WorldWideScience

Sample records for infected fibroblasts hsv-fs

  1. Evidence that neomycin inhibits human cytomegalovirus infection of fibroblasts.

    Science.gov (United States)

    Lobert, P E; Hober, D; Delannoy, A S; Wattré, P

    1996-01-01

    The effect of phosphoinositide-binding aminoglycosides, such as neomycin, gentamicin and streptomycin, on human cytomegalovirus (HCMV) infection of human fibroblasts MRC-5 was studied. The inhibition of HCMV infection was obtained with all of these molecules but neomycin was more effective than the others. We showed that the inoculation of the cells with cell-free viral suspension in presence of neomycin concentrations above 5 mM at 37 degrees C, inhibited more than 98% the HCMV infection. However, the preincubation of the fibroblasts with neomycin at 4 degrees C, before the removal of the drug and the inoculation of the cells, induced only a 30% decrease in the number of infected cells. Addition of neomycin after the HCMV-binding at 4 degrees C or the infection of the cells was less efficient to inhibit HCMV infection than the standard incubation of neomycin during inoculation of the fibroblasts. Indeed, 1 hour after the inoculation of the cells at 37 degrees C, neomycin still inhibited HCMV infection, but 4 hours after the inoculation, this drug had no effect on HCMV infection. Our findings demonstrated that neomycin must be present at the time of infection in order to exert a full inhibiting effect. The effect of neomycin on the HCMV infection was almost immediate upon the addition of the drug (binding and/or internalization) and after the virus internalization (inhibition of immediate-early events). We suggest that neomycin and other aminoglycoside antibiotics may interact with HCMV glycoproteins for binding to similar structural features of cell surface heparan sulfate proteoglycans and may inhibit HCMV infection in fibroblasts by disrupting phosphoinositide-mediated events in the cells.

  2. Mucosal stromal fibroblasts markedly enhance HIV infection of CD4+ T cells

    Science.gov (United States)

    Kohgadai, Nargis; Müller, Janis A.; Laustsen, Anders; Thavachelvam, Karthiga; Stürzel, Christina M.; Jones, Jennifer J.; Somsouk, Ma; Garcia, Maurice M.; Smith, James F.; Greenblatt, Ruth M.; Münch, Jan; Jakobsen, Martin R.; Giudice, Linda C.; Greene, Warner C.; Roan, Nadia R.

    2017-01-01

    Understanding early events of HIV transmission within mucosal tissues is vital for developing effective prevention strategies. Here, we report that primary stromal fibroblasts isolated from endometrium, cervix, foreskin, male urethra, and intestines significantly increase HIV infection of CD4+ T cells–by up to 37-fold for R5-tropic HIV and 100-fold for X4-tropic HIV–without themselves becoming infected. Fibroblasts were more efficient than dendritic cells at trans-infection and mediate this response in the absence of the DC-SIGN and Siglec-1 receptors. In comparison, mucosal epithelial cells secrete antivirals and inhibit HIV infection. These data suggest that breaches in the epithelium allow external or luminal HIV to escape an antiviral environment to access the infection-favorable environment of the stromal fibroblasts, and suggest that resident fibroblasts have a central, but previously unrecognized, role in HIV acquisition at mucosal sites. Inhibiting fibroblast-mediated enhancement of HIV infection should be considered as a novel prevention strategy. PMID:28207890

  3. IFN-Dependent and -Independent Reduction in West Nile Virus Infectivity in Human Dermal Fibroblasts

    Directory of Open Access Journals (Sweden)

    Lisa I. Hoover

    2014-03-01

    Full Text Available Although dermal fibroblasts are one of the first cell types exposed to West Nile virus (WNV during a blood meal by an infected mosquito, little is known about WNV replication within this cell type. Here, we demonstrate that neuroinvasive, WNV-New York (WNV-NY, and nonneuroinvasive, WNV-Australia (WNV-AUS60 strains are able to infect and replicate in primary human dermal fibroblasts (HDFs. However, WNV-AUS60 replication and spread within HDFs was reduced compared to that of WNV-NY due to an interferon (IFN-independent reduction in viral infectivity early in infection. Additionally, replication of both strains was constrained late in infection by an IFN-β-dependent reduction in particle infectivity. Overall, our data indicates that human dermal fibroblasts are capable of supporting WNV replication; however, the low infectivity of particles produced from HDFs late in infection suggests that this cell type likely plays a limited role as a viral reservoir in vivo.

  4. In vitro cytotoxic activity of equine lymphocytes on equine herpesvirus-1 infected allogenic fibroblasts

    OpenAIRE

    Edens, Lucy Marie

    1994-01-01

    The objectives of this study were to: 1) develop a technique to analyze the in vitro cytotoxic activity of lymphocytes from adult horses against equine herpes virus-1 (EHV-1) infected allogenic equine dermal fibroblasts (EDF); 2) evaluate the ability of a 72 hour in vitro incubation with interleukin-2 (I L-2) to enhance the lymphocytic cytolytic activity against EHV-1 infected EDF; 3) compare the cytotoxic activity among lymphocytes isolated from pregnant mares and non-pregnant...

  5. Deciphering the differential response of two human fibroblast cell lines following Chikungunya virus infection

    Directory of Open Access Journals (Sweden)

    Thon-Hon Vincent G

    2012-09-01

    Full Text Available Abstract Background Chikungunya virus (CHIKV is an arthritogenic member of the Alphavirus genus (family Togaviridae transmitted by Aedes mosquitoes. CHIKV is now known to target non hematopoietic cells such as epithelial, endothelial cells, fibroblasts and to less extent monocytes/macrophages. The type I interferon (IFN response is an early innate immune mechanism that protects cells against viral infection. Cells express different pattern recognition receptors (including TLR7 and RIG-I to sense viruses and to induce production of type I IFNs which in turn will bind to their receptor. This should result in the phosphorylation and translocation of STAT molecules into the nucleus to promote the transcription of IFN-stimulated antiviral genes (ISGs. We herein tested the capacity of CHIKV clinical isolate to infect two different human fibroblast cell lines HS 633T and HT-1080 and we analyzed the resulting type I IFN innate immune response. Methods Indirect immunofluorescence and quantitative RT-PCR were used to test for the susceptibility of both fibroblast cell lines to CHIKV. Results Interestingly, the two fibroblast cell lines HS 633T and HT-1080 were differently susceptible to CHIKV infection and the former producing at least 30-fold higher viral load at 48 h post-infection (PI. We found that the expression of antiviral genes (RIG-I, IFN-β, ISG54 and ISG56 was more robust in the more susceptible cell line HS 633T at 48 h PI. Moreover, CHIKV was shown to similarly interfere with the nuclear translocation of pSTAT1 in both cell lines. Conclusion Critically, CHIKV can control the IFN response by preventing the nuclear translocation of pSTAT1 in both fibroblast cell lines. Counter-intuitively, the relative resistance of HT-1080 cells to CHIKV infection could not be attributed to more robust innate IFN- and ISG-dependent antiviral responses. These cell lines may prove to be valuable models to screen for novel mechanisms mobilized differentially by

  6. Gene Expression Profiles of Chicken Embryo Fibroblasts in Response to Salmonella Enteritidis Infection.

    Directory of Open Access Journals (Sweden)

    Ama Szmolka

    Full Text Available The response of chicken to non-typhoidal Salmonella infection is becoming well characterised but the role of particular cell types in this response is still far from being understood. Therefore, in this study we characterised the response of chicken embryo fibroblasts (CEFs to infection with two different S. Enteritidis strains by microarray analysis. The expression of chicken genes identified as significantly up- or down-regulated (≥3-fold by microarray analysis was verified by real-time PCR followed by functional classification of the genes and prediction of interactions between the proteins using Gene Ontology and STRING Database. Finally the expression of the newly identified genes was tested in HD11 macrophages and in vivo in chickens. Altogether 19 genes were induced in CEFs after S. Enteritidis infection. Twelve of them were also induced in HD11 macrophages and thirteen in the caecum of orally infected chickens. The majority of these genes were assigned different functions in the immune response, however five of them (LOC101750351, K123, BU460569, MOBKL2C and G0S2 have not been associated with the response of chicken to Salmonella infection so far. K123 and G0S2 were the only 'non-immune' genes inducible by S. Enteritidis in fibroblasts, HD11 macrophages and in the caecum after oral infection. The function of K123 is unknown but G0S2 is involved in lipid metabolism and in β-oxidation of fatty acids in mitochondria.

  7. GROWTH REGULATION IN ROUS SARCOMA VIRUS INFECTED CHICKEN EMBRYO FIBROBLASTS: THE ROLE OF THE src GENE

    Energy Technology Data Exchange (ETDEWEB)

    Parry, G.; Bartholomew, J.A.; Blssell, M.J.

    1980-07-01

    We report here a study of the mechanisms leading to loss of growth control in chicken embryo fibroblasts transformed by Rous sarcoma virus (RSV). We have been particularly concerned with the role of the src gene in this process, and have used RSV mutants temperature sensitive (ts) for transformation to investigate the nature of the growth regulatory lesion. The two principal findings were (1) the stationary phase of the cell cycle (G{sub 1}) in chick embryo fibroblasts seems to have two distinct regulatory compartments (using the terminology of Brooks et al. we refer to these as 'Q' and 'A' states). When rendered stationary at 41.5 C by serum deprivation, normal cells enter a Q state, but cells infected with the ts-mutant occupy an A state. (2) Whereas normal cells can occupy either state depending on culture conditions, the ts-infected cells, at 41.5 C, do not seem to enter Q even though a known src gene product, a kinase, is reported to be inactive at this temperature. We discuss the possibility that viral factors other than the active src protein kinase influence growth control in infected cultures.

  8. Newcastle disease virus infection in chicken embryonic fibroblasts but not duck embryonic fibroblasts is associated with elevated host innate immune response.

    Science.gov (United States)

    Kang, Yinfeng; Feng, Minsha; Zhao, Xiaqiong; Dai, Xu; Xiang, Bin; Gao, Pei; Li, Yulian; Li, Yanling; Ren, Tao

    2016-03-15

    Chickens and ducks are major hosts of Newcastle disease virus (NDV) with distinct responses to infection. However, whereas ducks are generally asymptomatic or exhibit only mild symptoms following NDV infection and are thus regarded as potential long-term reservoirs of the virus, chickens exhibit severe clinical lesions, transient infections and even death due to NDV infection. These differences may in part result from the host innate immune response to NDV infection. To better understand the host innate immune response to NDV infection in avian species, by using the quantitative real-time polymerase chain reaction method we examined the messenger RNA expression levels of immune-related genes in chicken embryonic fibroblasts (CEFs) and duck embryonic fibroblasts (DEFs) when infected with NDV of different pathogenicities. Gene expression profiles showed that the expression of IL-1beta, TNF-α-like factor (LITAF) and interferon (IFN)-beta was upregulated in both CEFs and DEFs infected with SS-10 and NH-10 viruses or treated with polyinosinic:polycytidylic acid [poly(I:C)], as well as that expression levels were greater in CEFs than in DEFs. The expression of TLR3, TLR7, IL-6, IFN-alpha, IFN-gamma, MHC-I and MHC-II, except for IL-8, were also greater in CEFs than in DEFs in response to infection to both viruses or treatment with poly(I:C). However, unlike moderate virulent NH-10, highly virulent SS-10 induced greater pattern recognition receptors and cytokines, except for IFNs, in CEFs and DEFs. Results show distinct expression patterns of cytokines, Toll-like receptors and IFNs associated with inflammatory immune responses to NDV between species and by virulence.

  9. Defensin Production by Human Limbo-Corneal Fibroblasts Infected with Mycobacteria

    Directory of Open Access Journals (Sweden)

    Julieta Luna-Herrera

    2013-02-01

    Full Text Available Epithelial cells of the cornea and the conjunctiva constitutively produce antimicrobial peptides; however, the production of defensins by other cell types located around the eye has not been investigated. We analyzed the production of beta-defensins (hBD and cathelicidin LL-37 during the infection of primary limbo-corneal fibroblasts with M. tuberculosis (MTB, M. abscessus (MAB, and M. smegmatis (MSM. The intracellular survival of each mycobacterium, the production of cytokines and the changes on the distribution of the actin filaments during the infection were also analyzed. Fibroblasts produce basal levels of hBD1 and LL-37 and under PMA stimulation they produce hBD2, hBD3 and overexpress hBD1 and LL-37. MAB induced the highest levels of hBD1 and LL-37 and intermediate levels of IL-6; however, MAB was not eliminated. In addition, MAB induced the greatest change to the distribution of the actin filaments. MTB also produced changes in the structure of the cytoskeleton and induced low levels of hBD1 and IL-6, and intermediate levels of LL-37. The balance of these molecules induced by MTB appeared to contribute to the non-replicative state observed in the limbo-corneal cells. MSM induced the lowest levels of hBD1 and LL-37 but the highest levels of IL-6; MSM was eliminated. The results suggest that mycobacterial infections regulate the production of antimicrobial peptides and cytokines, which in conjunction can contribute to the control of the bacilli.

  10. Canine Distemper Virus Utilizes Different Receptors to Infect Chicken Embryo Fibroblasts and Vero cells

    Institute of Scientific and Technical Information of China (English)

    Jun Chen; Xiu Liang; Pei-fu Chen

    2011-01-01

    Inducing animal viruses to adapt to chicken embryos or chicken embryo fibroblasts(CEF)is a common method to develop attenuated live vaccines with full security.Canine distemper virus(CDV)also does this,but the mechanisms and particular receptors remain unclear.Virus overlay protein blot assays were carried out on CEF membrane proteins,which were extracted respectively with a Mem-PERTM kit,a radioimmunoprecipitation assay buffer or a modified co-immunoprecipitation method,and revealed a common 57 kDa positive band that differed from the 42-kDa positive band in Vero cells and also from those receptors reported in lymphocytes and293 cells,indicating a receptor diversity of CDV and the possibility of the 57-kDa protein acting as a receptor that is involved in adaptive infection of CDV Kunming strain to CEF.

  11. Fibroblast growth factor-2 drives and maintains progressive corneal neovascularization following HSV-1 infection.

    Science.gov (United States)

    Gurung, H R; Carr, M M; Bryant, K; Chucair-Elliott, A J; Carr, D Jj

    2017-04-05

    Herpes simplex virus type 1 (HSV-1) infection of the cornea induces vascular endothelial growth factor A (VEGF-A)-dependent lymphangiogenesis that continues to develop well beyond the resolution of infection. Inflammatory leukocytes infiltrate the cornea and have been implicated to be essential for corneal neovascularization, an important clinically relevant manifestation of stromal keratitis. Here we report that cornea infiltrating leukocytes including neutrophils and T cells do not have a significant role in corneal neovascularization past virus clearance. Antibody-mediated depletion of these cells did not impact lymphatic or blood vessel genesis. Multiple pro-angiogenic factors including IL-6, angiopoietin-2, hepatocyte growth factor, fibroblast growth factor-2 (FGF-2), VEGF-A, and matrix metalloproteinase-9 were expressed within the cornea following virus clearance. A single bolus of dexamethasone at day 10 post infection (pi) resulted in suppression of blood vessel genesis and regression of lymphatic vessels at day 21 pi compared to control-treated mice. Whereas IL-6 neutralization had a modest impact on hemangiogenesis (days 14-21 pi) and lymphangiogenesis (day 21 pi) in a time-dependent fashion, neutralization of FGF-2 had a more pronounced effect on the suppression of neovascularization (blood and lymphatic vessels) in a time-dependent, leukocyte-independent manner. Furthermore, FGF-2 neutralization suppressed the expression of all pro-angiogenic factors measured and preserved visual acuity.Mucosal Immunology advance online publication 5 April 2017; doi:10.1038/mi.2017.26.

  12. Programming of Intestinal Epithelial Differentiation by IL-33 Derived from Pericryptal Fibroblasts in Response to Systemic Infection

    Directory of Open Access Journals (Sweden)

    Mousumi Mahapatro

    2016-05-01

    Full Text Available The intestinal epithelium constitutes an efficient barrier against the microbial flora. Here, we demonstrate an unexpected function of IL-33 as a regulator of epithelial barrier functions. Mice lacking IL-33 showed decreased Paneth cell numbers and lethal systemic infection in response to Salmonella typhimurium. IL-33 was produced upon microbial challenge by a distinct population of pericryptal fibroblasts neighboring the intestinal stem cell niche. IL-33 programmed the differentiation of epithelial progenitors toward secretory IEC including Paneth and goblet cells. Finally, IL-33 suppressed Notch signaling in epithelial cells and induced expression of transcription factors governing differentiation into secretory IEC. In summary, we demonstrate that gut pericryptal fibroblasts release IL-33 to translate bacterial infection into an epithelial response to promote antimicrobial defense.

  13. In vitro effects of new artemisinin derivatives in Neospora caninum-infected human fibroblasts.

    Science.gov (United States)

    Müller, Joachim; Balmer, Vreni; Winzer, Pablo; Rahman, Mahbubur; Manser, Vera; Haynes, Richard K; Hemphill, Andrew

    2015-07-01

    From a panel of 34 artemisinin derivatives tested in vitro, artemisone, GC007 and GC012 were most efficacious at inhibiting Neospora caninum replication (IC50 values of 3-54nM), did not notably impair the invasiveness of tachyzoites and were non-toxic for human foreskin fibroblasts (HFFs). Transmission electron microscopy of drug-treated N. caninum-infected HFFs demonstrated severe alterations in the parasite cytoplasm, changes in the composition of the matrix of the parasitophorous vacuole (PV) and diminished integrity of the PV membrane. To exert parasiticidal activity, parasites had to be cultured continuously in the presence of 5μM artemisone or GC007 for 3 weeks. N. caninum tachyzoites readily adapted to a stepwise increase in concentrations (0.5-10μM) of GC012, but not to artemisone or GC007. Drugs induced the expression of elevated levels of NcBAG1 and NcSAG4 mRNA, but only NcBAG1 could be detected by immunofluorescence. Thus, artemisinin derivatives represent interesting leads that should be investigated further.

  14. Detection of Marek's disease virus serotype 1 (MDV1) glycoprotein D in MDV1-infected chick embryo fibroblasts.

    Science.gov (United States)

    Ono, M; Jang, H K; Maeda, K; Kawaguchi, Y; Tohya, Y; Niikura, M; Mikami, T

    1996-08-01

    Chick embryo fibroblasts (CEFs) infected with three strains of Marek's disease virus serotype 1 (MDV1), GA, Md5 and JM, were subjected to indirect immunofluorescence assay with monoclonal antibodies (MAbs) against MDV1 homolog of glycoprotein D (MDV1 gD) of herpes simplex virus. By the MAbs, a number of MDV1 gD-positive cells were detected in CEFs infected with GA, whereas only a few and no positive cells were detected in CEFs infected with Md5 and JM, respectively. The MDV1 gD in GA-infected CEFs was recognized as the band of 64 kDa in immunoblot analysis using one of the MAbs. This is the first report that the MDV1 gD was detected in MDV1-infected cell cultures.

  15. The morphogenesis of herpes simplex virus type 1 in infected parental mouse L fibroblasts and mutant gro29 cells

    DEFF Research Database (Denmark)

    Jensen, Helle Lone; Norrild, Bodil

    2003-01-01

    Mutants of cell lines and viruses are important biological tools. The pathway of herpesvirus particle maturation and egress are contentious issues. The mutant gro29 line of mouse L cells is defective for egress of herpes simplex virus type 1 (HSV-1) virions, and a candidate for studies of virus......-cell interactions. The properties of uninfected and HSV-1-infected L fibroblasts and gro29 cells investigated by protein assay, immunoblot, titration assay, immunofluorescence light microscopy and immunogold cryosection electron microscopy are reported. The ultrastructure of both HSV-1-infected L and gro29 cells...

  16. Type III interferon gene expression in response to influenza virus infection in chicken and duck embryonic fibroblasts.

    Science.gov (United States)

    Zhang, Zhijie; Zou, Tingting; Hu, Xiaotong; Jin, Hong

    2015-12-01

    Type III interferons (IFN-λs) comprise a group of newly identified antiviral cytokines that are functionally similar to type I IFNs and elicit first-line antiviral responses. Recently, type III IFNs were identified in several species; however, little information is available about type III IFNs in ducks. We compared the expression of type III IFNs and their receptor in chicken embryonic fibroblasts (CEFs) and duck embryonic fibroblasts (DEFs) in response to influenza virus infection. The results showed that the expression of type III IFNs was upregulated in both DEFs and CEFs following infection with H1N1 influenza virus or treatment with poly (I:C), and expression levels were significantly higher in CEFs than in DEFs at each time point. The expression of the receptor for type III IFNs (IL-28Rα) was also upregulated following infection with H1N1 virus or treatment with poly (I:C) and was significantly higher in CEFs than in DEFs at each time point. The expression of the receptor for type III IFNs occurred from 8 hpi and remained at similar levels until 36 hpi in CEFs, but the expression level was elevated from 36 hpi in DEFs. These findings revealed the existence of distinct expression patterns for type III IFNs in chickens and ducks in response to influenza virus infection. The provided data are fundamentally useful in furthering our understanding of type III IFNs and innate antiviral responses in different species.

  17. Multiple mechanisms of N-(phosphonoacetyl)-L-aspartate drug resistance in SV40-infected precrisis human fibroblasts.

    Science.gov (United States)

    Schaefer, D I; Livanos, E M; White, A E; Tlsty, T D

    1993-10-15

    Normal and SV40-infected human fibroblasts were grown in the presence of the drug N-(phosphonoacetyl)-L-aspartate (PALA) and examined for evidence of genetic instability. Both cell populations were precrisis and showed a normal, diploid karyotype at early passage. In contrast to the normal IMR-90 cells, which showed growth arrest and did not form colonies in PALA, the SV40-infected IMR-90 cells formed colonies at a very high frequency and continued to cycle in the drug. The drug-resistant colonies senesced after continued growth in culture, indicating that this change in ability to amplify preceded immortalization. This is the first observation of mortal human cells overcoming the drug-induced growth arrest. Although all previously isolated PALA-resistant colonies demonstrated CAD gene amplification as the mechanism of the drug-resistant phenotype, these SV40-infected human cells also showed alternative mechanisms, including increases in gene copy number by aneuploidy and formation of an isochromosome 2p.

  18. Transcriptional Profiling of Host Gene Expression in Chicken Embryo Fibroblasts Infected with Reticuloendotheliosis Virus Strain HA1101.

    Directory of Open Access Journals (Sweden)

    Ji Miao

    Full Text Available Reticuloendotheliosis virus (REV, a member of the Gammaretrovirus genus in the Retroviridae family, causes an immunosuppressive, oncogenic and runting-stunting syndrome in multiple avian hosts. To better understand the host interactions at the transcriptional level, microarray data analysis was performed in chicken embryo fibroblast cells at 1, 3, 5, and 7 days after infection with REV. This study identified 1,785 differentially expressed genes that were classified into several functional groups including signal transduction, immune response, biological adhesion and endocytosis. Significant differences were mainly observed in the expression of genes involved in the immune response, especially during the later post-infection time points. These results revealed that differentially expressed genes IL6, STAT1, MyD88, TLRs, NF-κB, IRF-7, and ISGs play important roles in the pathogenicity of REV infection. Our study is the first to use microarray analysis to investigate REV, and these findings provide insights into the underlying mechanisms of the host antiviral response and the molecular basis of viral pathogenesis.

