascertained by profiling DNA damage via detection of ?H2AX expression. Our results suggested that both TiO(2) and Tb-Gd(2)O(3) nanoparticles induced cytotoxicity in a dose dependent way on BJ cells. These two nanomaterials also promoted genotoxicity via DNA damage. On the contrary, PLGA nanoparticles did not induce significant cytotoxic or genotoxic effects on BJ cells. PMID:22927021

  18. Differential gene expression induced by high LET charged particles in normal human fibroblasts

    International Nuclear Information System (INIS)

    We investigated differential gene expression of normal human skin HSF42 fibroblasts induced by heavy ions using cDNA microarray technology. Irradiation with 3 types of heavy ions was performed at Heavy Ion Medical Accelerator in Chiba (HIMAC) facility. Out of 7458 genes, we found 61 significant genes (40 up-regulated and 21 down-regulated) that distinguished between human skin fibroblast HSF42 cells non-irradiated and irradiated with 1 Gy of neon particles and 62 significant genes (48 up-regulated and 14 down-regulated) that distinguished between HSF42 cells non-irradiated and irradiated with 1 Gy of silicon particles. Furthermore, we are going to analyze profiles of HSF42 cells exposed to carbon particles and compare those profiles between different types of beams. (author)

  19. Study of Fibroblast and Platelet Adhesion on Multi―wallCarbon Nanotubes

    Directory of Open Access Journals (Sweden)

    ZHAO Meng-Li, YUE Yu-Chen, YUAN Li,LI De-Jun, L Xiao-Ying, HUANG Yan

    2010-07-01

    Full Text Available Abstract:Multi―walled nanotubes (MWCNTs) were grown on carbonpapers by chemical vapor deposition (CVD). Adhesion and proliferation of L929 mouse fibroblasts and plateletson MWCNTs and carbon papers were investigated. The influence of different bloodprotein adsorption on the human skin fibroblasts growth was also studied. The resultsshowed that mouse fibroblasts implanted on MWCNTs tended to grow more prolificallythan those on carbon papers. The cell concentration observed on MWCNTsincreased from 1.25105 /mL (cultured for 1 d) to 4.1105/mL (culture for 7 d). No toxicity reaction was observed during the culturingperiod.By employing the adsorptionassay, it was found albumin, fibrinogen, and immunoglobulinG adhered on materials could promotethe adhesion and growth of the human skin fibroblasts. Furthermore, plateletadhesion rate on MWCNTs is lower than that on the carbon papers. These resultsindicate that MWCNTs have better tissue compatibility and blood compatibility.

  20. PED/PEA-15 Controls Fibroblast Motility and Wound Closure by ERK1/2-Dependent Mechanisms

    Science.gov (United States)

    Buonomo, Roberta; Giacco, Ferdinando; Vasaturo, Angela; Caserta, Sergio; Guido, Stefano; Pagliara, Valentina; Garbi, Corrado; Mansueto, Gelsomina; Cassese, Angela; Perruolo, Giuseppe; Oriente, Francesco; Miele, Claudia; Beguinot, Francesco; Formisano, Pietro

    2012-01-01

    Cell migration is dependent on the control of signaling events that play significant roles in creating contractile force and in contributing to wound closure. We evaluated wound closure in fibroblasts from mice overexpressing (TgPED) or lacking ped/pea-15 (KO), a gene overexpressed in patients with type 2 diabetes. Cultured skin fibroblasts isolated from TgPED mice showed a significant reduction in the ability to recolonize wounded area during scratch assay, compared to control fibroblasts. This difference was observed both in the absence and in the presence of mytomicin C, an inhibitor of mitosis. In time-lapse experiments, TgPED fibroblasts displayed about twofold lower velocity and diffusion coefficient, as compared to controls. These changes were accompanied by reduced spreading and decreased formation of stress fibers and focal adhesion plaques. At the molecular level, TgPED fibroblasts displayed decreased RhoA activation and increased abundance of phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2). Inhibition of ERK1/2 activity by PD98059 restored RhoA activation, cytoskeleton organization and cell motility, and almost completely rescued wound closure of TgPED fibroblasts. Interestingly, skin fibroblasts isolated from KO mice displayed an increased wound closure ability. In vivo, healing of dorsal wounds was delayed in TgPED and accelerated in KO mice. Thus, PED/PEA-15 may affect fibroblast motility by a mechanism, at least in part, mediated by ERK1/2. J. Cell. Physiol. 227: 21062116, 2012. 2011 Wiley Periodicals, Inc. PMID:21780113