  19. Effect of Different Antibiotic Chemotherapies on Pseudomonas aeruginosa Infection In Vitro of Primary Human Corneal Fibroblast Cells

    Directory of Open Access Journals (Sweden)

    Maria del Mar Cendra

    2017-08-01

    Full Text Available Pseudomonas aeruginosa is a major cause of bacterial keratitis (BK worldwide. Inappropriate or non-optimal antibiotic chemotherapy can lead to corneal perforation and rapid sight loss. In this study, we tested the hypothesis that P. aeruginosa strain PAO1 invades primary human corneal fibroblasts (hCFs in vitro and persists intracellularly, despite chemotherapy with antibiotics used commonly to treat BK. In rank order, ciprofloxacin, levofloxacin and polymyxin B showed the highest activity against planktonic PAO1 growth (100% inhibitory concentration ≤10 μg/mL; 50% inhibitory concentration ≤1 μg/mL, followed by gentamicin and ofloxacin (100% inhibitory concentration ≤50 μg/mL; 50% inhibitory concentration ≤10 μg/mL. These bactericidal antibiotics (50–200 μg/mL concentrations all killed PAO1 in the extracellular environment of infected hCF monolayers. By contrast, the bactericidal antibiotic cefuroxime and the bacteriostatic antibiotic chloramphenicol failed to sterilize both PAO1 broth cultures, even at a concentration of ≥200 μg/mL and infected hCF monolayers. Statistically, all antibiotics were able to prevent LDH release from PAO1-infected hCF monolayers at both concentrations tested. Intracellular Pseudomonas were significantly reduced (>99%, P < 0.05 following treatment with ciprofloxacin, levofloxacin and ofloxacin, whereas gentamicin, polymyxin B and cefuroxime failed to clear intracellular bacteria over 24 h. Intracellular Pseudomonas infection was resistant to chloramphenicol, with hCF death observed by 9 h. Eventual growth of remaining intracellular Pseudomonas was observed in hCF after removal of all antibiotics, resulting in re-infection cycles and cell death by 48 h. All of the antibiotics reduced significantly (P < 0.05 IL-1β secretion by hCF infected with a Multiplicity Of Infection (MOI = 1 of PAO1. With higher MOI, no pro-inflammatory effects were observed with antibiotic treatment, expect with polymyxin B and

  20. Phospholipase C-related catalytically inactive protein participates in the autophagic elimination of Staphylococcus aureus infecting mouse embryonic fibroblasts.

    Directory of Open Access Journals (Sweden)

    Kae Harada-Hada

    Full Text Available Autophagy is an intrinsic host defense system that recognizes and eliminates invading bacterial pathogens. We have identified microtubule-associated protein 1 light chain 3 (LC3, a hallmark of autophagy, as a binding partner of phospholipase C-related catalytically inactive protein (PRIP that was originally identified as an inositol trisphosphate-binding protein. Here, we investigated the involvement of PRIP in the autophagic elimination of Staphylococcus aureus in infected mouse embryonic fibroblasts (MEFs. We observed significantly more LC3-positive autophagosome-like vacuoles enclosing an increased number of S. aureus cells in PRIP-deficient MEFs than control MEFs, 3 h and 4.5 h post infection, suggesting that S. aureus proliferates in LC3-positive autophagosome-like vacuoles in PRIP-deficient MEFs. We performed autophagic flux analysis using an mRFP-GFP-tagged LC3 plasmid and found that autophagosome maturation is significantly inhibited in PRIP-deficient MEFs. Furthermore, acidification of autophagosomes was significantly inhibited in PRIP-deficient MEFs compared to the wild-type MEFs, as determined by LysoTracker staining and time-lapse image analysis performed using mRFP-GFP-tagged LC3. Taken together, our data show that PRIP is required for the fusion of S. aureus-containing autophagosome-like vacuoles with lysosomes, indicating that PRIP is a novel modulator in the regulation of the innate immune system in non-professional phagocytic host cells.

  1. Fibroblast growth factor 23: associations with antiretroviral therapy in patients co-infected with HIV and hepatitis C.

    Science.gov (United States)

    Young, J; Mucsi, I; Rollet-Kurhajec, K C; Klein, M B

    2016-05-01

    Fibroblast growth factor 23 (FGF23) has been associated with cardiovascular mortality. We estimate associations between the level of plasma FGF23 and exposure to abacavir (ABC) and to other components of antiretroviral therapy in patients co-infected with HIV and hepatitis C. Both intact and c-terminal FGF23 were measured in plasma using commercial assays for a sub-cohort of 295 patients selected at random from the 1150 patients enrolled in the Canadian Co-infection Cohort. The multiplicative effects of antiretroviral drug exposures and covariates on median FGF23 were then estimated using a hierarchical Bayesian model. The median level of intact FGF23 was independent of either past or recent exposure to abacavir, with multiplicative ratios of 1.00 and 1.07, 95% credible intervals 0.90-1.12 and 0.94-1.23, respectively. Median intact FGF23 tended to increase with past use of both nonnucleoside reverse-transcriptase inhibitors and protease inhibitors, but tended to decrease with recent use of either tenofovir, efavirenz or lopinavir. There were no obvious associations between the median level of c-terminal FGF23 and individual drugs or drug classes. Age, female gender, smoking and the aspartate aminotransferase to platelet ratio index were all associated with a higher median c-terminal FGF23 but not with a higher median intact FGF23. The level of FGF23 in plasma was independent of exposure to ABC. Lower levels of intact FGF23 with recent use of tenofovir, efavirenz or lopinavir may reflect their adverse effects on bone and vitamin D metabolism relative to other drugs in their respective drug classes. © 2015 British HIV Association.

  2. Parapoxvirus orf virus infection induces an increase in interleukin-8, tumour necrosis factor-α, and decorin in goat skin fibroblast cells

    Directory of Open Access Journals (Sweden)

    Wang Lingling

    2016-09-01

    Full Text Available Introduction: Orf virus (ORFV is a prototype Parapoxvirus species in the Poxviridae family that causes serious zoonotic infectious disease. Goat skin fibroblast (GSF cells are the major host targets of ORFV. Interleukin 8 (IL-8 and tumour necrosis factor (TNF-α are known to play a vital role in immune response during viral infections. However, the manner of variation over time of their level of expression in GSF cells remains unclear.

  3. HCV derived from sera of HCV-infected patients induces pro-fibrotic effects in human primary fibroblasts by activating GLI2

    Science.gov (United States)

    Granato, M.; Zompetta, C.; Vescarelli, E.; Rizzello, C.; Cardi, A.; Valia, S.; Antonelli, G.; Marchese, C.; Torrisi, M. R.; Faggioni, A.; Cirone, M.

    2016-01-01

    Hepatitis C virus (HCV) infection is a leading cause of liver fibrosis, especially in developing countries. The process is characterized by the excess accumulation of ECM that may lead, over time, to hepatic cirrhosis, liver failure and also to hepatocarcinoma. The direct role of HCV in promoting fibroblasts trans-differentiation into myofibroblasts, the major fibrogenic cells, has not been fully clarified. In this study, we found that HCV derived from HCV-infected patients infected and directly induced the trans-differentiation of human primary fibroblasts into myofibroblasts, promoting fibrogenesis. This effect correlated with the activation of GLI2, one of the targets of Hedgehog signaling pathway previously reported to be involved in myofibroblast generation. Moreover, GLI2 activation by HCV correlated with a reduction of autophagy in fibroblasts, that may further promoted fibrosis. GLI2 inhibition by Gant 61 counteracted the pro-fibrotic effects and autophagy inhibition mediated by HCV, suggesting that targeting HH/GLI2 pathway might represent a promising strategy to reduce the HCV-induced fibrosis. PMID:27476557

  4. Viral proliferation and expression of tumor-related gene in different chicken embryo fibroblasts infected with different tumorigenic phenotypes of avian leukosis virus subgroup J.

    Science.gov (United States)

    Qu, Yajin; Liu, Litao; Niu, Yujuan; Qu, Yue; Li, Ning; Sun, Wei; Lv, Chuanwei; Wang, Pengfei; Zhang, Guihua; Liu, Sidang

    2016-10-01

    Subgroup J avian leukosis virus (ALV-J) causes a neoplastic disease in infected chickens. The ALV-J strain NX0101, which was isolated from broiler breeders in 2001, mainly induced formation of myeloid cell tumors. However, strain HN10PY01, which was recently isolated from laying hens, mainly induces formation of myeloid cell tumors and hemangioma. To identify the molecular pathological mechanism underlying changes in host susceptibility and tumor classification induced by these two types of ALV-J strains, chicken embryo fibroblasts derived from chickens with different genetic backgrounds (broiler breeders and laying hens) and an immortalized chicken embryo fibroblasts (DF-1) were prepared and infected with strain NX0101 or HN10PY01, respectively. The 50% tissue culture infective dose (TCID50) and levels of ALV group-specific antigen p27 and heat shock protein 70 in the supernatant collected from the ALV-J infected cells were detected. Moreover, mRNA expression levels of tumor-related genes p53, c-myc, and Bcl-2 in ALV-J-infected cells were quantified. The results indicated that the infection of ALV-J could significantly increase mRNA expression levels of p53, c-myc, and Bcl-2 Strain HN10PY01 exhibited a greater influence on the three tumor-related genes in each of the three types of cells when compared with strain NX0101, and the TCID50 and p27 levels in the supernatant collected from HN10PY01-infected cells were higher than those collected from NX0101-infected cells. These results indicate that the infection of the two ALV-J strains influenced the gene expression levels in the infected cells, while the newly isolated strain HN10PY01 showed higher replication ability in cells and induced higher expression levels of tumor-related genes in infected cells. Furthermore, virus titers and expression levels of tumor-related genes and cellular stress responses of cells with different genetic backgrounds when infected with each of the two ALV-J strain were different

  5. Vitamin D3 supplementation increases fibroblast growth factor-23 in HIV-infected youth treated with tenofovir disoproxil fumarate

    Science.gov (United States)

    Havens, Peter L; Hazra, Rohan; Stephensen, Charles B; Kiser, Jennifer J; Flynn, Patricia M; Wilson, Craig M; Rutledge, Brandy; Bethel, James; Pan, Cynthia G; Woodhouse, Leslie R; Van Loan, Marta D; Liu, Nancy; Lujan-Zilbermann, Jorge; Baker, Alyne; Kapogiannis, Bill G; Gordon, Catherine M; Mulligan, Kathleen

    2014-01-01

    Background Tenofovir (TDF) is associated with phosphaturia and elevated 1,25 dihydroxy vitamin D (1,25-OH(2)D). Fibroblast growth factor 23 (FGF23) causes phosphaturia and increases in response to elevated 1,25-OH(2)D. Vitamin D binding protein (VDBP) binds to 1,25-OH(2)D, decreasing its biologic activity, and is elevated in persons with higher plasma tenofovir concentrations. We compared FGF23 and VDBP before and after vitamin D3 (VITD) supplementation in youth treated with combination antiretroviral therapy (cART) containing or not containing TDF. Methods A randomized controlled trial in HIV+ youth ages 18–25 years enrolled participants based on cART treatment with TDF (TDF, N=118) or without TDF (no-TDF, N=85) and randomized within those groups to VITD (50,000 IU every four weeks) or placebo (PL). We measured FGF23 and VDBP and calculated free 1,25-OH(2)D at baseline and week 12, and compared changes by TDF treatment and VITD randomized group. Results At baseline, serum FGF23 concentration showed a quadratic relationship with 1,25-OH(2)D most pronounced in the TDF group. At week 12, total and free 1,25-OH(2)D increased in the VITD but not PL groups, independent of TDF use. FGF23 increased in the TDF group receiving VITD, but there was no FGF23 change in the no-TDF group receiving VITD or the PL groups. The adjusted mean change in FGF23 from baseline to week 12 was +7.7 pg/mL in the TDF/VITD group, compared to −1.7 (no-TDF/VITD, p=0.010); −1.3 (TDF/PL, p=0.006); and +1.1 (no-TDF/PL, p=0.035). Conclusions These results suggest that TDF-containing cART may alter the FGF23 response to vitamin D supplementation in HIV-infected youth. PMID:24535626

  6. Temporal activation of anti- and pro-apoptotic factors in human gingival fibroblasts infected with the periodontal pathogen, Porphyromonas gingivalis: potential role of bacterial proteases in host signalling

    Directory of Open Access Journals (Sweden)

    Takehara Tadamichi

    2006-03-01

    Full Text Available Abstract Background Porphyromonas gingivalis is the foremost oral pathogen of adult periodontitis in humans. However, the mechanisms of bacterial invasion and the resultant destruction of the gingival tissue remain largely undefined. Results We report host-P. gingivalis interactions in primary human gingival fibroblast (HGF cells. Quantitative immunostaining revealed the need for a high multiplicity of infection for optimal infection. Early in infection (2–12 h, P. gingivalis activated the proinflammatory transcription factor NF-kappa B, partly via the PI3 kinase/AKT pathway. This was accompanied by the induction of cellular anti-apoptotic genes, including Bfl-1, Boo, Bcl-XL, Bcl2, Mcl-1, Bcl-w and Survivin. Late in infection (24–36 h the anti-apoptotic genes largely shut down and the pro-apoptotic genes, including Nip3, Hrk, Bak, Bik, Bok, Bax, Bad, Bim and Moap-1, were activated. Apoptosis was characterized by nuclear DNA degradation and activation of caspases-3, -6, -7 and -9 via the intrinsic mitochondrial pathway. Use of inhibitors revealed an anti-apoptotic function of NF-kappa B and PI3 kinase in P. gingivalis-infected HGF cells. Use of a triple protease mutant P. gingivalis lacking three major gingipains (rgpA rgpB kgp suggested a role of some or all these proteases in myriad aspects of bacteria-gingival interaction. Conclusion The pathology of the gingival fibroblast in P. gingivalis infection is affected by a temporal shift from cellular survival response to apoptosis, regulated by a number of anti- and pro-apoptotic molecules. The gingipain group of proteases affects bacteria-host interactions and may directly promote apoptosis by intracellular proteolytic activation of caspase-3.

  7. Isolation of Endoplasmic Reticulum, Mitochondria, and Mitochondria-Associated Membrane and Detergent Resistant Membrane Fractions from Transfected Cells and from Human Cytomegalovirus-Infected Primary Fibroblasts.

    Science.gov (United States)

    Williamson, Chad D; Wong, Daniel S; Bozidis, Petros; Zhang, Aiping; Colberg-Poley, Anamaris M

    2015-09-01

    Increasingly mechanistic virology studies require dependable and sensitive methods for isolating purified organelles containing functional cellular sub-domains. The mitochondrial network is, in part, closely apposed to the endoplasmic reticulum (ER). The mitochondria-associated membrane (MAM) fraction provides direct physical contact between the ER and mitochondria. Characterization of the dual localization and trafficking of human cytomegalovirus (HCMV) UL37 proteins required establishing protocols in which the ER and mitochondria could be reliably separated. Because of its documented role in lipid and ceramide transfer from the ER to mitochondria, a method to purify MAM from infected cells was also developed. Two robust procedures were developed to efficiently isolate mitochondria, ER, and MAM fractions while providing substantial protein yields from HCMV-infected primary fibroblasts and from transfected HeLa cells. Furthermore, this unit includes protocols for isolation of detergent resistant membranes from subcellular fractions as well as techniques that allow visualization of the mitochondrial network disruption that occurs in permissively infected cells by their optimal resolution in Percoll gradients. Copyright © 2015 John Wiley & Sons, Inc.

  8. Sterol Regulatory Element-Binding Protein-1c Regulates Inflammasome Activation in Gingival Fibroblasts Infected with High-Glucose-Treated Porphyromonas gingivalis

    Science.gov (United States)

    Kuo, Hsing-Chun; Chang, Li-Ching; Chen, Te-Chuan; Lee, Ko-Chao; Lee, Kam-Fai; Chen, Cheng-Nan; Yu, Hong-Ren

    2016-01-01

    Background: Porphyromonas gingivalis is a major bacterial species implicated in the progression of periodontal disease, which is recognized as a common complication of diabetes. The interleukin (IL)-1β, processed by the NLR family pyrin domain containing 3 (NLRP3) inflammasome, has been identified as a target for pathogenic infection of the inflammatory response. However, the effect of P. gingivalis in a high-glucose situation in the modulation of inflammasome activation in human gingival fibroblasts (HGFs) is not well-understood. Methods: P. gingivalis strain CCUG25226 was used to study the mechanisms underlying the regulation of HGF NLRP3 expression by the infection of high-glucose-treated P. gingivalis (HGPg). Results: HGF infection with HGPg increases the expression of IL-1β and NLRP3. We further demonstrated that the upregulation of sterol regulatory element-binding protein (SREBP)-1c by activation of the Akt and p70S6K pathways is critical for HGPg-induced NLRP3 expression. We showed that the inhibition of Janus kinase 2 (JAK2) blocks the Akt- and p70S6K-mediated SREBP-1c, NLRP3, and IL-1β expression. The effect of HGPg on HGF signaling and NLRP3 expression is mediated by β1 integrin. In addition, gingival tissues from diabetic patients with periodontal disease exhibited higher NLRP3 and SREBP-1c expression. Conclusions: Our findings identify the molecular pathways underlying HGPg-dependent NLRP3 inflammasome expression in HGFs, providing insight into the effect of P. gingivalis invasion in HGFs. PMID:28083517

  9. 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.

  10. Anti-infective activities of Pelargonium sidoides (EPS® 7630): effects of induced NO production on Leishmania major in infected macrophages and antiviral effects as assessed in a fibroblast-virus protection assay.

    Science.gov (United States)

    Thäle, Carsten; Kiderlen, Albrecht Ferdinand; Kolodziej, Herbert

    2011-05-01

    EPs® 7630 is an aqueous-ethanolic extract of the roots of Pelargonium sidoides, employed in the treatment of upper respiratory tract infections. Its anti-infective activity is supposed to be associated with the activation of the nonspecific immune system. Using Leishmania major GFP-infected murine BMMΦ, the NO production of EPs® 7630-activated macrophages was correlated with the reduction of the GFP signal measured at single cell levels using flow cytometry. The anti-infectious effect of EPs® 7630 (3-10 µg/mL) on its own (NO production: 4-13 µM; signal reduction: 25-73 %) was less prominent than that in combination with IFN- γ (100 U/mL) (NO production: 20-27 µM; signal reduction: 35-78 %). Furthermore, supernatants of EPs® 7630-stimulated BMMΦ (10 µg/mL) significantly reduced the cytopathic effect of EMCV on L929 fibroblasts (antiviral activity 80 U/mL) when compared with an IFN- γ standard (100 U/mL). Direct addition of EPs® 7630 to L929 did not mediate cytoprotective effects. The antiviral components induced in BMMΦ by EPs® 7630 remain to be identified. Detection of any IFNs by ELISA was unsuccessful, which may be due to their very low concentrations in cell supernatants. The current data provide convincing support for the induction of anti-infectious responses by EPs® 7630.

  11. Preventing infection of osseointegrated transcutaneous implants: Incorporation of silver into preconditioned fibronectin-functionalized hydroxyapatite coatings suppresses Staphylococcus aureus colonization while promoting viable fibroblast growth in vitro.

    Science.gov (United States)

    Chimutengwende-Gordon, Mukai; Pendegrass, Catherine; Bayston, Roger; Blunn, Gordon

    2014-09-01

    The success of transcutaneous implants depends on the achievement of a soft tissue seal by enabling fibroblasts to win the race for the surface against bacteria. Fibronectin-functionalized hydroxyapatite coatings (HAFn) have been shown to improve dermal tissue ingrowth and attachment. However, during the early postoperative period before a soft tissue seal has formed, bacterial colonization may occur. This study explored the incorporation of silver, a broad spectrum antimicrobial agent, into HAFn coatings with the aim of reducing bacterial colonization. Silver is known to have dose-dependent cytotoxic effects. Therefore, the effects of silver incorporation into HAFn coatings on both in vitro human dermal fibroblast viability and Staphylococcus aureus colonization were assessed. An electrochemical deposition technique was used to codeposit hydroxyapatite and silver (HAAg) and fibronectin was adsorbed onto this to produce HAAgFn coatings. Surfaces were preconditioned with serum to mimic the in vivo environment. Nonpreconditioned HAAg and HAAgFn coatings suppressed bacterial colonization but were cytotoxic. After serum-preconditioning, more than 90% of fibroblasts that grew on all HAAg and HAAgFn coatings were viable. The highest silver content coatings tested (HAAg100 and HAAgFn100) resulted in a greater than 99% reduction in biofilm and planktonic bacterial numbers compared to HA and HAFn controls. Although HAAg100 had greater antibacterial activity than HAAgFn100, the findings of this study indicate that fibroblasts would win the race for the surface against S aureus on both HAAg100 and HAAgFn100 after serum-preconditioning.

  12. Proteomic analyses of human cytomegalovirus strain AD169 derivatives reveal highly conserved patterns of viral and cellular proteins in infected fibroblasts.

    Science.gov (United States)

    Reyda, Sabine; Büscher, Nicole; Tenzer, Stefan; Plachter, Bodo

    2014-01-07

    Human cytomegalovirus (HCMV) particle morphogenesis in infected cells is an orchestrated process that eventually results in the release of enveloped virions. Proteomic analysis has been employed to reveal the complexity in the protein composition of these extracellular particles. Only limited information is however available regarding the proteome of infected cells preceding the release of HCMV virions. We used quantitative mass spectrometry to address the pattern of viral and cellular proteins in cells, infected with derivatives of the AD169 laboratory strain. Our analyses revealed a remarkable conservation in the patterns of viral and of abundant cellular proteins in cells, infected for 2 hours, 2 days, or 4 days. Most viral proteins increased in abundance as the infection progressed over time. Of the proteins that were reliably detectable by mass spectrometry, only IE1 (pUL123), pTRS1, and pIRS1 were downregulated at 4 days after infection. In addition, little variation of viral proteins in the virions of the different viruses was detectable, independent of the expression of the major tegument protein pp65. Taken together these data suggest that there is little variation in the expression program of viral and cellular proteins in cells infected with related HCMVs, resulting in a conserved pattern of viral proteins ultimately associated with extracellular virions.

  13. In vivo fluorescent labeling of corneal wound healing fibroblasts.

    Science.gov (United States)

    Gatlin, Joel; Melkus, Michael W; Padgett, Angela; Petroll, W Matthew; Cavanagh, H Dwight; Garcia, J Victor; Jester, James V

    2003-03-01

    Numerous studies have shown that fibroblasts play an important role in corneal wound healing, however, the dynamic cellular events underlying wound tissue organization and contraction remain unclear. The purpose of this study was to develop a system to enable live cell imaging of corneal wound healing fibroblasts in situ. To this end, concentrated preparations of an RD114 pseudotyped MLV-based vector expressing the enhanced green fluorescent protein (EGFP) were evaluated in vitro for gene transfer efficiency using cultured rabbit corneal keratocytes. Primary rabbit keratocytes were efficiently labeled in vitro (up to 50% EGFP(+)) at a low multiplicity of infection (MOI=10). To evaluate this gene transfer vector in vivo, rabbit corneal fibroblasts were transduced by direct application of vector supernatant to injured corneas following lamellar keratectomy. Fluorescent fibroblasts were then visualized in situ using epifluorescence microscopy and multiphoton confocal microscopy of excised fresh tissue at multiple time points from 14 days to four months following gene transfer. Fourteen days post-transduction, labeled fibroblasts expressing EGFP were readily detectable by fluorescence microscopy. Detectable fluorescence was noted up to eight weeks post-transduction. Labeled fibroblasts were detected in clusters located predominantly along the margin circumscribing the wound and to a lesser extent within the wound area. Cell growth in clusters was suggestive of the expansion of individual transduced clones. High-resolution imaging showed fluorescent fibroblasts to have a broad, flattened, dendritic morphology, distinct from the spindle shape of cultured fibroblasts. Utilizing multiphoton confocal microscopy, three-dimensional imaging of viable, labeled cells showed wound healing fibroblasts to be extensively interconnected and multi-layered within the corneal wound. These results demonstrate that rabbit corneal fibroblasts can be efficiently transduced in vitro and in

  14. [Dynamics of local expression of connexin-43 and basic fibroblast growth factor receptors in patients with skin and soft-tissue infections against the background of diabetes mellitus type II].

    Science.gov (United States)

    Vinnik, Iu S; Salmina, A B; Tepliakova, O V; Drobushevskaia, A I; Malinovskaia, N A; Pozhilenkova, E A; Morgun, A V; Gitlina, A G

    2014-01-01

    Clinical results of wound healing dynamics were studied in 60 patients with soft-tissue infection against the background of diabetes mellitus type II. At the same time the study considered indices of intercellular contacts protein tissue expression such as connexin 43 (Cx43) and basic fibroblast growth factor receptors (bFGFR). The basic therapy of biopsy material of wound borders was applied. The reduction of bFGFR expression and the minor growth of Cx43 expression were observed. The pain syndrome proceeded for a long time and there were signs of perifocal inflammation, retard wound healing with granulation tissue. The application of combined method of ozone therapy which included autohemotherapy with ozone and an external management of wound by ozone-oxygen mixture facilitated to considerable shortening of inflammatory phase and regeneration. It was associated with increased Cx43 expression (in 1.9 times) in comparison with initial level and bFGFR was enlarged in 1.7 times to eighth day of postoperative period.