  1. Dermal fibroblasts in Hutchinson-Gilford progeria syndrome with the lamin A G608G mutation have dysmorphic nuclei and are hypersensitive to heat stress

    Directory of Open Access Journals (Sweden)

    Worman Howard J

    2005-06-01

    Full Text Available Abstract Background Hutchinson-Gilford progeria syndrome (HGPS, OMIM 176670 is a rare sporadic disorder with an incidence of approximately 1 per 8 million live births. The phenotypic appearance consists of short stature, sculptured nose, alopecia, prominent scalp veins, small face, loss of subcutaneous fat, faint mid-facial cyanosis, and dystrophic nails. HGPS is caused by mutations in LMNA, the gene that encodes nuclear lamins A and C. The most common mutation in subjects with HGPS is a de novo single-base pair substitution, G608G (GGC>GGT, within exon 11 of LMNA. This creates an abnormal splice donor site, leading to expression of a truncated protein. Results We studied a new case of a 5 year-old girl with HGPS and found a heterozygous point mutation, G608G, in LMNA. Complementary DNA sequencing of RNA showed that this mutation resulted in the deletion of 50 amino acids in the carboxyl-terminal tail domain of prelamin A. We characterized a primary dermal fibroblast cell line derived from the subject's skin. These cells expressed the mutant protein and exhibited a normal growth rate at early passage in primary culture but showed alterations in nuclear morphology. Expression levels and overall distributions of nuclear lamins and emerin, an integral protein of the inner nuclear membrane, were not dramatically altered. Ultrastructural analysis of the nuclear envelope using electron microscopy showed that chromatin is in close association to the nuclear lamina, even in areas with abnormal nuclear envelope morphology. The fibroblasts were hypersensitive to heat shock, and demonstrated a delayed response to heat stress. Conclusion Dermal fibroblasts from a subject with HGPS expressing a mutant truncated lamin A have dysmorphic nuclei, hypersensitivity to heat shock, and delayed response to heat stress. This suggests that the mutant protein, even when expressed at low levels, causes defective cell stability, which may be responsible for phenotypic abnormalities in the disease.

  2. Fibroblast growth factor 2 can replace ectodermal signaling for feather development.

    OpenAIRE

    Song, H; Wang, Y.; Goetinck, P F

    1996-01-01

    The initiation and morphogenesis of cutaneous appendages depend on a series of reciprocal signaling events between the epithelium and mesenchyme of the embryonic skin. In the development of feather germs, early dermal signals induce the formation of epidermal placodes that in turn signal the mesoderm to form dermal condensations immediately beneath them. We find a spatially and temporally restricted pattern of transcription for the genes that encode fibroblast growth factor (FGF) 2 and FGF re...

  3. Fatty Acid Accumulation and Resulting PPAR? Activation in Fibroblasts due to Trifunctional Protein Deficiency

    OpenAIRE

    Masato Wakabayashi; Yuji Kamijo; Takero Nakajima; Naoki Tanaka; Eiko Sugiyama; Tian Yangyang; Takefumi Kimura; Toshifumi Aoyama

    2012-01-01

    To examine fatty acid accumulation and its toxic effects in cells, we analyzed skin fibroblasts from six patients with mitochondrial trifunctional protein deficiency, who had abnormalities in the second through fourth reactions in fatty acid ?-oxidation system. We found free fatty acid accumulation, enhanced three acyl-CoA dehydrogenases, catalyzing the first reaction in the ?-oxidation system and being assumed to have normal activities in these patients, and PPAR? activation that was confirm...

  4. CCN2: a mechanosignaling sensor modulating integrin-dependent connective tissue remodeling in fibroblasts?

    OpenAIRE

    Leask, Andrew

    2013-01-01

    Tensegrity (tensional integrity) is an emerging concept governing the structure of the body. Integrin-mediated mechanical tension is essential for connective tissue function in vivo. For example, in adult skin fibroblasts, the integrin β1 subunit mediates adhesion to collagen and fibronectin. Moreover, integrin β1, through its abilities to activate latent TGFβ1 and promote collagen production through focal adhesion kinase/rac1/nicotinamide adenine dinucleotide phosphate oxidase (NOX)/reactive...

  5. Collagen Synthesis Is Suppressed in Dermal Fibroblasts by the Human Antimicrobial Peptide LL-37

    OpenAIRE

    Park, Hyun Jeong; Cho, Dae Ho; Kim, Hee Jung; Lee, Jun Young; Cho, Baik Kee; Bang, Sa Ik; Song, Sang Yong; Yamasaki, Kenshi; Nardo, Anna Di; Gallo, Richard L.

    2008-01-01

    LL-37 is a human cathelicidin antimicrobial peptide that is released in the skin after injury and acts to defend against infection and modulate the local cellular immune response. We observed in human dermal keloids that fibrosis was inversely related to the expression of cathelicidin and sought to determine how LL-37 influenced expression of types I and III collagen genes in dermal fibroblasts. At nano-molar concentrations, LL-37 inhibited baseline and transforming growth factor-β-induced co...

  6. Extracellular low pH affects circadian rhythm expression in human primary fibroblasts

    OpenAIRE

    Lee, Sang Kil; Achieng, Elsie; Maddox, Connie; Chen, Suephy C; Iuvone, Michael; Fukuhara, Chiaki

    2011-01-01

    Circadian rhythm is a fundamental biological system involved in the regulation of various physiological functions. However, little is known about a nature or function of circadian clock in human primary cells. In the present study, we have applied in vitro real time circadian rhythm monitoring to study human clock properties using primary skin fibroblasts. Among factors that affect human physiology, slightly lower extracellular pH was chosen to test its effects on circadian rhythm expression....

  7. A genetic defect in the biosynthesis of dermatan sulfate proteoglycan: galactosyltransferase I deficiency in fibroblasts from a patient with a progeroid syndrome.

    OpenAIRE

    Quentin, E; Gladen, A; Rodén, L; Kresse, H.

    1990-01-01

    A small proteoglycan that contains only a single dermatan sulfate chain is the main proteoglycan synthesized by skin fibroblasts. Fibroblasts from a patient with progeroidal appearance and symptoms of the Ehlers-Danlos syndrome have a reduced ability of converting the core protein of this proteoglycan into a mature glycosaminoglycan chain-bearing species. This abnormality is the consequence of a deficiency in galactosyltransferase I (xylosylprotein 4-beta-galactosyltransferase; EC 2.4.1.133),...

  8. Age-Related Changes in the Mechanical Properties of Human Fibroblasts and Its Prospective Reversal After Anti-Wrinkle Tripeptide Treatment

    OpenAIRE

    Dulińska-Molak, Ida; Pasikowska, Monika; Pogoda, Katarzyna; Lewandowska, Małgorzata; Eris, Irena; Lekka, Małgorzata

    2013-01-01

    One of an essential characteristic of human skin are time dependent mechanical properties. Here, we demonstrate that stiffness of human dermal fibroblast correlates with age and it can be restored after anti-wrinkle tripeptide treatment. The stiffness of human fibroblasts isolated from donors of 30-, 40- and 60 years old were examined. Additionally the effect of anti- wrinkle tripeptide of latter cells was investigated. The atomic force microscopy measurements were performed on untreated fibr...

  9. Human lung fibroblasts may modulate dendritic cell phenotype and function: results from a pilot in vitro study.

    Science.gov (United States)

    Freynet, Olivia; Marchal-Sommé, Joëlle; Jean-Louis, Francette; Mailleux, Arnaud; Crestani, Bruno; Soler, Paul; Michel, Laurence

    2016-01-01

    In human lung fibrotic lesions, fibroblasts were shown to be closely associated with immature dendritic cell (DC) accumulation. The aim of the present pilot study was to characterize the role of pulmonary fibroblasts on DC phenotype and function, using co-culture of lung fibroblasts from patients with idiopathic pulmonary fibrosis (IPF) and from control patients, with a DC cell line MUTZ-3. We observed that co-culture of lung control and IPF fibroblasts with DCs reduced the expression of specific DC markers and down-regulated their T-cell stimulatory activity. This suggests that pulmonary fibroblasts might sustain chronic inflammation in the fibrotic lung by maintaining in situ a pool of immature DCs. PMID:27044262