  15. 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

  16. Activation of the innate immune response against DENV in normal non-transformed human fibroblasts.

    Directory of Open Access Journals (Sweden)

    José Bustos-Arriaga

    2011-12-01

    Full Text Available BACKGROUND: When mosquitoes infected with DENV are feeding, the proboscis must traverse the epidermis several times ("probing" before reaching a blood vessel in the dermis. During this process, the salivary glands release the virus, which is likely to interact first with cells of the various epidermal and dermal layers, cells which could be physiologically relevant to DENV infection and replication in humans. However, important questions are whether more abundant non-hematopoietic cells such as fibroblasts become infected, and whether they play any role in antiviral innate immunity in the very early stages of infection, or even if they might be used by DENV as primary replication cells. METHODOLOGY/PRINCIPAL FINDINGS: Fibroblasts freshly released from healthy skin and infected 12 hours after their isolation show a positive signal for DENV. In addition, when primary skin fibroblast cultures were established and subsequently infected, we showed DENV-2 antigen-positive intracellular signal at 24 hours and 48 hours post-infection. Moreover, the fibroblasts showed productive infection in a conventional plaque assay. The skin fibroblasts infected with DENV-2 underwent potent signaling through both TLR3 and RIG- 1, but not Mda5, triggering up-regulation of IFNβ, TNFα, defensin 5 (HB5 and β defensin 2 (HβD2. In addition, DENV infected fibroblasts showed increased nuclear translocation of interferon (IFN regulatory factor 3 (IRF3, but not interferon regulatory factor 7 (IRF7, when compared with mock-infected fibroblasts. CONCLUSIONS/SIGNIFICANCE: In this work, we demonstrated the high susceptibility to DENV infection by primary fibroblasts from normal human skin, both in situ and in vitro. Our results suggest that these cells may contribute to the pro-inflammatory and anti-viral microenvironment in the early stages of interaction with DENV-2. Furthermore, the data suggest that fibroblast may also be used as a primary site of DENV replication and

  17. Increased fibroblast functionality on CNN2-loaded titania nanotubes.

    Science.gov (United States)

    Wei, Hongbo; Wu, Shuyi; Feng, Zhihong; Zhou, Wei; Dong, Yan; Wu, Guofeng; Bai, Shizhu; Zhao, Yimin

    2012-01-01

    Infection and epithelial downgrowth are major problems associated with maxillofacial percutaneous implants. These complications are mainly due to the improper closure of the implant-skin interface. Therefore, designing a percutaneous implant that better promotes the formation of a stable soft tissue biologic seal around percutaneous sites is highly desirable. Additionally, the fibroblast has been proven to play an important role in the formation of biologic seals. In this study, titania nanotubes were filled with 11.2 kDa C-terminal CCN2 (connective tissue growth factor) fragment, which could exert full CCN2 activity to increase the biological functionality of fibroblasts. This drug delivery system was fabricated on a titanium implant surface. CCN2 was loaded into anodized titania nanotubes using a simplified lyophilization method and the loading efficiency was approximately 80%. Then, the release kinetics of CCN2 from these nanotubes was investigated. Furthermore, the influence of CCN2-loaded titania nanotubes on fibroblast functionality was examined. The results revealed increased fibroblast adhesion at 0.25, 0.5, 1, 2, 4, and 24 hours, increased fibroblast viability over the course of 5 days, as well as enhanced actin cytoskeleton organization on CCN2-loaded titania nanotubes surfaces compared to uncoated, unmodified counterparts. Therefore, the results from this in vitro study demonstrate that CCN2-loaded titania nanotubes have the ability to increase fibroblast functionality and should be further studied as a method of promoting the formation of a stable soft tissue biologic seal around percutaneous sites.

  18. Infections

    Science.gov (United States)

    ... Does My Child Need? How to Safely Give Acetaminophen Is It a Cold or the Flu? Is the Flu Vaccine a Good Idea for Your Family? Too Late for the Flu Vaccine? Common Childhood Infections Can Chronic Ear Infections Cause Long-Term Hearing Loss? Chickenpox Cold Sores Common Cold Diarrhea Fever and ...

  19. The hallmarks of fibroblast ageing.

    Science.gov (United States)

    Tigges, Julia; Krutmann, Jean; Fritsche, Ellen; Haendeler, Judith; Schaal, Heiner; Fischer, Jens W; Kalfalah, Faiza; Reinke, Hans; Reifenberger, Guido; Stühler, Kai; Ventura, Natascia; Gundermann, Sabrina; Boukamp, Petra; Boege, Fritz

    2014-06-01

    Ageing is influenced by the intrinsic disposition delineating what is maximally possible and extrinsic factors determining how that frame is individually exploited. Intrinsic and extrinsic ageing processes act on the dermis, a post-mitotic skin compartment mainly consisting of extracellular matrix and fibroblasts. Dermal fibroblasts are long-lived cells constantly undergoing damage accumulation and (mal-)adaptation, thus constituting a powerful indicator system for human ageing. Here, we use the systematic of ubiquitous hallmarks of ageing (Lopez-Otin et al., 2013, Cell 153) to categorise the available knowledge regarding dermal fibroblast ageing. We discriminate processes inducible in culture from phenomena apparent in skin biopsies or primary cells from old donors, coming to the following conclusions: (i) Fibroblasts aged in culture exhibit most of the established, ubiquitous hallmarks of ageing. (ii) Not all of these hallmarks have been detected or investigated in fibroblasts aged in situ (in the skin). (iii) Dermal fibroblasts aged in vitro and in vivo exhibit additional features currently not considered ubiquitous hallmarks of ageing. (iv) The ageing process of dermal fibroblasts in their physiological tissue environment has only been partially elucidated, although these cells have been a preferred model of cell ageing in vitro for decades. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  20. Infections

    Directory of Open Access Journals (Sweden)

    Virginia Vanzzini Zago

    2012-01-01

    Full Text Available This is a retrospective, and descriptive study about the support that the laboratory of microbiology aids can provide in the diagnosis of ocular infections in patients whom were attended a tertiary-care hospital in México City in a 10-year-time period. We describe the microbiological diagnosis in palpebral mycose; in keratitis caused by Fusarium, Aspergillus, Candida, and melanized fungi; endophthalmitis; one Histoplasma scleritis and one mucormycosis. Nowadays, ocular fungal infections are more often diagnosed, because there is more clinical suspicion and there are easy laboratory confirmations. Correct diagnosis is important because an early medical treatment gives a better prognosis for visual acuity. In some cases, fungal infections are misdiagnosed and the antifungal treatment is delayed.

  1. Increased fibroblast functionality on CNN2-loaded titania nanotubes

    Directory of Open Access Journals (Sweden)

    Wei HB

    2012-02-01

    Full Text Available Hongbo Wei*, Shuyi Wu*, Zhihong Feng, Wei Zhou, Yan Dong, Guofeng Wu, Shizhu Bai, Yimin Zhao Department of Prosthodontics, School of Stomatology, Fourth Military Medical University, Xi'an, People's Republic of China *These authors contributed equally to this workAbstract: Infection and epithelial downgrowth are major problems associated with maxillofacial percutaneous implants. These complications are mainly due to the improper closure of the implant–skin interface. Therefore, designing a percutaneous implant that better promotes the formation of a stable soft tissue biologic seal around percutaneous sites is highly desirable. Additionally, the fibroblast has been proven to play an important role in the formation of biologic seals. In this study, titania nanotubes were filled with 11.2 kDa C-terminal CCN2 (connective tissue growth factor fragment, which could exert full CCN2 activity to increase the biological functionality of fibroblasts. This drug delivery system was fabricated on a titanium implant surface. CCN2 was loaded into anodized titania nanotubes using a simplified lyophilization method and the loading efficiency was approximately 80%. Then, the release kinetics of CCN2 from these nanotubes was investigated. Furthermore, the influence of CCN2-loaded titania nanotubes on fibroblast functionality was examined. The results revealed increased fibroblast adhesion at 0.25, 0.5, 1, 2, 4, and 24 hours, increased fibroblast viability over the course of 5 days, as well as enhanced actin cytoskeleton organization on CCN2-loaded titania nanotubes surfaces compared to uncoated, unmodified counterparts. Therefore, the results from this in vitro study demonstrate that CCN2-loaded titania nanotubes have the ability to increase fibroblast functionality and should be further studied as a method of promoting the formation of a stable soft tissue biologic seal around percutaneous sites.Keywords: anodization, titania nanotubes, adhesion, connective

  2. Dysregulated proinflammatory and fibrogenic phenotype of fibroblasts in cystic fibrosis.

    Directory of Open Access Journals (Sweden)

    François Huaux

    Full Text Available Morbi-mortality in cystic fibrosis (CF is mainly related to chronic lung infection and inflammation, uncontrolled tissue rearrangements and fibrosis, and yet the underlying mechanisms remain largely unknown. We evaluated inflammatory and fibrosis responses to bleomycin in F508del homozygous and wild-type mice, and phenotype of fibroblasts explanted from mouse lungs and skin. The effect of vardenafil, a cGMP-specific phosphodiesterase type 5 inhibitor, was tested in vivo and in culture. Responses of proinflammatory and fibrotic markers to bleomycin were enhanced in lungs and skin of CF mice and were prevented by treatment with vardenafil. Purified lung and skin fibroblasts from CF mice proliferated and differentiated into myofibroblasts more prominently and displayed higher sensitivity to growth factors than those recovered from wild-type littermates. Under inflammatory stimulation, mRNA and protein expression of proinflammatory mediators were higher in CF than in wild-type fibroblasts, in which CFTR expression reached similar levels to those observed in other non-epithelial cells, such as macrophages. Increased proinflammatory responses in CF fibroblasts were reduced by half with submicromolar concentrations of vardenafil. Proinflammatory and fibrogenic functions of fibroblasts are upregulated in CF and are reduced by vardenafil. This study provides compelling new support for targeting cGMP signaling pathway in CF pharmacotherapy.

  3. Cardiac Fibroblasts Aggravate Viral Myocarditis: Cell Specific Coxsackievirus B3 Replication

    Directory of Open Access Journals (Sweden)

    Diana Lindner

    2014-01-01

    Full Text Available Myocarditis is an inflammatory disease caused by viral infection. Different subpopulations of leukocytes enter the cardiac tissue and lead to severe cardiac inflammation associated with myocyte loss and remodeling. Here, we study possible cell sources for viral replication using three compartments of the heart: fibroblasts, cardiomyocytes, and macrophages. We infected C57BL/6j mice with Coxsackievirus B3 (CVB3 and detected increased gene expression of anti-inflammatory and antiviral cytokines in the heart. Subsequently, we infected cardiac fibroblasts, cardiomyocytes, and macrophages with CVB3. Due to viral infection, the expression of TNF-α, IL-6, MCP-1, and IFN-β was significantly increased in cardiac fibroblasts compared to cardiomyocytes or macrophages. We found that in addition to cardiomyocytes cardiac fibroblasts were infected by CVB3 and displayed a higher virus replication (132-fold increase compared to cardiomyocytes (14-fold increase between 6 and 24 hours after infection. At higher virus concentrations, macrophages are able to reduce the viral copy number. At low virus concentration a persistent virus infection was determined. Therefore, we suggest that cardiac fibroblasts play an important role in the pathology of CVB3-induced myocarditis and are another important contributor of virus replication aggravating myocarditis.

  4. Infection

    Science.gov (United States)

    2010-09-01

    Interactions between biofilms and the environment. FEMS Microbiol Rev. 1997;20:291–303. 4. Webb LX, Wagner W, Carroll D, et al. Osteomyelitis and...treatment of osteomyelitis . Biomed Mater. 2008;3: 034114. 6. Gristina AG. Biomaterial-centered infection: microbial adhesion versus tissue integration...vertebral osteomyelitis . Spine. 2007;32: 2996–3006. 15. Beckham JD, Tuttle K, Tyler KL. Reovirus activates transforming growth factor ß and bone

  5. Direct Conversion of Fibroblasts to Megakaryocyte Progenitors

    Directory of Open Access Journals (Sweden)

    Julian Pulecio

    2016-10-01

    Full Text Available Current sources of platelets for transfusion are insufficient and associated with risk of alloimmunization and blood-borne infection. These limitations could be addressed by the generation of autologous megakaryocytes (MKs derived in vitro from somatic cells with the ability to engraft and differentiate in vivo. Here, we show that overexpression of a defined set of six transcription factors efficiently converts mouse and human fibroblasts into MK-like progenitors. The transdifferentiated cells are CD41+, display polylobulated nuclei, have ploidies higher than 4N, form MK colonies, and give rise to platelets in vitro. Moreover, transplantation of MK-like murine progenitor cells into NSG mice results in successful engraftment and further maturation in vivo. Similar results are obtained using disease-corrected fibroblasts from Fanconi anemia patients. Our results combined demonstrate that functional MK progenitors with clinical potential can be obtained in vitro, circumventing the use of hematopoietic progenitors or pluripotent stem cells.

  6. Connexin43 expression levels influence intercellular coupling and cell proliferation of native murine cardiac fibroblasts.

    Science.gov (United States)

    Zhang, Yan; Kanter, Evelyn M; Laing, James G; Aprhys, Colette; Johns, David C; Kardami, Elissavet; Yamada, Kathryn A

    2008-09-01

    Little is known about connexin expression and function in murine cardiac fibroblasts. The authors isolated native ventricular fibroblasts from adult mice and determined that although they expressed both connexin43 (Cx43) and connexin45 (Cx45), the relative abundance of Cx45 was greater than that of Cx43 in fibroblasts compared to myocytes, and the electrophoretic mobility of both Cx43 and Cx45 differed in fibroblasts and in myocytes. Increasing Cx43 expression by adenoviral infection increased intercellular coupling, whereas decreasing Cx43 expression by genetic ablation decreased coupling. Interestingly, increasing Cx43 expression reduced fibroblast proliferation, whereas decreasing Cx43 expression increased proliferation. These data demonstrate that native fibroblasts isolated from the mouse heart exhibit intercellular coupling via gap junctions containing both Cx43 and Cx45. Fibroblast proliferation is inversely related to the expression level of Cx43. Thus, connexin expression and remodeling is likely to alter fibroblast function, maintenance of the extracellular matrix, and ventricular remodeling in both normal and diseased hearts.

  7. Fibroblastic osteosarcoma of the mandible

    Energy Technology Data Exchange (ETDEWEB)

    Daffner, Richard H. [Department of Diagnostic Radiology, Allegheny General Hospital, Pittsburgh, PA (United States); Fox, Karl R. [Department of Laboratory Medicine, Allegheny General Hospital, Pittsburgh, PA (United States); Galey, Kent R. [Department of Surgery, Division of Oral and Maxillofacial Surgery, Allegheny General Hospital, Pittsburgh, PA (United States)

    2002-02-01

    Osteosarcoma of the mandible is a rare lesion. We report the case of a 16-year-old male who developed a fibroblastic osteosarcoma at the site of a wisdom tooth extraction. The lesion followed an aggressive course of recurrence and diffuse disseminated osteosarcomatosis. We speculate on the causal factors that resulted in this rapid course. (orig.)

  8. [Reconstruction of rabbit corneal stroma with skin fibroblasts].

    Science.gov (United States)

    Zhang, Yan-qing; Zhang, Wen-jie; Liu, Wei; Hu, Xiao-jie; Zhou, Guang-dong; Cui, Lei; Cao, Yi-lin

    2009-09-01

    To explore whether skin fibroblasts could be used as a cell source for reconstruction of the corneal stroma. It was an experimental study. Skin fibroblast cells were isolated from newborn rabbits, cultured and expanded in vitro. Cells were labeled with green fluorescence protein (GFP) gene by retro-viral infection. Fibroblasts at passage 3 were seeded on polyglycolic acid (PGA) non-woven fibers to form a cell-scaffold construct. Constructs were then implanted into the adult rabbit corneal stroma layer after being cultured in vitro for 1 week. Engineered stroma were observed continuously and harvested after 8 weeks of transplantation for gross, histological evaluation and Keratocan examination. PGA alone was used as control. The engineered tissue in the cornea became transparent gradually over a period of 8 weeks. Histological analysis showed that engineered stromal lamellar was relatively regular and the orientation of fibers was parallel to the surface of cornea, which is similar to normal cornea. The implanted cells were confirmed by GFP expression under fluorescent microscope, which also express Keratocan. By transmission electron microscopy examination, no significant difference in the diameter of collagen fiber was observed between engineered stroma (33.08 + or - 2.47) nm and normal stroma (t = 1.80, P = 0.0771). Skin fibroblast cells could be used as seed cells for reconstruction of the corneal stroma.

  9. A fibroblast-associated antigen: Characterization in fibroblasts and immunoreactivity in smooth muscle differentiated stromal cells

    DEFF Research Database (Denmark)

    Rønnov-Jessen, Lone; Celis, Julio E.; van Deurs, Bo

    1992-01-01

    Fibroblasts with smooth muscle differentiation are frequently derived from human breast tissue. Immunofluorescence cytochemistry of a fibroblast-associated antigen recognized by a monoclonal antibody (MAb), 1B10, was analyzed with a view to discriminating smooth muscle differentiated fibroblasts...... from vascular smooth muscle cells. The antigen was detected on the cell surface and in cathepsin D-positive and acridine orange-accumulating vesicular compartments of fibroblasts. Ultrastructurally, the antigen was revealed in coated pits and in endosomal and lysosomal structures. 1B10 recognized three...... immunoreactivity was specific to fibroblasts and smooth muscle differentiated fibroblasts within the context of vascular smooth muscle cells....

  10. Cardiac fibroblast in development and wound healing.

    Science.gov (United States)

    Deb, Arjun; Ubil, Eric

    2014-05-01

    Cardiac fibroblasts are the most abundant cell type in the mammalian heart and comprise approximately two-thirds of the total number of cardiac cell types. During development, epicardial cells undergo epithelial-mesenchymal-transition to generate cardiac fibroblasts that subsequently migrate into the developing myocardium to become resident cardiac fibroblasts. Fibroblasts form a structural scaffold for the attachment of cardiac cell types during development, express growth factors and cytokines and regulate proliferation of embryonic cardiomyocytes. In post natal life, cardiac fibroblasts play a critical role in orchestrating an injury response. Fibroblast activation and proliferation early after cardiac injury are critical for maintaining cardiac integrity and function, while the persistence of fibroblasts long after injury leads to chronic scarring and adverse ventricular remodeling. In this review, we discuss the physiologic function of the fibroblast during cardiac development and wound healing, molecular mediators of activation that could be possible targets for drug development for fibrosis and finally the use of reprogramming technologies for reversing scar. This article is part of a Special Issue entitled "Myocyte-Fibroblast Signalling in Myocardium." Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Gastrointestinal Fibroblasts Have Specialized, Diverse Transcriptional Phenotypes: A Comprehensive Gene Expression Analysis of Human Fibroblasts.

    Directory of Open Access Journals (Sweden)

    Youichi Higuchi

    Full Text Available Fibroblasts are the principal stromal cells that exist in whole organs and play vital roles in many biological processes. Although the functional diversity of fibroblasts has been estimated, a comprehensive analysis of fibroblasts from the whole body has not been performed and their transcriptional diversity has not been sufficiently explored. The aim of this study was to elucidate the transcriptional diversity of human fibroblasts within the whole body.Global gene expression analysis was performed on 63 human primary fibroblasts from 13 organs. Of these, 32 fibroblasts from gastrointestinal organs (gastrointestinal fibroblasts: GIFs were obtained from a pair of 2 anatomical sites: the submucosal layer (submucosal fibroblasts: SMFs and the subperitoneal layer (subperitoneal fibroblasts: SPFs. Using hierarchical clustering analysis, we elucidated identifiable subgroups of fibroblasts and analyzed the transcriptional character of each subgroup.In unsupervised clustering, 2 major clusters that separate GIFs and non-GIFs were observed. Organ- and anatomical site-dependent clusters within GIFs were also observed. The signature genes that discriminated GIFs from non-GIFs, SMFs from SPFs, and the fibroblasts of one organ from another organ consisted of genes associated with transcriptional regulation, signaling ligands, and extracellular matrix remodeling.GIFs are characteristic fibroblasts with specific gene expressions from transcriptional regulation, signaling ligands, and extracellular matrix remodeling related genes. In addition, the anatomical site- and organ-dependent diversity of GIFs was also discovered. These features of GIFs contribute to their specific physiological function and homeostatic maintenance, and create a functional diversity of the gastrointestinal tract.

  12. 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 ...

  13. Interactions between fibroblastic reticular cells and B cells promote mesenteric lymph node lymphangiogenesis.

    Science.gov (United States)

    Dubey, Lalit Kumar; Karempudi, Praneeth; Luther, Sanjiv A; Ludewig, Burkhard; Harris, Nicola L

    2017-08-28

    Lymphatic growth (lymphangiogenesis) within lymph nodes functions to promote dendritic cell entry and effector lymphocyte egress in response to infection or inflammation. Here we demonstrate a crucial role for lymphotoxin-beta receptor (LTβR) signaling to fibroblastic reticular cells (FRCs) by lymphotoxin-expressing B cells in driving mesenteric lymph node lymphangiogenesis following helminth infection. LTβR ligation on fibroblastic reticular cells leads to the production of B-cell-activating factor (BAFF), which synergized with interleukin-4 (IL-4) to promote the production of the lymphangiogenic factors, vascular endothelial growth factors (VEGF)-A and VEGF-C, by B cells. In addition, the BAFF-IL-4 synergy augments expression of lymphotoxin by antigen-activated B cells, promoting further B cell-fibroblastic reticular cell interactions. These results underlie the importance of lymphotoxin-dependent B cell-FRC cross talk in driving the expansion of lymphatic networks that function to promote and maintain immune responsiveness.The growth of lymph nodes in response to infection requires lymphangiogenesis. Dubey et al. show that the mesenteric lymph node lymphangiogenesis upon helminth infection depends on the signaling loop between the B and fibroblastic reticular cells (FRCs), whereby the FRCs respond to lymphotoxin secreted by B cells by releasing B cell activating factor.

  14. Cytotoxicity of selenium nanoparticles in rat dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    Ramos JF

    2012-07-01

    Full Text Available Joseph F Ramos,1 Thomas J Webster21School of Engineering, Center of Biomedical Engineering, 2School of Engineering and Department of Orthopedics, Brown University, Providence, RI, USABackground: Ventilator-associated pneumonia is a deadly nosocomial infection caused by contaminated endotracheal tubes. It has been shown that polyvinyl chloride (PVC, the endotracheal tube substrate coated with elemental selenium nanoparticles reduces bacterial adherence and proliferation on PVC by over 99%. However, it is not known if selenium nanoparticles elicit a cytotoxic effect in vitro. The purpose of this study was to investigate the cytotoxic effects of PVC coated with selenium nanoparticles on fibroblasts, which are mammalian cells central to endotracheal tube intubation.Methods: Different concentrations of selenium nanoparticles were precipitated onto the PVC surface by reduction of selenium salts using glutathione. Characterization of PVC coated with selenium nanoparticles was done by scanning electron microscopy, energy dispersive x-ray, and contact angle measurements. For the cytotoxicity experiments, fibroblasts were seeded at a density of 5000 cm2 onto PVC coated with three different concentrations of selenium nanoparticles (high, medium, low and incubated for 4 hours (adhesion as well as for 24 hours and 72 hours (proliferation. The half-maximal inhibitory concentration (IC50 value was determined after 72 hours using an ultrahigh concentration. MTT assays were used to assess cell viability at the indicated time points.Results: The three concentrations of selenium nanoparticles did not elicit a cytotoxic effect after 72 hours (P < 0.01, n = 3. It was found that the IC50 value was at the ultrahigh concentration of selenium nanoparticles. The nanoparticulate elemental selenium concentration previously shown to decrease the function of bacteria was shown not to cause a cytotoxic effect on fibroblasts in vitro.Conclusion: These findings demonstrate

  15. Aedes Aegypti saliva enhances chikungunya virus replication in human skin fibroblasts via inhibition of the type I interferon signaling pathway.

    Science.gov (United States)

    Wichit, Sineewanlaya; Diop, Fodé; Hamel, Rodolphe; Talignani, Loïc; Ferraris, Pauline; Cornelie, Sylvie; Liegeois, Florian; Thomas, Frédéric; Yssel, Hans; Missé, Dorothée

    2017-09-01

    Chikungunya virus (CHIKV) transmission occurs through the bite of an infected Aedes mosquito which injects virus-containing saliva into the skin of the human host during blood feeding. In the present study, we have determined the effect of Aedes aegypti saliva on CHIKV replication in human skin fibroblasts, a major cell type for viral entry, which mimics the events that occur during natural transmission. A significant increase in the expression of viral transcripts and infectious viral particles was observed in fibroblasts infected with CHIKV in the presence of saliva, as compared with those infected with virus alone. CHIKV-infected human fibroblasts were found to express significantly increased levels of various type I IFN-responsive genes, as demonstrated by specific PCR array analysis. In contrast, the expression of these genes was markedly decreased in cells infected with CHIKV in the presence of mosquito saliva. Moreover, Western blotting analysis revealed that STAT2 and its phosphorylated form were down-regulated in the presence of mosquito saliva. Our data demonstrate for the first time the significance of Aedes aegypti saliva in promoting CHIKV infection via down-regulation of several type I IFN-responsive genes in infected human skin fibroblasts via the JAK-STAT signaling pathway. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Establishing Primary Adult Fibroblast Cultures From Rodents

    Science.gov (United States)

    Seluanov, Andrei; Vaidya, Amita; Gorbunova, Vera

    2010-01-01

    The importance of using primary cells, rather than cancer cell lines, for biological studies is becoming widely recognized. Primary cells are preferred in studies of cell cycle control, apoptosis, and DNA repair, as cancer cells carry mutations in genes involved in these processes. Primary cells cannot be cultured indefinitely due to the onset of replicative senescence or aneuploidization. Hence, new cultures need to be established regularly. The procedure for isolation of rodent embryonic fibroblasts is well established, but isolating adult fibroblast cultures often presents a challenge. Adult rodent fibroblasts isolated from mouse models of human disease may be a preferred control when comparing them to fibroblasts from human patients. Furthermore, adult fibroblasts are the only available material when working with wild rodents where pregnant females cannot be easily obtained. Here we provide a protocol for isolation and culture of adult fibroblasts from rodent skin and lungs. We used this procedure successfully to isolate fibroblasts from over twenty rodent species from laboratory mice and rats to wild rodents such as beaver, porcupine, and squirrel. PMID:20972406

  17. [Heterogeneity of pulmonary fibroblasts in tuberculosis].

    Science.gov (United States)

    Kogan, E A; Sekamova, S M; Bogadel'nikova, I V; Vitukhnovskaia, L A; Fipps, R; Perel'man, M I; Serov, V V

    1997-01-01

    Chronic inflammation and pneumofibrosis are the central events in tuberculosis morphogenesis. It was suggested that a certain type of fibroblasts may play a role in chronization of the inflammation and development of sclerosis in tuberculosis. Fibrous tissue from the foci of secondary tuberculosis (fibrous-cavernous tuberculosis and tuberculomas) of 35 patients were studied light- and electron-microscopically and immunohistochemically. (THY 1-)fibroblasts non-containing lipids and producing insulin-like growth factor 2 (ILGF 2), binding proteins 2 and 4 and epidermal growth factor receptors were found in the foci of secondary tuberculosis close to the granulomatous inflammation and in the new and scarrous fibrous connective tissue of the tuberculoma capsule and caverna walls. These fibroblasts are able for auto- and paracrine regulation of the proliferation of fibroblasts, epithelium and other cells in the inflammatory foci. (THY 1+) fibroblasts containing lipids were observed in the foci of old sclerotic changes among the rough collagen fibres. Thus, (THY 1-) fibroblasts probably play a key role in chronization of inflammation, proliferation and pretumorous dysplasia of pulmonary epithelium in secondary tuberculosis. (THY 1+) fibroblasts containing lipids may show more pronounced collagenesis and may persist under hypoxia condition in the collagenous scars for a long time.

  18. Wound healing and the role of fibroblasts.

    Science.gov (United States)

    Bainbridge, P

    2013-08-01

    Fibroblasts are critical in supporting normal wound healing, involved in key processes such as breaking down the fibrin clot, creating new extra cellular matrix (ECM) and collagen structures to support the other cells associated with effective wound healing, as well as contracting the wound. This article explores and summarises the research evidence on the role of fibroblasts, their origins and activation, and how they navigate the wound bed, as well as how their activity leads to wound contraction. This article also explores the local conditions at the wound site, which activate, regulate and ultimately reduce the fibroblast activity as the skin's integrity returns on healing.

  19. Cigarette Smoke-Exposed Candida albicans Increased Chitin Production and Modulated Human Fibroblast Cell Responses

    Directory of Open Access Journals (Sweden)

    Humidah Alanazi

    2014-01-01

    Full Text Available The predisposition of cigarette smokers for development of respiratory and oral bacterial infections is well documented. Cigarette smoke can also contribute to yeast infection. The aim of this study was to investigate the effect of cigarette smoke condensate (CSC on C. albicans transition, chitin content, and response to environmental stress and to examine the interaction between CSC-pretreated C. albicans and normal human gingival fibroblasts. Following exposure to CSC, C. albicans transition from blastospore to hyphal form increased. CSC-pretreated yeast cells became significantly (P<0.01 sensitive to oxidation but significantly (P<0.01 resistant to both osmotic and heat stress. CSC-pretreated C. albicans expressed high levels of chitin, with 2- to 8-fold recorded under hyphal conditions. CSC-pretreated C. albicans adhered better to the gingival fibroblasts, proliferated almost three times more and adapted into hyphae, while the gingival fibroblasts recorded a significantly (P<0.01 slow growth rate but a significantly higher level of IL-1β when in contact with CSC-pretreated C. albicans. CSC was thus able to modulate both C. albicans transition through the cell wall chitin content and the interaction between C. albicans and normal human gingival fibroblasts. These findings may be relevant to fungal infections in the oral cavity in smokers.

  20. Cigarette Smoke-Exposed Candida albicans Increased Chitin Production and Modulated Human Fibroblast Cell Responses

    Science.gov (United States)

    Alanazi, Humidah; Semlali, Abdelhabib; Perraud, Laura; Chmielewski, Witold; Zakrzewski, Andrew

    2014-01-01

    The predisposition of cigarette smokers for development of respiratory and oral bacterial infections is well documented. Cigarette smoke can also contribute to yeast infection. The aim of this study was to investigate the effect of cigarette smoke condensate (CSC) on C. albicans transition, chitin content, and response to environmental stress and to examine the interaction between CSC-pretreated C. albicans and normal human gingival fibroblasts. Following exposure to CSC, C. albicans transition from blastospore to hyphal form increased. CSC-pretreated yeast cells became significantly (P < 0.01) sensitive to oxidation but significantly (P < 0.01) resistant to both osmotic and heat stress. CSC-pretreated C. albicans expressed high levels of chitin, with 2- to 8-fold recorded under hyphal conditions. CSC-pretreated C. albicans adhered better to the gingival fibroblasts, proliferated almost three times more and adapted into hyphae, while the gingival fibroblasts recorded a significantly (P < 0.01) slow growth rate but a significantly higher level of IL-1β when in contact with CSC-pretreated C. albicans. CSC was thus able to modulate both C. albicans transition through the cell wall chitin content and the interaction between C. albicans and normal human gingival fibroblasts. These findings may be relevant to fungal infections in the oral cavity in smokers. PMID:25302312

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

    DEFF Research Database (Denmark)

    Lepekhin, Eugene; Grøn, Birgitte; Berezin, Vladimir

    2002-01-01

    at these sites can be explained by differences in the motile behavior of their respective fibroblast populations. The migratory characteristics were studied in a two-dimensional culture system. The migration of single cells was time-lapse video recorded at intervals of 15 min for a period of 6 h using a computer...... displacement of periodontal and skin fibroblasts. The decreased cellular displacement of the buccal fibroblasts was found to be due to both lower cellular speed and less persistence in direction. The buccal fibroblasts also displayed smaller areas and longer processes. The differences in cellular morphology...

  2. Measuring the strength of dermal fibroblast attachment to functionalized titanium alloys in vitro.

    Science.gov (United States)

    Pendegrass, Catherine J; Middleton, Claire A; Gordon, David; Jacob, Josh; Blunn, Gordon W

    2010-03-01

    The success of intraosseous transcutaneous amputation prostheses (ITAP) relies on soft tissue attachment to prevent infection which leads to implant failure. Fibronectin (Fn) has been shown to enhance dermal fibroblast attachment in vitro, however measurement of cell attachment strength has been indirect; using cell area and immunolocalization of focal adhesion components. In this study, we have developed a flow apparatus to assess the biophysical strength of cell attachment to biomaterials used in ITAP. We have demonstrated that dermal fibroblast attachment strength increases significantly up to 96 h and that data from direct and indirect methods of assessing cell attachment strength have a significant positive correlation. Additionally, we have used direct and indirect assessment methods to demonstrate that dermal fibroblast attachment strength is significantly greater on fibronectin-coated titanium alloy compared with uncoated controls at 1, 4, and 24 hours.

  3. Fibroblasts in fibrosis: novel roles and mediators

    Directory of Open Access Journals (Sweden)

    Ryan Thomas Kendall

    2014-05-01

    Full Text Available Fibroblasts are the most common cell type of the connective tissues found throughout the body and the principal source of the extensive extracellular matrix (ECM characteristic of these tissues. They are also the central mediators of the pathological fibrotic accumulation of ECM and the cellular proliferation and differentiation that occurs in response to prolonged tissue injury and chronic inflammation. The transformation of the fibroblast cell lineage involves classical developmental signaling programs and includes a surprisingly diverse range of precursor cell types—most notably, myofibroblasts that are the apex of the fibrotic phenotype. Myofibroblasts display exaggerated ECM production; constitutively secrete and are hypersensitive to chemical signals such as cytokines, chemokines, and growth factors; and are endowed with a contractile apparatus allowing them to manipulate the ECM fibers physically to close open wounds. In addition to ECM production, fibroblasts have multiple concomitant biological roles, such as in wound healing, inflammation, and angiogenesis, which are each interwoven with the process of fibrosis. We now recognize many common fibroblast-related features across various physiological and pathological protracted processes. Indeed, a new appreciation has emerged for the role of noncancerous fibroblast interactions with tumors in cancer progression. Although the predominant current clinical treatments of fibrosis involve nonspecific immunosuppressive and anti-proliferative drugs, a variety of potential therapies under investigation specifically target fibroblast biology.

  4. Hematopoietic Origin of Murine Lung Fibroblasts

    Directory of Open Access Journals (Sweden)

    Lindsay T. McDonald

    2015-01-01

    Full Text Available Multiple origins, including the bone marrow, have been suggested to contribute to fibroblast populations in the lung. Using bone marrow reconstitution strategies, the present study tested the hypothesis that the bone marrow hematopoietic stem cell (HSC gives rise to lung tissue fibroblasts in vivo. Data demonstrate that the nonadherent bone marrow fraction is enriched for CD45+ HSC-derived cells and was able to reconstitute hematopoiesis in lethally irradiated animals. Analysis of peripheral blood and lung tissues from engrafted mice demonstrated the ability of this population to give rise to CD45+/Discoidin-Domain Receptor-2+ (DDR2 circulating fibroblast precursors (CFPs in blood and fibroblast populations in lung. An HSC origin for lung fibroblasts was confirmed using a novel clonal cell transplantation method in which the bone marrow is reconstituted by a clonal population derived from a single HSC. Together, these findings provide evidence for an HSC contribution to lung fibroblasts and demonstrate a circulating intermediate through the CD45+/DDR2+ HSC-derived CFP.

  5. A pilot study on indunction and differentiation of human fibroblasts to iPS cells and iPS-RPE cells mediated by retrovirus infection%逆转录病毒感染法对RP患者人体细胞向iPS细胞和iPS-RPE细胞诱导分化的研究

    Institute of Scientific and Technical Information of China (English)

    田媛媛; 蒋超; 陈雪; 丁思加; 徐敏; 赵晨

    2016-01-01

    ,zonula occludens protein 1 (ZO-1) and lecithin retinol acyltransferase (LRAT).Results Cultured human dermal fibroblasts showed fusiform or polygon shape and intercellular close arrangement,and Vimentin was positively expressed in the cells.Small cell colonies were harvested 5-7 days after infected by retroviruses,and the morphology changed from spindle into round mass.The hESC-like iPSCs clonies appeared 20 days after cultivation,and the positive expressions of hESC-specific surface antigens including SSEA3,SSEA4,TRA-1-60,TRA-1-81 and Nanog were found in the cells 25-30 days after cultivation,and the positive staining of AP was obtained 12 weeks after cultivation.A teratoma was formed at the injection site of iPSCs suspension in SCID mouse.Immunofluorescence technique showed that RPE cell-specific proteins including RPE65,ZO-1 and LRAT proteins were positively expressed in iPS-RPE cells at 30 days after differentiation.Conclusions Mutation SNRNP200 p.S1087L of RP patient-specific iPSCs can be induced from human dermal fibroblast by retrovirus infection method.The function and morphology of the iPSCs were similar to hESCs.Human iPSCs cell line generated from the dermal fibroblasts of non-RP individuals can differentiate into iPS-RPE cells.

  6. MicroRNAs Reprogram Normal Fibroblasts into Cancer Associated Fibroblasts in Ovarian Cancer

    Science.gov (United States)

    Mitra, Anirban K.; Zillhardt, Marion; Hua, Youjia; Tiwari, Payal; Murmann, Andrea E.; Peter, Marcus E.; Lengyel, Ernst

    2013-01-01

    Cancer associated fibroblasts (CAFs) are a major constituent of the tumor stroma, but little is known about how cancer cells transform normal fibroblasts into CAFs. miRNAs are small noncoding RNA molecules that negatively regulate gene expression at a posttranscriptional level. While it is clearly established that miRNAs are deregulated in human cancers, it is not known whether miRNA expression in resident fibroblasts is affected by their interaction with cancer cells. We found that in ovarian CAFs, miR-31 and miR-214 are downregulated while miR-155 is upregulated when compared to normal or tumor-adjacent fibroblasts. Mimicking this deregulation by transfecting miRNAs and miRNA inhibitors induced a functional conversion of normal fibroblasts into CAFs, and the reverse experiment resulted in the reversion of CAFs into normal fibroblasts. The miRNA-reprogrammed normal fibroblasts and patient-derived CAFs shared a large number of upregulated genes highly enriched in chemokines, which are known to be important for CAF function. The most highly upregulated chemokine, CCL5, was found to be a direct target of miR-214. These results indicate that ovarian cancer cells reprogram fibroblasts to become CAFs through the action of miRNAs. Targeting these miRNAs in stromal cells could have therapeutic benefit. PMID:23171795

  7. Effect of lipopolysaccharide on glucocorticoid receptor function in control nasal mucosa fibroblasts and in fibroblasts from patients with chronic rhinosinusitis with nasal polyps and asthma.

    Directory of Open Access Journals (Sweden)

    Laura Fernández-Bertolín

    Full Text Available Chronic rhinosinusitis with nasal polyps (CRSwNP is a chronic inflammatory disease of the upper airways frequently associated with asthma. Bacterial infection is a feature of CRSwNP that can aggravate the disease and the response to glucocorticoid treatment.We examined whether the bacterial product lipopolysaccharide (LPS reduces glucocorticoid receptor (GR function in control nasal mucosa (NM fibroblasts and in nasal polyp (NP fibroblasts from patients with CRSwNP and asthma.NP (n = 12 and NM fibroblasts (n = 10 were in vitro pre-incubated with LPS (24 hours prior to the addition of dexamethasone. Cytokine/chemokine secretion was measured by ELISA and Cytometric Bead Array. GRα, GRβ, mitogen-activated protein-kinase phosphatase-1 (MKP-1 and glucocorticoid-induced leucine zipper (GILZ expression was measured by RT-PCR and immunoblotting, GRα nuclear translocation by immunocytochemistry, and GRβ localization by immunoblotting. The role of MKP-1 and GILZ on dexamethasone-mediated cytokine inhibition was analyzed by small interfering RNA silencing.Pre-incubation of nasal fibroblasts with LPS enhanced the secretion of IL-6, CXCL8, RANTES, and GM-CSF induced by FBS. FBS-induced CXCL8 secretion was higher in NP than in NM fibroblasts. LPS effects on IL-6 and CXCL8 were mediated via activation of p38α/β MAPK and IKK/NF-κB pathways. Additionally, LPS pre-incubation: 1 reduced dexamethasone's capacity to inhibit FBS-induced IL-6, CXCL8 and RANTES, 2 reduced dexamethasone-induced GRα nuclear translocation (only in NM fibroblasts, 3 did not alter GRα/GRβ expression, 4 decreased GILZ expression, and 5 did not affect dexamethasone's capacity to induce MKP-1 and GILZ expression. MKP-1 knockdown reduced dexamethasone's capacity to suppress FBS-induced CXCL8 release.The bacterial product LPS negatively affects GR function in control NM and NP fibroblasts by interfering with the capacity of the activated receptor to inhibit the production of pro

  8. Corneal Fibroblasts as Sentinel Cells and Local Immune Modulators in Infectious Keratitis

    Directory of Open Access Journals (Sweden)

    Ken Fukuda

    2017-08-01

    Full Text Available The cornea serves as a barrier to protect the eye against external insults including microbial pathogens and antigens. Bacterial infection of the cornea often results in corneal melting and scarring that can lead to severe visual impairment. Not only live bacteria but also their components such as lipopolysaccharide (LPS of Gram-negative bacteria contribute to the development of inflammation and subsequent corneal damage in infectious keratitis. We describe the important role played by corneal stromal fibroblasts (activated keratocytes as sentinel cells, immune modulators, and effector cells in infectious keratitis. Corneal fibroblasts sense bacterial infection through Toll-like receptor (TLR–mediated detection of a complex of LPS with soluble cluster of differentiation 14 (CD14 and LPS binding protein present in tear fluid. The cells then initiate innate immune responses including the expression of chemokines and adhesion molecules that promote the recruitment of inflammatory cells necessary for elimination of the infecting bacteria. Infiltrated neutrophils are activated by corneal stromal collagen and release mediators that stimulate the production of pro–matrix metalloproteinases by corneal fibroblasts. Elastase produced by Pseudomonas aeruginosa (P. aeruginosa activates these released metalloproteinases, resulting in the degradation of stromal collagen. The modulation of corneal fibroblast activation and of the interaction of these cells with inflammatory cells and bacteria is thus important to minimize corneal scarring during treatment of infectious keratitis. Pharmacological agents that are able to restrain such activities of corneal fibroblasts without allowing bacterial growth represent a potential novel treatment option for prevention of excessive scarring and tissue destruction in the cornea.

  9. N-cadherin is overexpressed in Crohn's stricture fibroblasts and promotes intestinal fibroblast migration.

    LENUS (Irish Health Repository)

    Burke, John P

    2012-02-01

    BACKGROUND: Intestinal fibroblasts mediate stricture formation in Crohn\\'s disease (CD). Transforming growth factor-beta (TGF-beta) is important in fibroblast activation, while cell attachment and migration is regulated by the adhesion molecule N-cadherin. The aim of this study was to investigate the expression and function of N-cadherin in intestinal fibroblasts in patients with fibrostenosing CD. METHODS: Intestinal fibroblasts were cultured from seromuscular biopsies from patients undergoing resection for terminal ileal fibrostenosing CD (n = 14) or controls patients (n = 8). N-cadherin expression was assessed using Western blot and quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Fibroblasts were stimulated with TGF-beta and selective pathway inhibitors Y27632, PD98050, and LY294002 were used to examine the Rho\\/ROCK, ERK-1\\/2, and Akt signaling pathways, respectively. Cell migration was assessed using a scratch wound assay. N-cadherin was selectively overexpressed using a plasmid. RESULTS: Fibroblasts from fibrostenosing CD express increased constitutive N-cadherin mRNA and protein and exhibit enhanced basal cell migration relative to those from directly adjacent normal bowel. Control fibroblasts treated with TGF-beta induced N-cadherin in a dose-dependent manner which was inhibited by Rho\\/ROCK and Akt pathway modulation. Control fibroblasts exhibited enhanced cell migration in response to treatment with TGF-beta or transfection with an N-cadherin plasmid. CONCLUSIONS: Fibroblasts from strictures in CD express increased constitutive N-cadherin and exhibit enhanced basal cell migration. TGF-beta is a potent inducer of N-cadherin in intestinal fibroblasts resulting in enhanced cell migration. The TGF-beta-mediated induction of N-cadherin may potentiate Crohn\\'s stricture formation.

  10. MicroRNAs reprogram normal fibroblasts into cancer-associated fibroblasts in ovarian cancer.

    Science.gov (United States)

    Mitra, Anirban K; Zillhardt, Marion; Hua, Youjia; Tiwari, Payal; Murmann, Andrea E; Peter, Marcus E; Lengyel, Ernst

    2012-12-01

    Cancer-associated fibroblasts (CAF) are a major constituent of the tumor stroma, but little is known about how cancer cells transform normal fibroblasts into CAFs. microRNAs (miRNA) are small noncoding RNA molecules that negatively regulate gene expression at a posttranscriptional level. Although it is clearly established that miRNAs are deregulated in human cancers, it is not known whether miRNA expression in resident fibroblasts is affected by their interaction with cancer cells. We found that in ovarian CAFs, miR-31 and miR-214 were downregulated, whereas miR-155 was upregulated when compared with normal or tumor-adjacent fibroblasts. Mimicking this deregulation by transfecting miRNAs and miRNA inhibitors induced a functional conversion of normal fibroblasts into CAFs, and the reverse experiment resulted in the reversion of CAFs into normal fibroblasts. The miRNA-reprogrammed normal fibroblasts and patient-derived CAFs shared a large number of upregulated genes highly enriched in chemokines, which are known to be important for CAF function. The most highly upregulated chemokine, CCL5, (C-C motif ligand 5) was found to be a direct target of miR-214. These results indicate that ovarian cancer cells reprogram fibroblasts to become CAFs through the action of miRNAs. Targeting these miRNAs in stromal cells could have therapeutic benefit. The mechanism by which quiescent fibroblasts are converted into CAFs is unclear. The present study identifies a set of 3 miRNAs that reprogram normal fibroblasts to CAFs. These miRNAs may represent novel therapeutic targets in the tumor microenvironment. ©2012 AACR.

  11. Molecular mechanism of fluoroquinolones modulation on corneal fibroblast motility.

    Science.gov (United States)

    Chen, Tsan-Chi; Tsai, Tzu-Yun; Chang, Shu-Wen

    2016-04-01

    Topical fluoroquinolones are widely used to prevent ocular infections after ophthalmic surgery. However, they have been shown to affect the corneal cell motility, whose mechanism remains indefinite. The purpose of this study was to investigate how fluoroquinolones affect corneal stromal cell motility. Human corneal fibroblasts (HCFs) were incubated in ciprofloxacin (CIP), levofloxacin (LEV), or moxifloxacin (MOX) at 0, 10, 50, and 100 μg/ml for up to 3 days. Effect of CIP, LEV, or MOX on HCF migration was monitored using migration assay. HCF viability was determined by WST-1 assay. Expression of focal adhesion kinase (FAK), paxillin (PXN), and their phosphorylated forms were analyzed by immunoblotting. Binding affinity between FAK and PXN was determined by co-immunoprecipitation. Our results revealed that CIP and MOX, but not LEV, noticeably retarded HCF migration. HCF proliferation was significantly reduced by CIP (38.2%), LEV (29.5%), and MOX (21.3%), respectively (p = 0.002). CIP and MOX suppressed the phosphorylation of PXN at tyrosines (10.2 ± 4.3%, p MOX diminished the binding affinity between FAK and PXN (8.2 ± 1.8%, p MOX, but not LEV, might delay corneal fibroblast migration via interfering with recruitment of PXN to focal adhesions and dephosphorylation of PXN at the tyrosines.