  10. Senescent Fibroblasts Promote Neoplastic Transformation of Partially Transformed Ovarian Epithelial Cells in a Three-dimensional Model of Early Stage Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Kate Lawrenson

    2010-04-01

    Full Text Available Most epithelial ovarian cancers are diagnosed postmenopausally, although the well-established epidemiological risk factors (parity, oral contraceptive use are premenopausal. We hypothesized that accumulation of senescent fibroblasts, together with concomitant loss of presenescent fibroblasts within the ovarian cortex, promotes initiation and early development of ovarian cancer from ovarian surface epithelial (OSE cells. To test this, we established immortalized OSE (IOSE cell lines that mimic early neoplastic transformation by overexpressing the CMYC oncogene (IOSECMYC and normal ovarian presenescent (PSN and senescent (SEN fibroblast cell lines. We then evaluated the ability of PSN and SEN fibroblasts to transform IOSE and IOSECMYC after coculture. SEN fibroblasts significantly enhanced neoplastic development of IOSECMYC cells; there was an up to 15-fold increase in migration of IOSECMYC cells cocultured with SEN fibroblasts compared with PSN fibroblasts. Conditioned medium from SEN fibroblasts promoted anchorage-independent growth of IOSECMYC cells. We studied fibroblast-epithelial cell interactions in heterotypic three-dimensional spheroid models. Dual immunohistochemical staining of spheroids for a proliferation marker (MIB-1 and cytokeratin-18 indicated that SEN fibroblasts induce approximately a five-fold increase in proliferation of IOSECMYC cells relative to cocultures with PSN fibroblasts. SEN, but not PSN fibroblasts, also induced nuclear atypia in epithelial cells in three-dimensional spheroids. These data suggest for the first time that the accumulation of senescent, or loss of presenescent fibroblasts, can promote neoplastic development of partially transformed OSE cells in vitro and illustrates the power of using three-dimensional heterotypic modeling to gain better insights into the etiology underlying the development of epithelial ovarian cancer.

  11. Keratinocytes in tissue engineering of human skin: invitro and in vivo studies

    OpenAIRE

    Fredriksson, Camilla

    2008-01-01

    Full thickness wounds, such as deep burns, need restoration of both the dermal and epidermal layers of the skin. In normal wound healing, re-epithelialization occurs by migration and proliferation of keratinocytes from the wound edges and by differentiation of stem cells from remaining hair follicles. Restoration of dermis occurs by influx of growth factors secreted by macrophages, platelets, and fibroblasts; by fibroblast proliferation and subsequent synthesis and remodeling of collagenous d...

  12. Tissue engineered skin substitutes created by laser-assisted bioprinting form skin-like structures in the dorsal skin fold chamber in mice.

    Science.gov (United States)

    Michael, Stefanie; Sorg, Heiko; Peck, Claas-Tido; Koch, Lothar; Deiwick, Andrea; Chichkov, Boris; Vogt, Peter M; Reimers, Kerstin

    2013-01-01

    Tissue engineering plays an important role in the production of skin equivalents for the therapy of chronic and especially burn wounds. Actually, there exists no (cellularized) skin equivalent which might be able to satisfactorily mimic native skin. Here, we utilized a laser-assisted bioprinting (LaBP) technique to create a fully cellularized skin substitute. The unique feature of LaBP is the possibility to position different cell types in an exact three-dimensional (3D) spatial pattern. For the creation of the skin substitutes, we positioned fibroblasts and keratinocytes on top of a stabilizing matrix (Matriderm). These skin constructs were subsequently tested in vivo, employing the dorsal skin fold chamber in nude mice. The transplants were placed into full-thickness skin wounds and were fully connected to the surrounding tissue when explanted after 11 days. The printed keratinocytes formed a multi-layered epidermis with beginning differentiation and stratum corneum. Proliferation of the keratinocytes was mainly detected in the suprabasal layers. In vitro controls, which were cultivated at the air-liquid-interface, also exhibited proliferative cells, but they were rather located in the whole epidermis. E-cadherin as a hint for adherens junctions and therefore tissue formation could be found in the epidermis in vivo as well as in vitro. In both conditions, the printed fibroblasts partly stayed on top of the underlying Matriderm where they produced collagen, while part of them migrated into the Matriderm. In the mice, some blood vessels could be found to grow from the wound bed and the wound edges in direction of the printed cells. In conclusion, we could show the successful 3D printing of a cell construct via LaBP and the subsequent tissue formation in vivo. These findings represent the prerequisite for the creation of a complex tissue like skin, consisting of different cell types in an intricate 3D pattern. PMID:23469227