  12. Human cytomegalovirus induces alteration of (-actin mRNA and microfilaments in human embryo fibroblast cells

    Institute of Scientific and Technical Information of China (English)

    林茂芳; 魏国庆; 黄河; 蔡真

    2004-01-01

    Objective: To investigate the infection of human embryo fibroblast cell line HF cells by CMV as well as the effects of CMV on β-actin mRNA and microfilaments. Methods: HF cells shape was observed after the infection of CMV. RT-PCR assay was used to detect the mRNA expression of CMV immediate early (IE) gene, β-actin and GAPDH genes of HF cells infected by CMV. CMV particles and cell microfilaments were detected with electron microscope. Results: Shape of HF cell changed after the infection by CMV. HF cells infected by CMV could express IE mRNA and the expression of β-actin mRNA decreased in a time- and titer-dependent manner compared with the uninfected HF cells whose expression of GAPDH mRNA did not change much. CMV particles were found with electron microscope in the cells. Microfilaments were ruptured and shortened after the infection of CMV. Conclusion: CMV can not only infect human embryo fibroblast cells line HF cells and replicate in the cells, but can also affect the expression of β-actin mRNA and the microfilaments.

  13. Fetal intestinal fibroblasts respond to insulin-like growth factor (IGF-II better than adult intestinal fibroblasts

    Directory of Open Access Journals (Sweden)

    Fillenwarth Michael J

    2006-01-01

    Full Text Available Abstract Background We compared IGF responses of fetal and adult intestinal fibroblasts to identify a developmental difference in the IGF-axis. Intestinal fibroblasts were isolated from maternal and fetal jejunum. Media was conditioned at confluence and one week afterwards. The proliferative response at confluence to 5 nM IGF-I or -II was compared. Results There were no significant differences in IGFBP expression at confluence. Post-confluence, fetal fibroblasts had no significant changes in IGFBP-2 and IGFBP-3 expression. Post-confluent maternal fibroblasts had increased IGFBP-3 levels that were significant compared to the fetal fibroblasts. IGF-I increased in post-confluent fetal fibroblasts, while in maternal fibroblasts it decreased (p Conclusion Fetal intestinal fibroblasts respond to IGF-II with greater proliferation and do not have the increased IGFBPs seen post-confluence in adult intestinal fibroblasts.

  14. FIBROBLAST INVOLVEMENT IN SOFT CONNECTIVE TISSUE CALCIFICATION

    Directory of Open Access Journals (Sweden)

    Ivonne eRonchetti

    2013-03-01

    Full Text Available Soft connective tissue calcification is not a passive process, but the consequence of metabolic changes of local mesenchymal cells that, depending on both genetic and environmental factors, alter the balance between pro- and anti-calcifying pathways. While the role of smooth muscle cells and pericytes in ectopic calcifications has been widely investigated, the involvement of fibroblasts is still elusive. Fibroblasts isolated from the dermis of PXE patients and of patients exhibiting PXE-like clinical and histopathological findings offer an attractive model to investigate the mechanisms leading to the precipitation of mineral deposits within elastic fibres and to explore the influence of the genetic background and of the extracellular environment on fibroblast-associated calcifications, thus improving the knowledge on the role of mesenchymal cells on pathologic mineralization.

  15. Chikungunya virus exploits miR-146a to regulate NF-κB pathway in human synovial fibroblasts.

    Directory of Open Access Journals (Sweden)

    Sakthi Priya Selvamani

    Full Text Available OBJECTIVES: Chikungunya virus causes chronic infection with manifestations of joint pain. Human synovial fibroblasts get infected with CHIKV and could lead to pro-inflammatory responses. MicroRNAs have potentials to regulate the gene expression of various anti-viral and pro-inflammatory genes. The study aims to investigate the role of miR-146a in modulation of inflammatory responses of human synovial fibroblasts by Chikungunya virus. METHODS: To study the role of miR-146a in CHIKV pathogenesis in human synovial cells and underlying inflammatory manifestations, we performed CHIKV infection in primary human synovial fibroblasts. Western blotting, real-time PCR, luciferase reporter assay, overexpression and knockdown of cellular miR-146a strategies have been employed to validate the role of miR-146a in regulation of pro-inflammatory NF-κB pathway. RESULTS: CHIKV infection induced the expression of cellular miR-146a, which resulted into down-regulation of TRAF6, IRAK1, IRAK2 and increased replication of CHIKV in human synovial fibroblasts. Exogenous expression of miR-146a in human synovial fibroblasts led to decreased expression of TRAF6, IRAK1, IRAK2 and decreased replication of CHIKV. Inhibition of cellular miR-146a by anti-miR-146a restored the expression levels of TRAF6, IRAK1 and IRAK2. Downregulation of TRAF6, IRAK1 and IRAK2 led to downstream decreased NF-κB activation through negative feedback loop. CONCLUSION: This study demonstrated the mechanism of exploitation of cellular miR-146a by CHIKV in modulating the host antiviral immune response in primary human synovial fibroblasts.

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

    Energy Technology Data Exchange (ETDEWEB)

    Schweikert, H.U.; Hein, H.J.; Romijn, J.C.; Schroeder, F.H.

    1982-02-01

    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.

  17. Myocardial fibroblast-matrix interactions and potential therapeutic targets.

    Science.gov (United States)

    Goldsmith, Edie C; Bradshaw, Amy D; Zile, Michael R; Spinale, Francis G

    2014-05-01

    The cardiac extracellular matrix (ECM) is a dynamic structure, adapting to physiological and pathological stresses placed on the myocardium. Deposition and organization of the matrix fall under the purview of cardiac fibroblasts. While often overlooked compared to myocytes, fibroblasts play a critical role in maintaining ECM homeostasis under normal conditions and in response to pathological stimuli assume an activated, myofibroblast phenotype associated with excessive collagen accumulation contributing to impaired cardiac function. Complete appreciation of fibroblast function is hampered by the lack of fibroblast-specific reagents and the heterogeneity of fibroblast precursors. This is further complicated by our ability to dissect the role of myofibroblasts versus fibroblasts in myocardial in remodeling. This review highlights critical points in the regulation of collagen deposition by fibroblasts, the current panel of molecular tools used to identify fibroblasts and the role of fibroblast-matrix interactions in fibroblast function and differentiation into the myofibroblast phenotype. The clinical potential of exploiting differences between fibroblasts and myofibroblasts and using them to target specific fibroblast populations is also discussed. This article is part of a Special Issue entitled "Myocyte-Fibroblast Signalling in Myocardium." Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Zinc and propolis reduces cytotoxicity and proliferation in skin fibroblast cell culture: total polyphenol content and antioxidant capacity of propolis.

    Science.gov (United States)

    Tyszka-Czochara, Małgorzata; Paśko, Paweł; Reczyński, Witold; Szlósarczyk, Marek; Bystrowska, Beata; Opoka, Włodzimierz

    2014-07-01

    It has been demonstrated that zinc exerts its beneficial influence on skin fibroblasts. Propolis, a complex mixture of plant-derived and bees' products, was reported to stimulate cicatrization processes in skin and prevent infections. The aim of this study was to find out how zinc and propolis influence human skin fibroblasts in cell culture and to compare the effect of individual compounds to the effect of a mixture of zinc and propolis. In this study, zinc, as zinc aspartate, at a concentration of 16 μM, increased human fibroblasts proliferation in cell culture, whereas propolis at a concentration of 0.01% (w/v) revealed antiproliferative and cytotoxic action followed by mild cell necrosis. In culture, zinc was effectively transported into fibroblasts, and propolis inhibited the amount of zinc incorporated into the cells. An addition of propolis to the medium caused a decrease in the Zn(II) amount incorporated into fibroblasts. The obtained results also indicate an appreciable antioxidant property of propolis and revealed its potential as a supplement when applied at doses lower than 0.01% (w/v). In conclusion, the present study showed that zinc had a protective effect on human cultured fibroblasts' viability, although propolis revealed its antiproliferative action and caused mild necrosis.

  19. Avidin is induced in chicken embryo fibroblasts by viral transformation and cell damage.

    OpenAIRE

    Korpela, J; Kulomaa, M; Tuohimaa, P; Vaheri, A.

    1983-01-01

    Synthesis and secretion of avidin was studied in cultured chicken embryo fibroblasts infected with transforming retroviruses (Rous sarcoma virus, its mutants temperature-sensitive for transformation, OK-10 virus) or a nontransforming retrovirus (RAV-1). Avidin was detectable in both transformed and untransformed cultures, and was identical to chicken egg white avidin by several criteria: biotin-binding, heat-induced biotin exchange, subunit size (mol. wt. 15 600), immunoprecipitation of metab...

  20. [The effect of alloxan on the multiplication of different types of normal or cancerous cells, and on the transformation of chick embryo fibroblasts by Rous virus in vitro].

    Science.gov (United States)

    Grobon, P; Latarjet, R

    1975-05-26

    The action of alloxan was studied in vitro, on different categories of normal or cancerous cells. At concentrations of 250 and 350 gamma/ml, alloxan does not significantly inhibit the growth of normal cells, whereas it does inhibit the growth of cancerous cells. Furthermore, alloxan inhibits infection and transformation of chicken-embryo fibroblasts infected by the Rous virus.

  1. Extracellular Matrix and Dermal Fibroblast Function in the Healing Wound

    OpenAIRE

    Tracy, Lauren E.; Minasian, Raquel A.; Caterson, E.J.

    2016-01-01

    Significance: Fibroblasts play a critical role in normal wound healing. Various extracellular matrix (ECM) components, including collagens, fibrin, fibronectin, proteoglycans, glycosaminoglycans, and matricellular proteins, can be considered potent protagonists of fibroblast survival, migration, and metabolism.

  2. Transcriptomic profiles of human foreskin fibroblast cells in response to orf virus.

    Science.gov (United States)

    Chen, Daxiang; Long, Mingjian; Xiao, Bin; Xiong, Yufeng; Chen, Huiqin; Chen, Yu; Kuang, Zhenzhan; Li, Ming; Wu, Yingsong; Rock, Daniel L; Gong, Daoyuan; Wang, Yong; He, Haijian; Liu, Fang; Luo, Shuhong; Hao, Wenbo

    2017-08-29

    Orf virus has been utilized as a safe and efficient viral vector against not only diverse infectious diseases, but also against tumors. However, the nature of the genes triggered by the vector in human cells is poorly characterized. Using RNA sequencing technology, we compared specific changes in the transcriptomic profiles in human foreskin fibroblast cells following infection by the orf virus. The results indicated that orf virus upregulates or downregulates expression of a variety of genes, including genes involved in antiviral immune response, apoptosis, cell cycle and a series of signaling pathways, such as the IFN and p53-signaling pathways. The orf virus stimulates or inhibits immune gene expression such as chemokines, chemokine receptors, cytokines, cytokine receptors, and molecules involved in antigen uptake and processing after infection. Expression of pro-apoptotic genes increased at 8 hours post-infection. The p53 signaling pathway was activated to induce apoptosis at the same time. However, the cell cycle program was promoted after infection, which may be due to the immunomodulatory genes of the orf virus. This presents the first description of transcription profile changes in human foreskin fibroblast cells after orf virus infection and provides an in-depth analysis of the interaction between the host and orf virus. These data offer new insights into the understanding of the mechanisms of infection by orf virus and identify potential targets for future studies.

  3. Efficacy of Nano Germicidal Light Therapy on Wound Related Infections

    Science.gov (United States)

    2016-09-12

    wound healing through leukocytes and stimulation of fibroblasts and epithelial cells. Specifically, the cytokines include platelet derived growth...infection and to serve as a central component releasing cytokine of wound healing process.29 During the proliferative phase, cells such as fibroblasts ... healing : Part II. Role in normal and chronic wound healing . Am J Surg. 1993;166(1):74-81. 29. Bowler PG. Wound pathophysiology, infection and

  4. Fibroblast growth factors in neurodevelopment and psychopathology

    NARCIS (Netherlands)

    Terwisscha van Scheltinga, Afke F; Bakker, Steven C; Kahn, René S; Kas, Martien J H

    2013-01-01

    In psychiatric disorders, the effect of genetic and environmental factors may converge on molecular pathways and brain circuits related to growth factor functioning. In this review, we describe how disturbances in fibroblast growth factors (FGFs) and their receptors influence behavior by affecting b

  5. Dissecting the Functions of Autophagy in Breast Cancer Associated Fibroblasts

    Science.gov (United States)

    2014-10-01

    unclear. This proposal seeks to understand how stromal fibroblast specific ATG deletion effects mammary tumor progression, and seeks to determine the...the formation of a double membrane organelle, the autophagosome, which sequesters cytoplasmic contents and fuses with the lysosome for degradation...fibroblasts affects mammary tumor development, progression, and metastasis. Task 1: To determine the effects of stromal fibroblast specific atg deletion on

  6. RNA-Guided Activation of Pluripotency Genes in Human Fibroblasts

    DEFF Research Database (Denmark)

    Xiong, Kai; Zhou, Yan; Blichfeld, Kristian Aabo

    2017-01-01

    fibroblasts. This SAM-mediated activation of LOS can be stably maintained for over 20 days in fibroblasts cultured in either fibroblasts or stem cell medium. However, when attempting to use the SAM-LOS activation as an approach for induced pluripotent stem cells-reprogramming, no embryonic stem-like colonies...

  7. The nitric oxide pathway provides innate antiviral protection in conjunction with the type I interferon pathway in fibroblasts.

    Directory of Open Access Journals (Sweden)

    Devangi R Mehta

    Full Text Available The innate host response to virus infection is largely dominated by the production of type I interferon and interferon stimulated genes. In particular, fibroblasts respond robustly to viral infection and to recognition of viral signatures such as dsRNA with the rapid production of type I interferon; subsequently, fibroblasts are a key cell type in antiviral protection. We recently found, however, that primary fibroblasts deficient for the production of interferon, interferon stimulated genes, and other cytokines and chemokines mount a robust antiviral response against both DNA and RNA viruses following stimulation with dsRNA. Nitric oxide is a chemical compound with pleiotropic functions; its production by phagocytes in response to interferon-γ is associated with antimicrobial activity. Here we show that in response to dsRNA, nitric oxide is rapidly produced in primary fibroblasts. In the presence of an intact interferon system, nitric oxide plays a minor but significant role in antiviral protection. However, in the absence of an interferon system, nitric oxide is critical for the protection against DNA viruses. In primary fibroblasts, NF-κB and interferon regulatory factor 1 participate in the induction of inducible nitric oxide synthase expression, which subsequently produces nitric oxide. As large DNA viruses encode multiple and diverse immune modulators to disable the interferon system, it appears that the nitric oxide pathway serves as a secondary strategy to protect the host against viral infection in key cell types, such as fibroblasts, that largely rely on the type I interferon system for antiviral protection.

  8. Transmission of HIV-1 by primary human uterine epithelial cells and stromal fibroblasts.

    Science.gov (United States)

    Asin, Susana N; Fanger, Michael W; Wildt-Perinic, Dunja; Ware, Patricia L; Wira, Charles R; Howell, Alexandra L

    2004-07-15

    Women can become infected with human immunodeficiency virus type 1 (HIV-1) after the heterosexual transmission of virus from an infected male partner. To understand the events that result in transmission of HIV-1 across the female reproductive tract, we characterized the life-cycle events of HIV-1 in primary cultures of human uterine epithelial cells and stromal fibroblasts. Epithelial cells and stromal fibroblasts released virus particles after exposure to either X4- or R5-tropic strains of HIV-1. Virus released by these cells was able to infect CD4(+) T cells. When exposed to an X4-tropic strain of HIV-1, these cells supported HIV-1 reverse transcription, integration, and viral DNA transcription. When exposed to an R5-tropic strain, however, these cells released unmodified virus. These data suggest that uterine cells are targets for productive infection with X4-tropic strains and release unmodified R5-tropic viruses that would then be able to infect submucosal target cells, including T cells and macrophages.

  9. The effect of tranilast on fibroblast activation protein α (FAP-α expression in normal and keloid fibroblasts in vitro

    Directory of Open Access Journals (Sweden)

    Paweł P. Antończak

    2017-07-01

    Full Text Available Introduction . Tranilast (N-(3’,4’-demethoxycinnamoyl-anthranilic acid is an anti-allergic drug. Its mechanism of action is based on the inhibition of antigen-induced release of chemical mediators from mast cells and basophils. It also reveals antifibroproliferative activities. These properties of tranilast are used in the treatment of hypertrophic scars and keloids. Keloids are characterized by incorrect extracellular matrix components turnover. Fibroblasts derived from keloids reveal overproduction of collagen type I and decreased degradation of extracellular matrix in comparison with normal fibroblasts. Fibroblast activation protein α (FAP-α may play an important role in remodeling of extracellular matrix and the invasive properties of keloids. Objective . In the present study, the effect of tranilast on expression of FAP-α gene and its protein was evaluated in normal human dermal fibroblasts and fibroblasts derived from keloids cultured in vitro . Materials and methods. In the first stage of the study, the influence of tranilast on cell viability was estimated. The second stage of the study included the quantitative evaluation of FAP-α mRNA expression in normal and keloid fibroblasts treated with tranilast. The third stage of the study comprised fibroblast activation protein α expression analysis in the examined cells treated with tranilast. Results and conclusions . The expression of FAP-α gene and fibroblast activation protein α is higher in keloid fibroblasts. Tranilast at concentrations of 3 μM and 30 μM up-regulated mRNA FAP-α expression in normal fibroblasts but did not influence keloid fibroblasts. The drug, at concentrations of 30 μM and 300 μM up-regulated fibroblast activation protein α expression in normal fibroblasts and did not influence keloid fibroblasts. Tranilast antiproliferative effect is not associated with FAP-α expression in keloid fibroblasts.

  10. Fibroblast growth factor 23 and its receptors.

    Science.gov (United States)

    Yu, Xijie; White, Kenneth E

    2005-08-01

    Fibroblast growth factor 23 (FGF23) is a circulating factor that plays critical roles in phosphate and vitamin D metabolism, as evidenced by the fact that FGF23 missense mutations cause autosomal dominant hypophosphatemic rickets (ADHR). Autosomal dominant hypophosphatemic rickets is characterized by hypophosphatemia with inappropriately normal 1,25-dihydroxyvitamin D concentrations, as well as bone pain, fracture and rickets. This phenotype parallels that of patients with tumor induced osteomalacia (TIO), X-linked hypophosphatemic rickets (XLH), and fibrous dysplasia (FD), in whom elevated serum FGF23 levels are often observed. The fibroblast growth factor receptors (FGFR1-4) play key roles in skeletal development, as well as in normal metabolic processes. Several FGFR isoforms that potentially mediate the activity of FGF23 have been implicated. In the short term, these findings will lead to further understanding of FGF23 function, and potentially in the long term, to targeted therapies in disorders of hypo- and hyperphosphatemia that involve FGF23.

  11. Myxoinflammatory fibroblastic sarcoma of the face.

    Science.gov (United States)

    Gómez Martín, Cristina; Ortega, María I; Aramburu, José A; Fernández-Cañamaque, José L

    2012-08-01

    Myxoinflammatory fibroblastic sarcoma (MIFS) is a rare low-grade tumor of modified fibroblasts, with tendency to local recurrence. This unusual entity typically presents as a slow-growing painless mass in the distal extremities of middle-aged subjects. A 48-year-old woman presented to our clinic with a painless subcutaneous mass in the right temporal region. Excisional biopsy made the rare diagnosis of MIFS. Histologic examination showed the unique features that characterize this lesion: a myxoid component with a superimposed inflammatory infiltrate and the presence of distinctive, large, and bizarre Reed-Stemberg-like cells. A second wide tumor bed resection was performed, achieving clear margins. No adjuvant therapy was administered, and the patient is free of disease at 18 months postoperatively. To the best of our knowledge, this is the first reported case of MIFS presenting in the face. This adds another possibility for differential diagnoses of soft tissue tumors of the face.

  12. Fibroblast growth factor 23--et fosfatregulerende hormon

    DEFF Research Database (Denmark)

    Beck-Nielsen, Signe Sparre; Pedersen, Susanne Møller; Kassem, Moustapha

    2010-01-01

    Fibroblast growth factor 23 (FGF23) is a recently identified phosphatonin. Its main physiological functions are to maintain serum phosphate within its reference range and to counter regulate the effects of vitamin D. Diseases correlated to high serum values of FGF23 are hypophosphatemic rickets......, fibrous dysplasia, and tumour-induced osteomalacia. In contrast, hyperphosphatemic tumoral calcinosis is associated with accelerated degradation of FGF23. Measuring FGF23 serves as a differential diagnostic tool in elucidating conditions of long-lasting hypophosphatemia....

  13. Fibroblast growth factor 19 entry into brain

    OpenAIRE

    Hsuchou, Hung; Pan, Weihong; Kastin, Abba J.

    2013-01-01

    Background Fibroblast growth factor (FGF)-19, an endocrine FGF protein mainly produced by the ileum, stimulates metabolic activity and alleviates obesity. FGF19 modulates metabolism after either intravenous or intracerebroventricular injection, and its receptor FGFR4 is present in the hypothalamus. This led to the question whether blood-borne FGF19 crosses the blood-brain barrier (BBB) to exert its metabolic effects. Methods We determined the pharmacokinetics of FGF19 permeation from blood to...

  14. 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.

  15. 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.

  16. Fibroblasts as architects of cancer pathogenesis.

    Science.gov (United States)

    Marsh, Timothy; Pietras, Kristian; McAllister, Sandra S

    2013-07-01

    Studies of epithelial cancers (i.e., carcinomas) traditionally focused on transformation of the epithelium (i.e., the cancer cells) and how aberrant signaling within the cancer cells modulates the surrounding tissue of origin. In more recent decades, the normal cells, blood vessels, molecules, and extracellular components that surround the tumor cells, collectively known as the "tumor microenvironment" or "stroma", have received increasing attention and are now thought to be key regulators of tumor initiation and progression. Of particular relevance to the work reviewed herein are the fibroblasts, which make up the major cell type within the microenvironment of most carcinomas. Due to their inherent heterogeneity, plasticity, and function, it is perhaps not surprising that fibroblasts are ideal modulators of normal and cancerous epithelium; however, these aspects also present challenges if we are to interrupt their tumor-supportive functions. Here, we review the current body of knowledge and the many questions that still remain about the special entity known as the cancer-associated fibroblast. This article is part of a Special Issue entitled: Fibrosis: Translation of basic research to human disease.

  17. Avidin is induced in chicken embryo fibroblasts by viral transformation and cell damage.

    Science.gov (United States)

    Korpela, J; Kulomaa, M; Tuohimaa, P; Vaheri, A

    1983-01-01

    Synthesis and secretion of avidin was studied in cultured chicken embryo fibroblasts infected with transforming retroviruses (Rous sarcoma virus, its mutants temperature-sensitive for transformation, OK-10 virus) or a nontransforming retrovirus (RAV-1). Avidin was detectable in both transformed and untransformed cultures, and was identical to chicken egg white avidin by several criteria: biotin-binding, heat-induced biotin exchange, subunit size (mol. wt. 15 600), immunoprecipitation of metabolically labeled proteins and immunoblotting. Transformation increased the production of avidin up to 50-fold, but several experiments suggested that the induction was not a direct consequence of virus-induced cell transformation. The production of avidin seemed to relate to cellular damage both in cultures of virus-transformed and of normal fibroblasts. It may represent a response to cellular damage and viral transformation may activate the process. Images Fig. 2. Fig. 3. PMID:6315397

  18. Avidin is induced in chicken embryo fibroblasts by viral transformation and cell damage.

    Science.gov (United States)

    Korpela, J; Kulomaa, M; Tuohimaa, P; Vaheri, A

    1983-01-01

    Synthesis and secretion of avidin was studied in cultured chicken embryo fibroblasts infected with transforming retroviruses (Rous sarcoma virus, its mutants temperature-sensitive for transformation, OK-10 virus) or a nontransforming retrovirus (RAV-1). Avidin was detectable in both transformed and untransformed cultures, and was identical to chicken egg white avidin by several criteria: biotin-binding, heat-induced biotin exchange, subunit size (mol. wt. 15 600), immunoprecipitation of metabolically labeled proteins and immunoblotting. Transformation increased the production of avidin up to 50-fold, but several experiments suggested that the induction was not a direct consequence of virus-induced cell transformation. The production of avidin seemed to relate to cellular damage both in cultures of virus-transformed and of normal fibroblasts. It may represent a response to cellular damage and viral transformation may activate the process.

  19. Hereditary orotic aciduria, Lesch-Nyhan syndrome, and xeroderma pigmentosum probed by herpes simplex virus: /sup 125/I-iododeoxycytidine incorporation as an assay for viral growth. [Human fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Campisi, J.; Hafner, J.; Boorstein, R.; Pardee, A.B.

    1983-01-01

    /sup 125/I-Iododeoxycytidine (/sup 125/IdC) incorporation into acid-insoluble material was a sensitive, rapid, and quantitative assay for the growth of herpes simplex virus type 1 (HSV-1) in human fibroblasts. Cellular utilization of the isotope was 10 to 25% of the incorporation by infected cells and could be 80% inhibited by tetrahydrouridine (THU). Viral utilization was inhibited by acycloguanosine, thioguanine (TG), and cytosine arabinoside. Isotope was incorporated equally well by growing or quiescent infected cells. HSV-1 was used to probe the metabolic capabilities of three mutant human fibroblast strains. /sup 125/IdC incorporation quantitatively measured the ability of the virus to grow in these cells. Viral /sup 125/IdC incorporation was sensitive to TG in normal fibroblasts but showed a 8- to 10-fold greater resistance to TG in fibroblasts derived from patients with Lesch-Nyhan syndrome (LN). Similarly, the growth of ultraviolet irradiated HSV-1 in normal fibroblasts was 5-fold greater than in fibroblasts derived from patients with xeroderma pigmentosum. In fibroblasts derived from patients with hereditary orotic aciduria, viral /sup 125/IdC incorporation was sensitive to adenosine (AD) at concentrations which were slightly stimulatory in normal fibroblasts. This was a 2-fold difference in AD sensitivity, which the radioassay reliably and quantitatively documented. HSV-1 infected cells could be individually identified by their incorporated /sup 125/IdC; such cells had blackened nuclei in autoradiograms prepared 12 hr after infection. Normal cells infected in the presence of TG had many fewer labeled nuclei than LN cells similarly infected in the presence of the drug. (JMT)

  20. Respiratory activity and growth of human skin derma fibroblasts.

    Science.gov (United States)

    Papa, F; Scacco, S; Vergari, R; Bucaria, V; Dioguardi, D; Papa, S

    1998-09-01

    A study has been made on the speed of growth and respiratory activity of fibroblast cultures from control derma, cheloid (hypertrophic) scar and stabilized scar taken from human skin. The speed of growth and the efficiency of plaque formation of fibroblasts from cheloid scar were greater in comparison with those of fibroblasts from stabilized scar and were stimulated by the addition to the culture medium of the exudate from post-traumatic ulcer. Measurement of the contents of cytochromes showed a decrease in the content of cytochromes b562 and c + c1 in the fibroblast culture from both cheloid and stabilized scar as compared to the fibroblast culture from control derma. Cytochrome aa3 content did not show significant difference among the three types of fibroblast cultures. The respiratory activities supported by pyruvate plus malate, succinate or ascorbate plus N,N,N',N'-tetramethyl-p-phenylenediamine did not show, however, significant difference among the three fibroblast cultures. These observations show that the speed of growth of skin fibroblasts does not depend on the overall respiratory capacity. The exudate stimulated the activity of cytochrome c oxidase in fibroblasts from control derma, and cheloid scar. This effect and the accompanying stimulation of fibroblast growth might be correlated with the balance of oxygen free radicals.

  1. 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.

  2. 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

  3. Study on Isolation, Passage, Cryopreservation and Histology of Mouse Fibroblasts

    Institute of Scientific and Technical Information of China (English)

    ZHANG Gui-xue; LIU Yan; HU Peng-fei

    2004-01-01

    The embryonic ages were determined for the best preparation of mouse fibroblasts. Four methods were adapted to verify cryopreservation of mouse fibroblasts. The results showed that embryonic cryopreserving method was best one with 0.86 of thawing viability. The embryos from 13-14 d pregnant mouse were superior to 11-12 d and 15-16 d in isolating, growing, laying and living. The first 6 generations were better than following ones in the same aspects above. Cell laying time became longer and vailable time became shorter after the sixth generation. With culture time increasing, fibroblast nuclear size became larger, fibrous filament appeared among fibroblasts, and macrocyst vesicle with fioccule appeared in the cells. Cyst vesicle structure with pyknotic granule appeared in 24 h cultured fibroblasts and macrocyst vesicle also appeared in passaging fibroblasts.

  4. Tumor-secreted LOXL2 activates fibroblasts through FAK signaling

    DEFF Research Database (Denmark)

    Barker, Holly E; Bird, Demelza; Lang, Georgina

    2013-01-01

    models. Here, we discovered that tumor-derived LOXL2 directly activated stromal fibroblasts in the tumor microenvironment. Genetic manipulation or antibody inhibition of LOXL2 in orthotopically grown mammary tumors reduced the expression of α-smooth muscle actin (α-SMA). Using a marker for reticular...... fibroblasts, it was determined that expression of α-SMA was localized to fibroblasts recruited from the host tissue. This marker also revealed that the matrix present in tumors with reduced levels of LOXL2 was more scattered compared with control tumors which exhibited matrices with dense, parallel alignments....... Importantly, in vitro assays revealed that tumor-derived LOXL2 and a recombinant LOXL2 protein induced fibroblast branching on collagen matrices, as well as increased fibroblast-mediated collagen contraction and invasion of fibroblasts through extracellular matrix. Moreover, LOXL2 induced the expression of α-SMA...

  5. Reduced CX3CL1 Secretion Contributes to the Susceptibility of Oral Leukoplakia-Associated Fibroblasts to Candida albicans

    Science.gov (United States)

    Cheng, Ran; Li, Duo; Shi, Xueke; Gao, Qinghong; Wei, Changlei; Li, Xiaoyu; Li, Yan; Zhou, Hongmei

    2016-01-01

    Candida leukoplakia (OLK) is a kind of oral leukoplakia combined with chronic candidal infection, which plays an important role in the malignant transformation of OLK. However, little is known about the etiology, including susceptibility of leukoplakia to candidal adhesion, invasion and infection. Some antimicrobial peptides secreted by oral epithelial cells or fibroblasts potentially have antifungal activities against Candida albicans (C. albicans). In this study, we established three co-culture models to simulate different C. albicans-fibroblasts interactions during progression of candida leukoplakia. The susceptibility of oral leukoplakia-associated fibroblasts (LKAFs) to C. albicans and its underlying mechanism were determined. Samples of 14 LKAFs and 10 normal fibroblasts (NFs) were collected. The co-culture models showed that LKAFs had promoted the adhesion, invasion, and survival of C. albicans compared with NFs. CX3CL1, a chemokine with antifungal activity, was less abundant in LKAFs than NFs. Overexpression of CX3CL1 via transfection in LKAFs could partly restore the resistance to C. albicans. We also showed that inhibition of ERK could suppress CX3CL1 secretion. While phosphor-ERK was inhibited in LKAFs compared with NFs. Besides, the mRNA expression of a shedding enzyme for CX3CL1, disintegrin and metalloproteinase domain (ADAM) 17 was decreased in LKAFs than NFs. In conclusion, LKAFs produced and secreted less CX3CL1 by inhibiting the ERK signaling pathway, thereby contributing to impaired cell resistance to C. albicans. PMID:27891323

  6. Reduced CX3CL1 secretion contributes to the susceptibility of oral leukoplakia-associated fibroblasts to Candida albicans

    Directory of Open Access Journals (Sweden)

    Ran Cheng

    2016-11-01

    Full Text Available Candida leukoplakia (OLK is a kind of oral leukoplakia combined with chronic candidal infection, which plays an important role in the malignant transformation of OLK. However, little is known about the etiology, including susceptibility of leukoplakia to candidal adhesion, invasion and infection. Some antimicrobial peptides secreted by oral epithelial cells or fibroblasts potentially have antifungal activities against Candida albicans (C. albicans. In this study, we established three co-culture models to simulate different C. albicans-fibroblasts interactions during progression of candida leukoplakia. The susceptibility of oral leukoplakia-associated fibroblasts (LKAFs to C. albicans and its underlying mechanism were determined. Samples of 14 LKAFs and 10 normal fibroblasts (NFs were collected. The co-culture models showed that LKAFs had promoted the adhesion, invasion, and survival of C. albicans compared with NFs. CX3CL1, a chemokine with antifungal activity, was less abundant in LKAFs than NFs. Overexpression of CX3CL1 via transfection in LKAFs could partly restore the resistance to C. albicans. We also showed that inhibition of ERK could suppress CX3CL1 secretion. While phosphor-ERK was inhibited in LKAFs compared with NFs. Besides, the expression of a shedding enzyme for CX3CL1, disintegrin and metalloproteinase domain (ADAM 17 was decreased in LKAFs than NFs. In conclusion, LKAFs produced and secreted less CX3CL1 by inhibiting the ERK signaling pathway, thereby contributing to impaired cell resistance to C. albicans.

  7. Fibroblasts and Myofibroblasts in Wound Healing: Force Generation and Measurement

    OpenAIRE

    Li, Bin; Wang, James H-C

    2009-01-01

    Fibroblasts are one of the most abundant cell types in connective tissues. These cells are responsible for tissue homeostasis under normal physiological conditions. When tissues are injured, fibroblasts become activated and differentiate into myofibroblasts, which generate large contractions and actively produce extracellular matrix (ECM) proteins to facilitate wound closure. Both fibroblasts and myofibroblasts play a critical role in wound healing by generating traction and contractile force...

  8. Cholesteatoma fibroblasts promote epithelial cell proliferation through overexpression of epiregulin.

    Directory of Open Access Journals (Sweden)

    Mamoru Yoshikawa

    Full Text Available To investigate whether keratinocytes proliferate in response to epiregulin produced by subepithelial fibroblasts derived from middle ear cholesteatoma. Tissue samples were obtained from patients undergoing tympanoplasty. The quantitative polymerase chain reaction and immunohistochemistry were performed to examine epiregulin expression and localization in cholesteatoma tissues and retroauricular skin tissues. Fibroblasts were cultured from cholesteatoma tissues and from normal retroauricular skin. These fibroblasts were used as feeder cells for culture with a human keratinocyte cell line (PHK16-0b. To investigate the role of epiregulin in colony formation by PHK16-0b cells, epiregulin mRNA expression was knocked down in fibroblasts by using short interfering RNA and epiregulin protein was blocked with a neutralizing antibody. Epiregulin mRNA expression was significantly elevated in cholesteatoma tissues compared with that in normal retroauricular skin. Staining for epiregulin was more intense in the epithelial cells and subepithelial fibroblasts of cholesteatoma tissues than in retroauricular skin. When PHK16-0b cells were cultured with cholesteatoma fibroblasts, their colony-forming efficiency was 50% higher than when these cells were cultured with normal skin fibroblasts. Also, knockdown of epiregulin mRNA in cholesteatoma fibroblasts led to greater suppression of colony formation than knockdown in skin fibroblasts. Furthermore, the colony-forming efficiency of PHK16-0b cells was significantly reduced after treatment with an epiregulin neutralizing antibody in co-culture with cholesteatoma fibroblasts, but not in co-culture with skin fibroblasts. These results suggest that keratinocyte hyperproliferation in cholesteatoma is promoted through overexpression of epiregulin by subepithelial fibroblasts via epithelial-mesenchymal interactions, which may play a crucial role in the pathogenesis of middle ear cholesteatoma.

  9. ZO-1: lamellipodial localization in a corneal fibroblast wound model.

    Science.gov (United States)

    Taliana, Lavinia; Benezra, Miriam; Greenberg, Roseanne S; Masur, Sandra K; Bernstein, Audrey M

    2005-01-01

    To explore the roles of ZO-1 in corneal fibroblasts and myofibroblasts in a model of wounding. Antibodies were used to identify ZO-1 in cultured rabbit corneal fibroblasts by immunocytochemistry, Western blot analysis, and immunoprecipitation. For colocalization studies, antibodies to beta-catenin, cadherins, connexins, integrins, alpha-actinin, and cortactin were used. G- and F-actin were identified by DNase and rhodamine phalloidin, respectively. To study ZO-1 localization during cell migration, confluent corneal fibroblasts were subjected to scrape-wounding and evaluated by immunocytochemistry. As predicted from previous studies, ZO-1 colocalized with cadherins and connexin 43 in intercellular junctions. The study revealed a new finding: ZO-1 was also detected at the leading edge of lamellipodia, especially in motile wounded fibroblasts and in freshly plated fibroblasts, before the formation of cell-cell contacts. In fibroblast lysates, ZO-1 largely partitioned to the detergent-soluble fraction compared with myofibroblast lysates, indicating that much of the fibroblast ZO-1 is not associated with insoluble structural components. Lamellipodial ZO-1 colocalized with G-actin, alpha-actinin, and cortactin, which are proteins involved with actin remodeling and cell migration. Integrins alpha5beta1 and alphavbeta3 also localized to the leading edge of migrating fibroblasts, and the association of ZO-1 with integrin was confirmed by immunoprecipitation. Finally, alkaline phosphatase treatment of fibroblast lysate decreased the molecular mass of ZO-1 in lysates of cells grown in serum, demonstrating that, in activated fibroblasts, ZO-1 is phosphorylated. ZO-1's appearance at the leading edge of migrating fibroblasts makes it a candidate for a role in the initiation and organization of integrin-dependent fibroblast adhesion complexes formed during migration and adhesion. Further, phosphorylation of ZO-1 may regulate its cellular localization.

  10. Cardiac fibroblasts regulate sympathetic nerve sprouting and neurocardiac synapse stability.

    Directory of Open Access Journals (Sweden)

    Céline Mias

    Full Text Available Sympathetic nervous system (SNS plays a key role in cardiac homeostasis and its deregulations always associate with bad clinical outcomes. To date, little is known about molecular mechanisms regulating cardiac sympathetic innervation. The aim of the study was to determine the role of fibroblasts in heart sympathetic innervation. RT-qPCR and western-blots analysis performed in cardiomyocytes and fibroblasts isolated from healthy adult rat hearts revealed that Pro-Nerve growth factor (NGF and pro-differentiating mature NGF were the most abundant neurotrophins expressed in cardiac fibroblasts while barely detectable in cardiomyocytes. When cultured with cardiac fibroblasts or fibroblast-conditioned medium, PC12 cells differentiated into/sympathetic-like neurons expressing axonal marker Tau-1 at neurites in contact with cardiomyocytes. This was prevented by anti-NGF blocking antibodies suggesting a paracrine action of NGF secreted by fibroblasts. When co-cultured with cardiomyocytes to mimic neurocardiac synapse, differentiated PC12 cells exhibited enhanced norepinephrine secretion as quantified by HPLC compared to PC12 cultured alone while co-culture with fibroblasts had no effect. However, when supplemented to PC12-cardiomyocytes co-culture, fibroblasts allowed long-term survival of the neurocardiac synapse. Activated fibroblasts (myofibroblasts isolated from myocardial infarction rat hearts exhibited significantly higher mature NGF expression than normal fibroblasts and also promoted PC12 cells differentiation. Within the ischemic area lacking cardiomyocytes and neurocardiac synapses, tyrosine hydroxylase immunoreactivity was increased and associated with local anarchical and immature sympathetic hyperinnervation but tissue norepinephrine content was similar to that of normal cardiac tissue, suggesting depressed sympathetic function. Collectively, these findings demonstrate for the first time that fibroblasts are essential for the setting of

  11. Cardiac Fibroblasts Regulate Sympathetic Nerve Sprouting and Neurocardiac Synapse Stability

    Science.gov (United States)

    Mias, Céline; Coatrieux, Christelle; Denis, Colette; Genet, Gaël; Seguelas, Marie-Hélène; Laplace, Nathalie; Rouzaud-Laborde, Charlotte; Calise, Denis; Parini, Angelo; Cussac, Daniel; Pathak, Atul; Sénard, Jean-Michel; Galés, Céline

    2013-01-01

    Sympathetic nervous system (SNS) plays a key role in cardiac homeostasis and its deregulations always associate with bad clinical outcomes. To date, little is known about molecular mechanisms regulating cardiac sympathetic innervation. The aim of the study was to determine the role of fibroblasts in heart sympathetic innervation. RT-qPCR and western-blots analysis performed in cardiomyocytes and fibroblasts isolated from healthy adult rat hearts revealed that Pro-Nerve growth factor (NGF) and pro-differentiating mature NGF were the most abundant neurotrophins expressed in cardiac fibroblasts while barely detectable in cardiomyocytes. When cultured with cardiac fibroblasts or fibroblast-conditioned medium, PC12 cells differentiated into/sympathetic-like neurons expressing axonal marker Tau-1 at neurites in contact with cardiomyocytes. This was prevented by anti-NGF blocking antibodies suggesting a paracrine action of NGF secreted by fibroblasts. When co-cultured with cardiomyocytes to mimic neurocardiac synapse, differentiated PC12 cells exhibited enhanced norepinephrine secretion as quantified by HPLC compared to PC12 cultured alone while co-culture with fibroblasts had no effect. However, when supplemented to PC12-cardiomyocytes co-culture, fibroblasts allowed long-term survival of the neurocardiac synapse. Activated fibroblasts (myofibroblasts) isolated from myocardial infarction rat hearts exhibited significantly higher mature NGF expression than normal fibroblasts and also promoted PC12 cells differentiation. Within the ischemic area lacking cardiomyocytes and neurocardiac synapses, tyrosine hydroxylase immunoreactivity was increased and associated with local anarchical and immature sympathetic hyperinnervation but tissue norepinephrine content was similar to that of normal cardiac tissue, suggesting depressed sympathetic function. Collectively, these findings demonstrate for the first time that fibroblasts are essential for the setting of cardiac sympathetic

  12. Fibrogenic Lung Injury Induces Non-Cell-Autonomous Fibroblast Invasion.

    Science.gov (United States)

    Ahluwalia, Neil; Grasberger, Paula E; Mugo, Brian M; Feghali-Bostwick, Carol; Pardo, Annie; Selman, Moisés; Lagares, David; Tager, Andrew M

    2016-06-01

    Pathologic accumulation of fibroblasts in pulmonary fibrosis appears to depend on their invasion through basement membranes and extracellular matrices. Fibroblasts from the fibrotic lungs of patients with idiopathic pulmonary fibrosis (IPF) have been demonstrated to acquire a phenotype characterized by increased cell-autonomous invasion. Here, we investigated whether fibroblast invasion is further stimulated by soluble mediators induced by lung injury. We found that bronchoalveolar lavage fluids from bleomycin-challenged mice or patients with IPF contain mediators that dramatically increase the matrix invasion of primary lung fibroblasts. Further characterization of this non-cell-autonomous fibroblast invasion suggested that the mediators driving this process are produced locally after lung injury and are preferentially produced by fibrogenic (e.g., bleomycin-induced) rather than nonfibrogenic (e.g., LPS-induced) lung injury. Comparison of invasion and migration induced by a series of fibroblast-active mediators indicated that these two forms of fibroblast movement are directed by distinct sets of stimuli. Finally, knockdown of multiple different membrane receptors, including platelet-derived growth factor receptor-β, lysophosphatidic acid 1, epidermal growth factor receptor, and fibroblast growth factor receptor 2, mitigated the non-cell-autonomous fibroblast invasion induced by bronchoalveolar lavage from bleomycin-injured mice, suggesting that multiple different mediators drive fibroblast invasion in pulmonary fibrosis. The magnitude of this mediator-driven fibroblast invasion suggests that its inhibition could be a novel therapeutic strategy for pulmonary fibrosis. Further elaboration of the molecular mechanisms that drive non-cell-autonomous fibroblast invasion consequently may provide a rich set of novel drug targets for the treatment of IPF and other fibrotic lung diseases.

  13. Trichomonas vaginalis and Tritrichomonas foetus: interaction with fibroblasts and muscle cells - new insights into parasite-mediated host cell cytotoxicity

    Directory of Open Access Journals (Sweden)

    Ricardo Chaves Vilela

    2012-09-01

    Full Text Available Trichomonas vaginalis and Tritrichomonas foetus are parasitic, flagellated protists that inhabit the urogenital tract of humans and bovines, respectively. T. vaginalis causes the most prevalent non-viral sexually transmitted disease worldwide and has been associated with an increased risk for human immunodeficiency virus-1 infection in humans. Infections by T. foetus cause significant losses to the beef industry worldwide due to infertility and spontaneous abortion in cows. Several studies have shown a close association between trichomonads and the epithelium of the urogenital tract. However, little is known concerning the interaction of trichomonads with cells from deeper tissues, such as fibroblasts and muscle cells. Published parasite-host cell interaction studies have reported contradictory results regarding the ability of T. foetus and T. vaginalis to interact with and damage cells of different tissues. In this study, parasite-host cell interactions were examined by culturing primary human fibroblasts obtained from abdominal biopsies performed during plastic surgeries with trichomonads. In addition, mouse 3T3 fibroblasts, primary chick embryo myogenic cells and L6 muscle cells were also used as models of target cells. The parasite-host cell cultures were processed for scanning and transmission electron microscopy and were tested for cell viability and cell death. JC-1 staining, which measures mitochondrial membrane potential, was used to determine whether the parasites induced target cell damage. Terminal deoxynucleotidyltransferase-mediated dUTP nick end labelling staining was used as an indicator of chromatin damage. The colorimetric crystal violet assay was performed to ana-lyse the cytotoxicity induced by the parasite. The results showed that T. foetus and T. vaginalis adhered to and were cytotoxic to both fibroblasts and muscle cells, indicating that trichomonas infection of the connective and muscle tissues is likely to occur; such

  14. Hsp90 regulation of fibroblast activation in pulmonary fibrosis

    Science.gov (United States)

    Sontake, Vishwaraj; Wang, Yunguan; Kasam, Rajesh K.; Sinner, Debora; Reddy, Geereddy B.; Naren, Anjaparavanda P.; McCormack, Francis X.; Jegga, Anil G.; Madala, Satish K.

    2017-01-01

    Idiopathic pulmonary fibrosis (IPF) is a severe fibrotic lung disease associated with fibroblast activation that includes excessive proliferation, tissue invasiveness, myofibroblast transformation, and extracellular matrix (ECM) production. To identify inhibitors that can attenuate fibroblast activation, we queried IPF gene signatures against a library of small-molecule-induced gene-expression profiles and identified Hsp90 inhibitors as potential therapeutic agents that can suppress fibroblast activation in IPF. Although Hsp90 is a molecular chaperone that regulates multiple processes involved in fibroblast activation, it has not been previously proposed as a molecular target in IPF. Here, we found elevated Hsp90 staining in lung biopsies of patients with IPF. Notably, fibroblasts isolated from fibrotic lesions showed heightened Hsp90 ATPase activity compared with normal fibroblasts. 17-N-allylamino-17-demethoxygeldanamycin (17-AAG), a small-molecule inhibitor of Hsp90 ATPase activity, attenuated fibroblast activation and also TGF-β–driven effects on fibroblast to myofibroblast transformation. The loss of the Hsp90AB, but not the Hsp90AA isoform, resulted in reduced fibroblast proliferation, myofibroblast transformation, and ECM production. Finally, in vivo therapy with 17-AAG attenuated progression of established and ongoing fibrosis in a mouse model of pulmonary fibrosis, suggesting that targeting Hsp90 represents an effective strategy for the treatment of fibrotic lung disease. PMID:28239659

  15. Oral fibroblasts produce more HGF and KGF than skin fibroblasts in response to co-culture with keratinocytes

    DEFF Research Database (Denmark)

    Grøn, Birgitte; Stoltze, Kaj; Andersson, Anders

    2002-01-01

    The production of hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF) in subepithelial fibroblasts from buccal mucosa, periodontal ligament, and skin was determined after co-culture with keratinocytes. The purpose was to detect differences between the fibroblast subpopulations...... that could explain regional variation in epithelial growth and wound healing. Normal human fibroblasts were cultured on polystyrene or maintained in collagen matrix and stimulated with keratinocytes cultured on membranes. The amount of HGF and KGF protein in the culture medium was determined every 24 h for 5...... days by ELISA. When cultured on polystyrene, the constitutive level of KGF and HGF in periodontal fibroblasts was higher than the level in buccal and skin fibroblasts. In the presence of keratinocytes, all three types of fibroblasts in general increased their HGF and KGF production 2-3 times. When...