  13. Effect of uranium on proliferation and mortality of the major constitutive cell types of the skin: influence on skin barrier integrity

    International Nuclear Information System (INIS)

    The skin is the initial barrier against mechanical, chemical or biological external stresses. It is a complex, multilayered organ. The upper layer is the epidermis that is mainly constituted by keratinocytes. The fibroblast is the major cell type of the dermis which is underlying the epidermis. In the case of an external contamination, uranium is able to diffuse through the skin [1-3] and can affect skin barrier integrity after chronic topical exposure[1, 3]. Our study tried to elucidate the cellular mechanisms leading to this skin alteration after uranyl nitrate contamination. Proliferation rate and mortality of primary cultures of rat skin fibroblasts and keratinocytes contaminated in vitro with different concentration of depleted-uranyl nitrate or 233-uranyl nitrate were measured. The huge difference between 233U and depleted-U specific activities, respectively 3.57 x 108Bq.g-1 and 1.45 x 104Bq.g-1', allowed to distinguish cellular radiotoxicity and chemotoxicity of uranium. Concerning fibroblasts, a significant radiotoxicity of the emitted alpha particles of 233U was observed with no chemotoxicity for the lowest concentrations, i.e. 2?M and 4?M, of uranyl nitrate. Keratinocytes were more sensitive to both uranium radiotoxicity and chemotoxicity than fibroblasts. This can be explained by the about three times higher ability of keratinocytes to incorporate uranium compared to fibroblasts. This greater capacity of epidermal cells than dermal cells to incorporate uranium was confirmed in vivo for the hairless rat following a uranyl nitrate topical contamination. As a conclusion, the important toxic effect of uranium on keratinocyte demonstrated in our study can explain the previous observations [1, 3] that epidermis was atrophied and so skin permeability increased after an in vivo chronic topical exposure of rat skin to uranyl nitrate. These results are of great importance concerning radiation protection of exposed workers and public, and are not taken into consideration by ICRP recommendations at the present time. (orig.)

  14. Wound healing properties of ethyl acetate fraction of Moringa oleifera in normal human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    Sivapragasam Gothai

    2016-03-01

    Full Text Available Background/Aim: Wounds are the outcome of injuries to the skin that interrupt the soft tissue. Healing of a wound is a complex and long-drawn-out process of tissue repair and remodeling in response to injury. A large number of plants are used by folklore traditions for treatment of cuts, wounds and burns. Moringa oleifera is an herb used as traditional folk medicine for the treatment of various skin wounds and associated diseases. The underlying mechanisms of wound healing activity of ethyl acetate fraction of M. oleifera leaves extract are completely unknown. Methods: In the current study, ethyl acetate fraction of Moringa oleifera leaves was investigated for its efficacy on cell viability, proliferation and migration (wound closure rate in human normal dermal fibroblast cells. Results: Results revealed that lower concentration (12.5 and micro;g/ml, 25 and micro;g/ml, and 50 and micro;g/ml of ethyl acetate fraction of M. oleifera leaves showed remarkable proliferative and migratory effect on normal human dermal fibroblasts. Conclusion: The present study suggested that ethyl acetate fraction of M. oleifera leaves might be a potential therapeutic agent for skin wound healing by promoting fibroblast proliferation and migration through increasing the wound closure rate corroborating its traditional use. [J Intercult Ethnopharmacol 2016; 5(1.000: 1-6

  15. Research Techniques Made Simple: Skin Carcinogenesis Models: Xenotransplantation Techniques.

    Science.gov (United States)