  16. Coupling of cytoskeleton functions for fibroblast locomotion

    DEFF Research Database (Denmark)

    Couchman, J R; Lenn, M; Rees, D A

    1985-01-01

    caused visible protrusions in projected positions at the leading edge. We conclude that fibroblast locomotion may be driven coordinately by a common set of motility mechanisms and that this coordination may be lost as a result of physical or pharmacological disturbance. Taking our evidence with results...... from other Laboratories, we propose the following cytoskeleton functions. (i) Protrusive activity, probably based on solation--gelation cycles of the actin based cytoskeleton and membrane recycling which provides cellular and membrane components for streaming through the cell body to the leading edge...

  17. 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.

  18. HMC and fibroblast illuminating experiments using microdisplay

    Science.gov (United States)

    Ou, Chung-Jen; Shen, Ching-I.; Ou, Chung-Ming

    2010-02-01

    Techniques like optical neural guiding, photodynamic therapy and photosynthesis of the cell all required specific spatial energy distribution. Influences factors like the wavelength, polarization, spatial intensity distribution are all required, and the appropriate illumination condition for the cells inside the incubator are required to meet more complicated conditions. We report the system that using of the spatial light modulator to provide a multi-points control for the cell culturing. This system is modified from the commercialized projection system to reduce the cost. It is now possible to apply it to other bio-culturing related applications. Results for Human Melanocyte HMC, Glia cell and fibroblast cell are discussed.

  19. Fibroblast growth factor 23 - et fosfatregulerende hormon

    DEFF Research Database (Denmark)

    Beck-Nielsen, Signe; Pedersen, Susanne Møller; Kassem, Moustapha

    2010-01-01

    Fibroblast growth factor 23 (FGF23) er et nyligt identificeret fosfatonin. FGF23's fysiologiske hovedfunktion er at opretholde normalt serumfosfat og at virke som et D-vitaminmodregulatorisk hormon. Sygdomme, der er koblet til forhøjet serum FGF23, er hypofosfatæmisk rakitis, fibrøs dysplasi og...... tumorinduceret osteomalaci. Hyperfosfatæmisk familiær tumoral calcinosis er derimod associeret med forhøjet nedbrydning af FGF23. Måling af FGF23 er et differentialdiagnostisk redskab ved udredning af tilstande med længerevarende hypofosfatæmi. Udgivelsesdato: 2010-May 17...

  20. SV40-mediated immortalization of human fibroblasts.

    Science.gov (United States)

    Ozer, H L; Banga, S S; Dasgupta, T; Houghton, J; Hubbard, K; Jha, K K; Kim, S H; Lenahan, M; Pang, Z; Pardinas, J R; Patsalis, P C

    1996-01-01

    We have identified a multistep mechanism by which the DNA virus SV40 overcomes cellular senescence. Expression of SV40 T antigen is required for both transient extension of life span and unlimited life span or immortalization. These effects are mediated through inactivation of function of growth suppressors pRB and p53 via complex formation with T antigen. However, immortalization additionally requires inactivation of a novel growth suppressor gene, which has recently been identified to be on the distal portion of the long arm of chromosome 6, designated SEN6. We propose that SEN6 is responsible for cellular senescence in fibroblasts and other cells.

  1. Gene Signature of Human Oral Mucosa Fibroblasts: Comparison with Dermal Fibroblasts and Induced Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Keiko Miyoshi

    2015-01-01

    Full Text Available Oral mucosa is a useful material for regeneration therapy with the advantages of its accessibility and versatility regardless of age and gender. However, little is known about the molecular characteristics of oral mucosa. Here we report the first comparative profiles of the gene signatures of human oral mucosa fibroblasts (hOFs, human dermal fibroblasts (hDFs, and hOF-derived induced pluripotent stem cells (hOF-iPSCs, linking these with biological roles by functional annotation and pathway analyses. As a common feature of fibroblasts, both hOFs and hDFs expressed glycolipid metabolism-related genes at higher levels compared with hOF-iPSCs. Distinct characteristics of hOFs compared with hDFs included a high expression of glycoprotein genes, involved in signaling, extracellular matrix, membrane, and receptor proteins, besides a low expression of HOX genes, the hDFs-markers. The results of the pathway analyses indicated that tissue-reconstructive, proliferative, and signaling pathways are active, whereas senescence-related genes in p53 pathway are inactive in hOFs. Furthermore, more than half of hOF-specific genes were similarly expressed to those of hOF-iPSC genes and might be controlled by WNT signaling. Our findings demonstrated that hOFs have unique cellular characteristics in specificity and plasticity. These data may provide useful insight into application of oral fibroblasts for direct reprograming.

  2. Human cytomegalovirus induces alteration of β-actin mRNA and microfilaments in human embryo fibroblast cells

    Institute of Scientific and Technical Information of China (English)

    林茂芳; 魏国庆; 黄河; 蔡真

    2004-01-01

    Objective: To investigate the infection of human embryo fibroblast cell line HF cells by CMV as well as the effects of CMV on β-actin mRNA and microfilaments. Methods: HF cells shape was observed after the infection of CMV.RT-PCR assay was used to detect the mRNA expression of CMV immediate early (IE) gene, β-actin and GAPDH genes of HF cells infected by CMV. CMV particles and cell microfilaments were detected with electron microscope. Results: Shape of HF cell changed after the infection by CMV. HF cells infected by CMV could express IE mRNA and the expression of β-actin mRNA decreased in a time-and titer-dependent manner compared with the uninfected HF cells whose expression of GAPDH mRNA did not change much. CMV particles were found with electron microscope in the cells. Microfilaments were ruptured and shortened after the infection of CMV. Conclusion: CMV can not only infect human embryo fibroblast cells line HF cells and replicate in the cells, but can also affect the expression of β-actin mRNA and the microfilaments.

  3. Dynamic changes of apoptosis in duck embryo fibroblasts induced by new type Gosling viral enteritis virus

    Institute of Scientific and Technical Information of China (English)

    Shun Chen; Anchun Cheng; Mingshu Wang; Xiaoyue Chen

    2008-01-01

    The monolayer duck embryo fibroblast (DEF) cells were experimentally infected with new type Gosling viral enteritis virus (NGVEV) and the dynamic changes of apoptosis were detected at different time points after NGVEV infection by transmission electron microscopy (TEM), DNA agarose gel electrophoresis and Annexin V-FITC/PI stained fluorescence-activated cell sorter (FACS). The result shows that NGVEV can induce infected cells undergoing apoptosis and changing regularly. A series of characteristic apoptotic morphological changes including shrinkage of the cells, chromatin condensation and margination, as well as formation of apoptotic bodies, wereobserved by TEM. The typical ladder pattern of DNA fragmentation was demonstrated by agarose gel electrophoresis. And using flow cytometry analysis of Annexin V-FITC/PI staining, the dead, viable, apoptotic and necrotic cells could be analyzed quantitatively.

  4. 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)

  5. Toll-like receptor 9 mediated responses in cardiac fibroblasts.

    Directory of Open Access Journals (Sweden)

    Ingrid Kristine Ohm

    Full Text Available Altered cardiac Toll-like receptor 9 (TLR9 signaling is important in several experimental cardiovascular disorders. These studies have predominantly focused on cardiac myocytes or the heart as a whole. Cardiac fibroblasts have recently been attributed increasing significance in mediating inflammatory signaling. However, putative TLR9-signaling through cardiac fibroblasts remains non-investigated. Thus, our aim was to explore TLR9-signaling in cardiac fibroblasts and investigate the consequence of such receptor activity on classical cardiac fibroblast cellular functions. Cultivated murine cardiac fibroblasts were stimulated with different TLR9 agonists (CpG A, B and C and assayed for the secretion of inflammatory cytokines (tumor necrosis factor α [TNFα], CXCL2 and interferon α/β. Expression of functional cardiac fibroblast TLR9 was proven as stimulation with CpG B and -C caused significant CXCL2 and TNFα-release. These responses were TLR9-specific as complete inhibition of receptor-stimulated responses was achieved by co-treatment with a TLR9-antagonist (ODN 2088 or chloroquine diphosphate. TLR9-stimulated responses were also found more potent in cardiac fibroblasts when compared with classical innate immune cells. Stimulation of cardiac fibroblasts TLR9 was also found to attenuate migration and proliferation, but did not influence myofibroblast differentiation in vitro. Finally, results from in vivo TLR9-stimulation with subsequent fractionation of specific cardiac cell-types (cardiac myocytes, CD45+ cells, CD31+ cells and cardiac fibroblast-enriched cell-fractions corroborated our in vitro data and provided evidence of differentiated cell-specific cardiac responses. Thus, we conclude that cardiac fibroblast may constitute a significant TLR9 responder cell within the myocardium and, further, that such receptor activity may impact important cardiac fibroblast cellular functions.

  6. Altered chromosome 6 in immortal human fibroblasts.

    Science.gov (United States)

    Hubbard-Smith, K; Patsalis, P; Pardinas, J R; Jha, K K; Henderson, A S; Ozer, H L

    1992-05-01

    Human diploid fibroblasts have a limited life span in vitro, and spontaneous immortalization is an extremely rare event. We have used transformation of human diploid fibroblasts by an origin-defective simian virus 40 genome to develop series of genetically matched immortal cell lines to analyze immortalization. Comparison of a preimmortal transformant (SVtsA/HF-A) with its uncloned and cloned immortalized derivatives (AR5 and HAL) has failed to reveal any major alteration involving the simian virus 40 genome. Karyotypic analysis, however, demonstrated that all of the immortal cell lines in this series have alterations of chromosome 6 involving loss of the portion distal to 6q21. The karyotypic analysis was corroborated by DNA analyses. Southern analysis demonstrated that only one copy of three proto-oncogene loci (ros1, c-myb, and mas1) on 6q was retained in immortal cells. Polymerase chain reaction analysis of the microsatellite polymorphism at 6q22 (D6S87) showed loss of heterozygosity. In addition, elevated expression of c-myb (6q22-23) was observed. We hypothesize that the region at and/or distal to 6q21 plays a role in immortalization, consistent with the presence of a growth suppressor gene.

  7. Altered chromosome 6 in immortal human fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Hubbard-Smith, K.; Pardinas, J.R.; Jha, K.K.; Ozer, H.L. (New Jersey Medical School, Newark, NJ (United States)); Patsalis, P.; Henderson, A.S. (City Univ. of New York, NY (United States))

    1992-05-01

    Human diploid fibroblasts have a limited life span in vitro, and spontaneous immortalization is an extremely rare event. We have used transformation of human diploid fibroblasts by an origin-defective simian virus 40 genome to develop series of genetically matched immortal cell lines to analyze immortalization. Comparison of a preimmortal transformant (SVtsA/HF-A) with its uncloned and cloned immortalized derivatives (AR5 and HAL) has failed to reveal any major alteration involving the simian virus 40 genome. Karyotypic analysis, however, demonstrated that all of the immortal cell lines in this series have alterations of chromosome 6 involving loss of the portion distal to 6q21. The karyotypic analysis was corroborated by DNA analyses. Southern analysis demonstrated that only one copy of three proto-oncogene loci (ros1, c-myb, and mas1) on 6q was retained in immortal cells. Polymerase chain reaction analysis of the microsatellite polymorphism at 6q22 (D6S87) showed loss of heterozygosity. In addition, elevated expression of c-myb (6q22-23) was observed. We hypothesize that the region at and/or distal to 6q21 plays a role in immortalization, consistent with the presence of a growth suppressor gene. 66 refs., 6 figs., 2 tabs.

  8. Povidone-iodine Solutions Inhibit Cell Migration and Survival of Osteoblasts, Fibroblasts, and Myoblasts.

    Science.gov (United States)

    Liu, James X; Werner, Jordan A; Buza, John A; Kirsch, Thorsten; Zuckerman, Joseph D; Virk, Mandeep S

    2017-04-12

    In vitro laboratory study. The purpose of this study was to identify the effect of dilute povidone-iodine (PVI) solutions on human osteoblast, fibroblast and myoblast cells in vitro. Dilute PVI wound lavage has been used successfully in spine and joint arthroplasty procedures to prevent post-operative surgical site infection, but their biologic effect on host cells is largely unknown. Human primary osteoblasts, fibroblasts, and myoblasts were expanded in cell culture and subjected to various concentrations of PVI (0%, 0.001%, 0.01%, 0.1%, 0.35%, 1%) for 3 minutes. To assess the effect of PVI on cell migration, a scratch assay was performed, in which a "scratch" was made by a standard pipette tip in a cell monolayer following PVI exposure, and time to closure of the scratch was evaluated. Cell survival and proliferation was measured 48 hours post-PVI exposure using a cell viability and cytotoxicity assay. Closure of the scratch defect in all cell monolayers was achieved in PVI concentrations PVI concentrations of ≥ 0.1%. PVI concentrations PVI ≥ 0.1% had cell survival rates of less than 6% (p PVI (0.35%) exerts a pronounced cytotoxic effect on osteoblasts, fibroblast, and myoblasts in vitro. Further investigation is required to systematically study the effect of PVI on tissue healing in vivo and also determine a safe and clinically potent concentration for PVI lavage. N/A.

  9. File list: Unc.Epd.50.AllAg.Dermal_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Epd.50.AllAg.Dermal_fibroblasts mm9 Unclassified Epidermis Dermal fibroblasts h...ttp://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Epd.50.AllAg.Dermal_fibroblasts.bed ...

  10. File list: Unc.Epd.20.AllAg.Dermal_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Epd.20.AllAg.Dermal_fibroblasts mm9 Unclassified Epidermis Dermal fibroblasts h...ttp://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Epd.20.AllAg.Dermal_fibroblasts.bed ...

  11. Differential effects of planktonic and biofilm MRSA on human fibroblasts.

    Science.gov (United States)

    Kirker, Kelly R; James, Garth A; Fleckman, Philip; Olerud, John E; Stewart, Philip S

    2012-01-01

    Bacteria colonizing chronic wounds often exist as biofilms, yet their role in chronic wound pathogenesis remains unclear. Staphylococcus aureus biofilms induce apoptosis in dermal keratinocytes, and given that chronic wound biofilms also colonize dermal tissue, it is important to investigate the effects of bacterial biofilms on dermal fibroblasts. The effects of a predominant wound pathogen, methicillin-resistant S. aureus, on normal, human, dermal fibroblasts were examined in vitro. Cell-culture medium was conditioned with equivalent numbers of either planktonic or biofilm methicillin-resistant S. aureus and then fed to fibroblast cultures. Fibroblast response was evaluated using scratch, viability, and apoptosis assays. The results suggested that fibroblasts experience the same fate when exposed to the soluble products of either planktonic or biofilm methicillin-resistant S. aureus, namely limited migration followed by death. Enzyme-linked immunosorbent assays demonstrated that fibroblast production of cytokines, growth factors, and proteases were differentially affected by planktonic and biofilm-conditioned medium. Planktonic-conditioned medium induced more interleukin-6, interleukin-8, vascular endothelial growth factor, transforming growth factor-β1, heparin-bound epidermal growth factor, matrix metalloproteinase-1, and metalloproteinase-3 production in fibroblasts than the biofilm-conditioned medium. Biofilm-conditioned medium induced more tumor necrosis factor-α production in fibroblasts compared with planktonic-conditioned medium, and suppressed metalloproteinase-3 production compared with controls.

  12. Analysis of primary cilia in directional cell migration in fibroblasts

    DEFF Research Database (Denmark)

    Christensen, Søren Tvorup; Veland, Iben; Schwab, Albrecht;

    2013-01-01

    Early studies of migrating fibroblasts showed that primary cilia orient in front of the nucleus and point toward the leading edge. Recent work has shown that primary cilia coordinate a series of signaling pathways critical to fibroblast cell migration during development and in wound healing. In p...

  13. Fibroblast subpopulations in intra-oral wound healing.

    NARCIS (Netherlands)

    Beurden, H.E. van; Snoek, P.A.; Hoff, J.W. Von den; Torensma, R.; Kuijpers-Jagtman, A.M.

    2003-01-01

    The objective of this study was to characterize fibroblasts at sequential time points during intra-oral wound healing in the rat. Experimental wounds were made at several time points in the mucoperiosteum of the palate of 35-day-old Wistar rats. Fibroblasts were cultured from the biopsies under

  14. Physiological ER Stress Mediates the Differentiation of Fibroblasts.

    Directory of Open Access Journals (Sweden)

    Shinsuke Matsuzaki

    Full Text Available Recently, accumulating reports have suggested the importance of endoplasmic reticulum (ER stress signaling in the differentiation of several tissues and cells, including myoblasts and osteoblasts. Secretory cells are easily subjected to ER stress during maturation of their secreted proteins. Skin fibroblasts produce and release several proteins, such as collagens, matrix metalloproteinases (MMPs, the tissue inhibitors of metalloproteinases (TIMPs and glycosaminoglycans (GAGs, and the production of these proteins is increased at wound sites. Differentiation of fibroblasts into myofibroblasts is one of the key factors for wound healing and that TGF-β can induce fibroblast differentiation into myofibroblasts, which express α-smooth muscle actin. Well-differentiated myofibroblasts show increased production of collagen and TGF-β, and bring about wound healing. In this study, we examined the effects of ER stress signaling on the differentiation of fibroblasts, which is required for wound healing, using constitutively ER stress-activated primary cultured fibroblasts. The cells expressed positive α-smooth muscle actin signals without TGF-β stimulation compared with control fibroblasts. Gel-contraction assays suggested that ER stress-treated primary fibroblasts caused stronger shrinkage of collagen gels than control cells. These results suggest that ER stress signaling could accelerate the differentiation of fibroblasts to myofibroblasts at injured sites. The present findings may provide important insights for developing therapies to improve wound healing.

  15. Upregulation of Mitochondrial Content in Cytochrome c Oxidase Deficient Fibroblasts.

    Science.gov (United States)

    Kogot-Levin, Aviram; Saada, Ann; Leibowitz, Gil; Soiferman, Devorah; Douiev, Liza; Raz, Itamar; Weksler-Zangen, Sarah

    2016-01-01

    Cytochrome-c-oxidase (COX) deficiency is a frequent cause of mitochondrial disease and is associated with a wide spectrum of clinical phenotypes. We studied mitochondrial function and biogenesis in fibroblasts derived from the Cohen (CDs) rat, an animal model of COX deficiency. COX activity in CDs-fibroblasts was 50% reduced compared to control rat fibroblasts (P<0.01). ROS-production in CDs fibroblasts increased, along with marked mitochondrial fragmentation and decreased mitochondrial membrane-potential, indicating mitochondrial dysfunction. Surprisingly, cellular ATP content, oxygen consumption rate (OCR) and the extracellular acidification rate (ECAR) were unchanged. To clarify the discrepancy between mitochondrial dysfunction and ATP production, we studied mitochondrial biogenesis and turnover. The content of mitochondria was higher in CDs-fibroblasts. Consistently, AMPK activity and the expression of NRF1-target genes, NRF2 and PGC1-α that mediate mitochondrial biogenesis were increased (P<0.01 vs control fibroblast). In CDs-fibrobalsts, the number of autophagosomes (LC3+ puncta) containing mitochondria in CDs fibroblasts was similar to that in control fibroblasts, suggesting that mitophagy was intact. Altogether, our findings demonstrate that mitochondrial dysfunction and oxidative stress are associated with an increase in mitochondrial biogenesis, resulting in preservation of ATP generation.

  16. Rac inhibition reverses the phenotype of fibrotic fibroblasts.

    Directory of Open Access Journals (Sweden)

    Shi-wen Xu

    Full Text Available BACKGROUND: Fibrosis, the excessive deposition of scar tissue by fibroblasts, is one of the largest groups of diseases for which there is no therapy. Fibroblasts from lesional areas of scleroderma patients possess elevated abilities to contract matrix and produce alpha-smooth muscle actin (alpha-SMA, type I collagen and CCN2 (connective tissue growth factor, CTGF. The basis for this phenomenon is poorly understood, and is a necessary prerequisite for developing novel, rational anti-fibrotic strategies. METHODS AND FINDINGS: Compared to healthy skin fibroblasts, dermal fibroblasts cultured from lesional areas of scleroderma (SSc patients possess elevated Rac activity. NSC23766, a Rac inhibitor, suppressed the persistent fibrotic phenotype of lesional SSc fibroblasts. NSC23766 caused a decrease in migration on and contraction of matrix, and alpha-SMA, type I collagen and CCN2 mRNA and protein expression. SSc fibroblasts possessed elevated Akt phosphorylation, which was also blocked by NSC23766. Overexpression of rac1 in normal fibroblasts induced matrix contraction and alpha-SMA, type I collagen and CCN2 mRNA and protein expression. Rac1 activity was blocked by PI3kinase/Akt inhibition. Basal fibroblast activity was not affected by NSC23766. CONCLUSION: Rac inhibition may be considered as a novel treatment for the fibrosis observed in SSc.

  17. Fibroblasts in myocardial infarction: a role in inflammation and repair

    Science.gov (United States)

    Shinde, Arti V.; Frangogiannis, Nikolaos G.

    2014-01-01

    Fibroblasts do not only serve as matrix-producing reparative cells, but exhibit a wide range of functions in inflammatory and immune responses, angiogenesis and neoplasia. The adult mammalian myocardium contains abundant fibroblasts enmeshed within the interstitial and perivascular extracellular matrix. The current review manuscript discusses the dynamic phenotypic and functional alterations of cardiac fibroblasts following myocardial infarction. Extensive necrosis of cardiomyocytes in the infarcted heart triggers an intense inflammatory reaction. In the early stages of infarct healing, fibroblasts become pro-inflammatory cells, activating the inflammasome and producing cytokines, chemokines and proteases. Pro-inflammatory cytokines (such as Interleukin-1) delay myofibroblast transformation, until the wound is cleared from dead cells and matrix debris. Resolution of the inflammatory infiltrate is associated with fibroblast migration, proliferation, matrix protein synthesis and myofibroblast conversion. Growth factors and matricellular proteins play an important role in myofibroblast activation during the proliferative phase of healing. Formation of a mature cross-linked scar is associated with clearance of fibroblasts, as poorly-understood inhibitory signals restrain the fibrotic response. However, in the non-infarcted remodeling myocardium, local fibroblasts may remain activated in response to volume and pressure overload and may promote interstitial fibrosis. Considering their abundance, their crucial role in cardiac inflammation and repair, and their involvement in myocardial dysfunction and arrhythmogenesis, cardiac fibroblasts may be key therapeutic targets in cardiac remodeling. PMID:24321195

  18. Rho A and the Rho kinase pathway regulate fibroblast contraction: Enhanced contraction in constitutively active Rho A fibroblast cells

    Energy Technology Data Exchange (ETDEWEB)

    Nobe, Koji, E-mail: kojinobe@pharm.showa-u.ac.jp [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan); Nobe, Hiromi [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan); Department of Physical Therapy, Bunkyo-Gakuin University (Japan); Yoshida, Hiroko [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan); Kolodney, Michael S. [Dermatology Division, Department of Medicine, UCLA, Los Angeles, CA (United States); Paul, Richard J. [Department of Molecular and Cellular Physiology, University of Cincinnati College of Medicine, Cincinnati, OH (United States); Honda, Kazuo [Department of Pharmacology, School of Pharmaceutical Sciences, Showa University, Tokyo (Japan)

    2010-08-20

    Research highlights: {yields} Mechanisms of fibroblast cell contraction in collagen matrix. {yields} Assessed an isometric force development using 3D-reconstituted-fibroblast fiber. {yields} Constitutively active Rho A induced the over-contraction of fibroblast cells. {yields} Rho A and Rho kinase pathway has a central role in fibroblast cell contraction. -- Abstract: Fibroblast cells play a central role in the proliferation phase of wound healing processes, contributing to force development. The intracellular signaling pathways regulating this non-muscle contraction are only partially understood. To study the relations between Rho A and contractile responses, constitutively active Rho A (CA-Rho A) fibroblast cells were reconstituted into fibers and the effects of calf serum (CS) on isometric force were studied. CS-induced force in CA-Rho A fibroblast fibers was twice as large as that in wild type (NIH 3T3) fibroblast fibers. During this response, the translocation of Rho A from the cytosol to the membrane was detected by Rho A activity assays and Western blot analysis. Pre-treatment with a Rho specific inhibitor (C3-exoenzyme) suppressed translocation as well as contraction. These results indicate that Rho A activation is essential for fibroblast contraction. The Rho kinase inhibitor ( (Y27632)) inhibited both NIH 3T3 and CA-Rho A fibroblast fiber contractions. Activation of Rho A is thus directly coupled with Rho kinase activity. We conclude that the translocation of Rho A from the cytosol to the membrane and the Rho kinase pathway can regulate wound healing processes mediated by fibroblast contraction.