    Mollo, Maria Rosaria; Antonini, Dario; Cirillo, Luisa; Missero, Caterina

    2016-02-01

    Xenotransplantation is a widely used technique to test the tumorigenic potential of human cells invivo using immunodeficient mice. Here we describe basic technologies and recent advances in xenotransplantation applied to study squamous cell carcinomas (SCCs) of the skin. SCC cells isolated from tumors can either be cultured to generate a cell line or injected directly into mice. Several immunodeficient mouse models are available for selection based on the experimental design and the type of tumorigenicity assay. Subcutaneous injection is the most widely used technique for xenotransplantation because it involves a simple procedure allowing the use of a large number of cells, although it may not mimic the original tumor environment. SCC cell injections at the epidermal-to-dermal junction or grafting of organotypic cultures containing human stroma have also been used to more closely resemble the tumor environment. Mixing of SCC cells with cancer-associated fibroblasts can allow the study of their interaction and reciprocal influence, which can be followed in real time by intradermal ear injection using conventional fluorescent microscopy. In this article, we will review recent advances in xenotransplantation technologies applied to study behavior of SCC cells and their interaction with the tumor environment invivo. PMID:26802242

  16. Adaptive skin detection based on online training

    Science.gov (United States)

    Zhang, Ming; Tang, Liang; Zhou, Jie; Rong, Gang

    2007-11-01

    Skin is a widely used cue for porn image classification. Most conventional methods are off-line training schemes. They usually use a fixed boundary to segment skin regions in the images and are effective only in restricted conditions: e.g. good lightness and unique human race. This paper presents an adaptive online training scheme for skin detection which can handle these tough cases. In our approach, skin detection is considered as a classification problem on Gaussian mixture model. For each image, human face is detected and the face color is used to establish a primary estimation of skin color distribution. Then an adaptive online training algorithm is used to find the real boundary between skin color and background color in current image. Experimental results on 450 images showed that the proposed method is more robust in general situations than the conventional ones.

  17. Skin Complications of IBD

    Science.gov (United States)

    ... Home > Resources > Skin Complications of IBD Go Back Skin Complications of IBD Email Print + Share After arthritis, ... about 5% of people with inflammatory bowel disease. SKIN DISORDERS COMMONLY SEEN IN IBD ERHTHEMA NODOSUM The ...

  18. Skin care and incontinence

    Science.gov (United States)

    Incontinence - skin care ... or bowels (called incontinence) are at risk of skin problems around the buttocks, hips, genitals, and the ... rectum (perineum). Excess moisture in these areas makes skin problems such as redness, peeling, irritation, and yeast ...

  19. Skin Cancer Foundation

    Science.gov (United States)

    ... Fundraising Event | About Us | Store The Skin Cancer Foundation The Skin Cancer Foundation is the only international organization devoted solely to ... About Us Contact Us © 2016 The Skin Cancer Foundation | 149 Madison Avenue Suite 901 New York, NY ...

  20. Fibroblast Activation Protein Cleaves and Inactivates Fibroblast Growth Factor 21.

    Science.gov (United States)

    Dunshee, Diana Ronai; Bainbridge, Travis W; Kljavin, Noelyn M; Zavala-Solorio, Jose; Schroeder, Amy C; Chan, Ruby; Corpuz, Racquel; Wong, Manda; Zhou, Wei; Deshmukh, Gauri; Ly, Justin; Sutherlin, Daniel P; Ernst, James A; Sonoda, Junichiro

    2016-03-11

    FGF21 is a stress-induced hormone with potent anti-obesity, insulin-sensitizing, and hepatoprotective properties. Although proteolytic cleavage of recombinant human FGF21 in preclinical species has been observed previously, the regulation of endogenously produced FGF21 is not well understood. Here we identify fibroblast activation protein (FAP) as the enzyme that cleaves and inactivates human FGF21. A selective chemical inhibitor, immunodepletion, or genetic deletion of Fap stabilized recombinant human FGF21 in serum. In addition, administration of a selective FAP inhibitor acutely increased circulating intact FGF21 levels in cynomolgus monkeys. On the basis of our findings, we propose selective FAP inhibition as a potential therapeutic approach to increase endogenous FGF21 activity for the treatment of obesity, type 2 diabetes, non-alcoholic steatohepatitis, and related metabolic disorders. PMID:26797127