  19. Intradermal adipocytes mediate fibroblast recruitment during skin wound healing

    Science.gov (United States)

    Schmidt, Barbara A.; Horsley, Valerie

    2013-01-01

    Acute wound healing in the skin involves the communication of multiple cell types to coordinate keratinocyte and fibroblast proliferation and migration for epidermal and dermal repair. Many studies have focused on the interplay between hematopoietic cells, keratinocytes and fibroblasts during skin wound healing, yet the possible roles for other cell types within the skin, such as intradermal adipocytes, have not been investigated during this process. Here, we identify that adipocyte lineage cells are activated and function during acute skin wound healing. We find that adipocyte precursor cells proliferate and mature adipocytes repopulate skin wounds following inflammation and in parallel with fibroblast migration. Functional analysis of mice with defects in adipogenesis demonstrates that adipocytes are necessary for fibroblast recruitment and dermal reconstruction. These data implicate adipocytes as a key component of the intercellular communication that mediates fibroblast function during skin wound healing. PMID:23482487

  20. Hyaluronic acid production by irradiated human synovial fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Yaron, M.; Yaron, I.; Levita, M.; Herzberg, M.

    1977-03-01

    Radioactive particles as well as x irradiation from an external source has been used in the treatment of rheumatoid arthritis, osteoarthritis, and ankylosing spondylitis. In order to clarify effects of ionizing irradiation on synovial cells, radioactive gold (/sup 198/Au) and yttrium (/sup 90/Y) were added to fibroblast cultures derived from human synovial membranes. Other cultures were irradiated by a Picker x-ray machine. Fibroblast growth and hyaluronic acid production were measured. Radioactive gold and yttrium particles induced a significant increase of hyaluronic acid synthesis rate (pg/cell/day) and inhibited fibroblast growth. Fibroblasts continued to overproduce hyaluronic acid and to show growth inhibition 3 weeks after irradiation with radioactive gold. Hydrocortisone inhibited hyaluronic acid overproduction induced by radioactive gold. Overproduction of hyaluronic acid induced by the x-ray machine was inhibited by hydrocortisone, actinomycin-D, and cycloheximide. Fibroblasts derived from normal and rheumatoid patients responded similarly to ionizing irradiation.

  1. EFFECT OF TASPINE ON WOUND HEALING AND FIBROBLAST PROLIFERATION

    Institute of Scientific and Technical Information of China (English)

    Dong Yalin; He Langchong; Chen Fang

    2005-01-01

    Objective To study the effect and mechanism of taspine on wound healing and fibroblast proliferation. Methods The effect of taspine on skin wound was observed in vivo. The different concentration of taspine hydrochloride was added to L929 fibroblast cultivated in vitro, and lactate dehydrogenase was detected and MTT method was applied to observe effect of taspine on fibroblast proliferation. Results The local application of taspine 3 mg/Ml and 1.5 mg/mL accelerated the healing of skin wounded. In vitro, 0.01~0.5 μg/mL of taspine hydrochloride showed no effect on the change of lactate dehydrogenase activity and fibroblast proliferation. Conclusion Taspine is a kind of active alkaloid from leontice robustum which can enhance wound healing, its mechanism on wound healing is not by means of accelerating the proliferation of fibroblast, other mechanisms are necessary for being further studied.

  2. Basic Fibroblast Growth Factor and Fibroblast Growth Factor Receptor-1in Human Meningiomas

    Institute of Scientific and Technical Information of China (English)

    YI Wei; CHEN Jian; Filimon H. Golwa; XUE Delin

    2005-01-01

    The expression of basic fibroblast growth factor (bFGF) and fibroblast growth factor receptor-1 (FGFR-1) in human meningiomas and the relationships between their expression and the tumors' histological features and angiogenesis were investigated by means of immunohistochemical technique. The expression of bFGF and FGFR-1 was detected by antibody of bFGF or FGFR-1.The tumors' angiogenesis was evaluated by microvascular density (MVD) and, which was observed by use of CD34-antibody immunohistochemically. The results showed that there were varied degrees of the expression of bFGF and FGFR-1 proteins in meningiomas. The expression was correlated with the tumors' histological characters and angiogenesis. It was concluded that bFGF and FGFR-1 might play important roles in meningiomas' angiogenesis and proliferation. The expression positive rate of bFGF and FGFR-1 may provide an indication of evaluating the histological and malignant degree of the tumor.

  3. Fibroblast growth factor 23 and bone mineralisation

    Institute of Scientific and Technical Information of China (English)

    Yu-Chen Guo; Quan Yuan

    2015-01-01

    Fibroblast growth factor 23 (FGF23) is a hormone that is mainly secreted by osteocytes and osteoblasts in bone. The critical role of FGF23 in mineral ion homeostasis was first identified in human genetic and acquired rachitic diseases and has been further characterised in animal models. Recent studies have revealed that the levels of FGF23 increase significantly at the very early stages of chronic kidney disease (CKD) and may play a critical role in mineral ion disorders and bone metabolism in these patients. Our recent publications have also shown that FGF23 and its cofactor, Klotho, may play an independent role in directly regulating bone mineralisation instead of producing a systematic effect. In this review, we will discuss the new role of FGF23 in bone mineralisation and the pathophysiology of CKD-related bone disorders.

  4. Beryllium induces premature senescence in human fibroblasts.

    Science.gov (United States)

    Coates, Shannon S A; Lehnert, Bruce E; Sharma, Sunil; Kindell, Susan M; Gary, Ronald K

    2007-07-01

    After cells have completed a sufficient number of cell divisions, they exit the cell cycle and enter replicative senescence. Here, we report that beryllium causes proliferation arrest with premature expression of the principal markers of senescence. After young presenescent human fibroblasts were treated with 3 microM BeSO(4) for 24 h, p21 cyclin-dependent kinase inhibitor mRNA increased by >200%. Longer periods of exposure caused mRNA and protein levels to increase for both p21 and p16(Ink4a), a senescence regulator that prevents pRb-mediated cell cycle progression. BeSO(4) also caused dose-dependent induction of senescence-associated beta-galactosidase activity (SA-beta-gal). Untreated cells had 48 relative fluorescence units (RFU)/microg/h of SA-beta-gal, whereas 3 microM BeSO(4) caused activity to increase to 84 RFU/microg/h. In chromatin immunoprecipitation experiments, BeSO(4) caused p53 protein to associate with its DNA binding site in the promoter region of the p21 gene, indicating that p53 transcriptional activity is responsible for the large increase in p21 mRNA elicited by beryllium. Forced expression of human telomerase reverse transcriptase (hTERT) rendered HFL-1 cells incapable of normal replicative senescence. However, there was no difference in the responsiveness of normal HFL-1 fibroblasts (IC(50) = 1.9 microM) and hTERT-immortalized cells (IC(50) = 1.7 microM) to BeSO(4) in a 9-day proliferation assay. The effects of beryllium resemble those of histone deacetylase-inhibiting drugs, which also cause large increases in p21. However, beryllium produced no changes in histone acetylation, suggesting that Be(2+) acts as a novel and potent pharmacological inducer of premature senescence.

  5. Global deletion of Ankrd1 results in a wound-healing phenotype associated with dermal fibroblast dysfunction.

    Science.gov (United States)

    Samaras, Susan E; Almodóvar-García, Karinna; Wu, Nanjun; Yu, Fang; Davidson, Jeffrey M

    2015-01-01

    The expression of ankyrin repeat domain protein 1 (Ankrd1), a transcriptional cofactor and sarcomeric component, is strongly elevated by wounding and tissue injury. We developed a conditional Ankrd1(fl/fl) mouse, performed global deletion with Sox2-cre, and assessed the role of this protein in cutaneous wound healing. Although global deletion of Ankrd1 did not affect mouse viability or development, Ankrd1(-/-) mice had at least two significant wound-healing phenotypes: extensive necrosis of ischemic skin flaps, which was reversed by adenoviral expression of ANKRD1, and delayed excisional wound closure, which was characterized by decreased contraction and reduced granulation tissue thickness. Skin fibroblasts isolated from Ankrd1(-/-) mice did not spread or migrate on collagen- or fibronectin-coated surfaces as efficiently as fibroblasts isolated from Ankrd1(fl/fl) mice. More important, Ankrd1(-/-) fibroblasts failed to contract three-dimensional floating collagen gels. Reconstitution of ANKRD1 by adenoviral infection stimulated both collagen gel contraction and actin fiber organization. These in vitro data were consistent with in vivo wound closure studies, and suggest that ANKRD1 is important for the proper interaction of fibroblasts with a compliant collagenous matrix both in vitro and in vivo. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  6. Microarray analyses of inflammation response of human dermal fibroblasts to different strains of Borrelia burgdorferi sensu stricto.

    Directory of Open Access Journals (Sweden)

    Frédéric Schramm

    Full Text Available In Lyme borreliosis, the skin is the key site of bacterial inoculation by the infected tick, and of cutaneous manifestations, erythema migrans and acrodermatitis chronica atrophicans. We explored the role of fibroblasts, the resident cells of the dermis, in the development of the disease. Using microarray experiments, we compared the inflammation of fibroblasts induced by three strains of Borrelia burgdorferi sensu stricto isolated from different environments and stages of Lyme disease: N40 (tick, Pbre (erythema migrans and 1408 (acrodermatitis chronica atrophicans. The three strains exhibited a similar profile of inflammation with strong induction of chemokines (CXCL1 and IL-8 and IL-6 cytokine mainly involved in the chemoattraction of immune cells. Molecules such as TNF-alpha and NF-κB factors, metalloproteinases (MMP-1, -3 and -12 and superoxide dismutase (SOD2, also described in inflammatory and cellular events, were up-regulated. In addition, we showed that tick salivary gland extracts induce a cytotoxic effect on fibroblasts and that OspC, essential in the transmission of Borrelia to the vertebrate host, was not responsible for the secretion of inflammatory molecules by fibroblasts. Tick saliva components could facilitate the early transmission of the disease to the site of injury creating a feeding pit. Later in the development of the disease, Borrelia would intensively multiply in the skin and further disseminate to distant organs.

  7. Effect of 660 nm Light-Emitting Diode on the Wound Healing in Fibroblast-Like Cell Lines

    Directory of Open Access Journals (Sweden)

    Myung-Sun Kim

    2015-01-01

    Full Text Available Light in the red to near-infrared (NIR range (630–1000 nm, which is generated using low energy laser or light-emitting diode (LED arrays, was reported to have a range of beneficial biological effects in many injury models. NIR via a LED is a well-accepted therapeutic tool for the treatment of infected, ischemic, and hypoxic wounds as well as other soft tissue injuries in humans and animals. This study examined the effects of exposure to 660 nm red LED light at intensities of 2.5, 5.5, and 8.5 mW/cm2 for 5, 10, and 20 min on wound healing and proliferation in fibroblast-like cells, such as L929 mouse fibroblasts and human gingival fibroblasts (HGF-1. A photo illumination-cell culture system was designed to evaluate the cell proliferation and wound healing of fibroblast-like cells exposed to 600 nm LED light. The cell proliferation was evaluated by MTT assay, and a scratched wound assay was performed to assess the rate of migrating cells and the healing effect. Exposure to the 660 nm red LED resulted in an increase in cell proliferation and migration compared to the control, indicating its potential use as a phototherapeutic agent.

  8. MAPs/bFGF-PLGA microsphere composite-coated titanium surfaces promote increased adhesion and proliferation of fibroblasts.

    Science.gov (United States)

    Wang, Zhongshan; Wu, Guofeng; Bai, Shizhu; Feng, Zhihong; Dong, Yan; Zhou, Jian; Qin, Haiyan; Zhao, Yimin

    2014-06-01

    Infection and epithelial downgrowth are two major problems with maxillofacial transcutaneous implants, and both are mainly due to lack of stable closure of soft tissues at transcutaneous sites. Fibroblasts have been shown to play a key role in the formation of biological seals. In this work, titanium (Ti) model surfaces were coated with mussel adhesive proteins (MAPs) utilizing its unique adhesion ability on diverse inorganic and organic surfaces in wet environments. Prepared basic fibroblast growth factor (bFGF)-poly(lactic-co-glycolic acid) (PLGA) microspheres can be easily synthesized and combined onto MAPs-coated Ti surfaces, due to the negative surface charges of microspheres in aqueous solution, which is in contrast to the positive charges of MAPs. Titanium model surfaces were divided into three groups. Group A: MAPs/bFGF-PLGA microspheres composite-coated Ti surfaces. Group B: MAPs-coated Ti surfaces. Group C: uncoated Ti surfaces. The effects of coated Ti surfaces on adhesion of fibroblasts, cytoskeletal organization, proliferation, and extracellular matrix (ECM)-related gene expressions were examined. The results revealed increased adhesion (P composite-coated Ti surfaces had the ability to increase fibroblast functionality. In addition, MAPs/bFGF-PLGA microsphere composite-coated Ti surfaces should be studied further as a method of promoting formation of stable biological seals around transcutaneous sites.

  9. Apoptosis of wound fibroblasts induced by oxidative stress.

    Science.gov (United States)

    Takahashi, Atsushi; Aoshiba, Kazutetsu; Nagai, Atsushi

    2002-06-01

    Irreversible lung parenchymal injury is usually healed by fibrosis, which depends on the abilities of fibroblasts to proliferate, migrate into the wound, and survive. Because the lung is frequently exposed to increased oxidative stress, which is thought to mediate apoptosis, we examined whether oxidative stress induces apoptosis in fibroblasts during wound healing. We performed an in vitro scratch wound assay where cultured fibroblast monolayers were exposed to H2O2 (10-500 microM) after artificial wounding. Apoptosis was evaluated by nuclear staining with Hoechst33342 or terminal deoxynucleotidyl transferase (TdT)-mediated nucleotide nick end-labeling (TUNEL). Intracellular oxidants were assessed with the peroxide-sensitive fluorochrome carboxydichlorodihydrofluorescein (CDCF). We found that repopulating fibroblasts at the wound margin, but not quiescent fibroblasts at the intact site, selectively underwent oxidant accumulation and apoptosis in response to H2O2 exposure. Some of the apoptotic cells had incorporated bromodeoxyuridine (BrdU), an indicator of proliferating cells. These results suggest that oxidative stress selectively induces apoptosis in fibroblasts that are stimulated to proliferate and/or migrate into the wound. Fibroblast apoptosis induced by oxidative stress during wound repopulation may be relevant to intractable wound healing.

  10. LIF Mediates Proinvasive Activation of Stromal Fibroblasts in Cancer

    Directory of Open Access Journals (Sweden)

    Jean Albrengues

    2014-06-01

    Full Text Available Signaling crosstalk between tumor cells and fibroblasts confers proinvasive properties to the tumor microenvironment. Here, we identify leukemia inhibitory factor (LIF as a tumor promoter that mediates proinvasive activation of stromal fibroblasts independent of alpha-smooth muscle actin (α-SMA expression. We demonstrate that a pulse of transforming growth factor β (TGF-β establishes stable proinvasive fibroblast activation by inducing LIF production in both fibroblasts and tumor cells. In fibroblasts, LIF mediates TGF-β-dependent actomyosin contractility and extracellular matrix remodeling, which results in collective carcinoma cell invasion in vitro and in vivo. Accordingly, carcinomas from multiple origins and melanomas display strong LIF upregulation, which correlates with dense collagen fiber organization, cancer cell collective invasion, and poor clinical outcome. Blockade of JAK activity by Ruxolitinib (JAK inhibitor counteracts fibroblast-dependent carcinoma cell invasion in vitro and in vivo. These findings establish LIF as a proinvasive fibroblast producer independent of α-SMA and may open novel therapeutic perspectives for patients with aggressive primary tumors.

  11. Bacterial lipopolysaccharide promotes profibrotic activation of intestinal fibroblasts.

    LENUS (Irish Health Repository)

    Burke, J P

    2012-02-01

    BACKGROUND: Fibroblasts play a critical role in intestinal wound healing. Lipopolysaccharide (LPS) is a cell wall component of commensal gut bacteria. The effects of LPS on intestinal fibroblast activation were characterized. METHODS: Expression of the LPS receptor, toll-like receptor (TLR) 4, was assessed in cultured primary human intestinal fibroblasts using flow cytometry and confocal microscopy. Fibroblasts were treated with LPS and\\/or transforming growth factor (TGF) beta1. Nuclear factor kappaB (NFkappaB) pathway activation was assessed by inhibitory kappaBalpha (IkappaBalpha) degradation and NFkappaB promoter activity. Fibroblast contractility was measured using a fibroblast-populated collagen lattice. Smad-7, a negative regulator of TGF-beta1 signalling, and connective tissue growth factor (CTGF) expression were assessed using reverse transcriptase-polymerase chain reaction and western blot. The NFkappaB pathway was inhibited by IkappaBalpha transfection. RESULTS: TLR-4 was present on the surface of intestinal fibroblasts. LPS treatment of fibroblasts induced IkappaBalpha degradation, enhanced NFkappaB promoter activity and increased collagen contraction. Pretreatment with LPS (before TGF-beta1) significantly increased CTGF production relative to treatment with TGF-beta1 alone. LPS reduced whereas TGF-beta1 increased smad-7 expression. Transfection with an IkappaBalpha plasmid enhanced basal smad-7 expression. CONCLUSION: Intestinal fibroblasts express TLR-4 and respond to LPS by activating NFkappaB and inducing collagen contraction. LPS acts in concert with TGF-beta1 to induce CTGF. LPS reduces the expression of the TGF-beta1 inhibitor, smad-7.

  12. Ets2 in tumor fibroblasts promotes angiogenesis in breast cancer.

    Directory of Open Access Journals (Sweden)

    Julie A Wallace

    Full Text Available Tumor fibroblasts are active partners in tumor progression, but the genes and pathways that mediate this collaboration are ill-defined. Previous work demonstrates that Ets2 function in stromal cells significantly contributes to breast tumor progression. Conditional mouse models were used to study the function of Ets2 in both mammary stromal fibroblasts and epithelial cells. Conditional inactivation of Ets2 in stromal fibroblasts in PyMT and ErbB2 driven tumors significantly reduced tumor growth, however deletion of Ets2 in epithelial cells in the PyMT model had no significant effect. Analysis of gene expression in fibroblasts revealed a tumor- and Ets2-dependent gene signature that was enriched in genes important for ECM remodeling, cell migration, and angiogenesis in both PyMT and ErbB2 driven-tumors. Consistent with these results, PyMT and ErbB2 tumors lacking Ets2 in fibroblasts had fewer functional blood vessels, and Ets2 in fibroblasts elicited changes in gene expression in tumor endothelial cells consistent with this phenotype. An in vivo angiogenesis assay revealed the ability of Ets2 in fibroblasts to promote blood vessel formation in the absence of tumor cells. Importantly, the Ets2-dependent gene expression signatures from both mouse models were able to distinguish human breast tumor stroma from normal stroma, and correlated with patient outcomes in two whole tumor breast cancer data sets. The data reveals a key function for Ets2 in tumor fibroblasts in signaling to endothelial cells to promote tumor angiogenesis. The results highlight the collaborative networks that orchestrate communication between stromal cells and tumor cells, and suggest that targeting tumor fibroblasts may be an effective strategy for developing novel anti-angiogenic therapies.

  13. Metabolites of Hypoxic Cardiomyocytes Induce the Migration of Cardiac Fibroblasts.

    Science.gov (United States)

    Shi, Huairui; Zhang, Xuehong; He, Zekun; Wu, Zhiyong; Rao, Liya; Li, Yushu

    2017-01-01

    The migration of cardiac fibroblasts to the infarct region plays a major role in the repair process after myocardial necrosis or damage. However, few studies investigated whether early hypoxia in cardiomyocytes induces the migration of cardiac fibroblasts. The purpose of this study was to assess the role of metabolites of early hypoxic cardiomyocytes in the induction of cardiac fibroblast migration. Neonatal rat heart tissue was digested with a mixture of trypsin and collagenase at an appropriate ratio. Cardiomyocytes and cardiac fibroblasts were cultured via differential adhesion. The cardiomyocyte cultures were subjected to hypoxia for 2, 4, 6, 8, 10, and 12 h. The supernatants of the cardiomyocyte cultures were collected to determine the differences in cardiac fibroblast migration induced by hypoxic cardiomyocyte metabolites at various time points using a Transwell apparatus. Meanwhile, ELISA was performed to measure TNF-α, IL-1β and TGF-β expression levels in the cardiomyocyte metabolites at various time points. The metabolites of hypoxic cardiomyocytes significantly induced the migration of cardiac fibroblasts. The induction of cardiac fibroblast migration was significantly enhanced by cardiomyocyte metabolites in comparison to the control after 2, 4, and 6 h of hypoxia, and the effect was most significant after 2 h. The expression levels of TNF-α, IL-1β, IL-6, and TGF-β were substantially increased in the metabolites of cardiomyocytes, and neutralization with anti-TNF-α and anti-IL-1β antibodies markedly reduced the induction of cardiac fibroblast migration by the metabolites of hypoxic cardiomyocytes. The metabolites of early hypoxic cardiomyocytes can induce the migration of cardiac fibroblasts, and TNF-α and IL-1β may act as the initial chemotactic inducers. © 2017 The Author(s) Published by S. Karger AG, Basel.

  14. HGF is released from buccal fibroblasts after smokeless tobacco stimulation

    DEFF Research Database (Denmark)

    Dabelsteen, S; Christensen, S; Gron, B

    2005-01-01

    To investigate the effect of smokeless tobacco (ST) on (1) HGF, KGF and GM-CSF expression by buccal fibroblasts and (2) on keratinocyte and fibroblast proliferation. Buccal fibroblasts were stimulated with different concentrations of ST extracts in a double dilution from 0.50% w/v to 0.03% w...... on exposure time and on concentration of the tobacco extract. High concentration increased production of HGF 4-fold. KGF production was doubled when high concentration of tobacco was used, low concentration did not stimulate cells. GM-CSF production was low in both stimulated and non-stimulated cells...

  15. Antibacterial activity of extract and fractions from branches of Protium spruceanum and cytotoxicity on fibroblasts.

    Science.gov (United States)

    Amparo, Tatiane Roquete; Rodrigues, Ivanildes Vasconcelos; Seibert, Janaína Brandão; Souza, Rafaella Hilda Zaniti; Oliveira, Amanda Ribeiro de; Cabral, Vivette Appolinário Rodrigues; Vieira, Paula Melo de Abreu; Brandão, Geraldo Célio; Okuma, Adriana Akemi; Filho, Sidney Augusto Vieira; Teixeira, Luiz Fernando Medeiros; Souza, Gustavo Henrique Bianco de

    2017-07-20

    The crude ethanol extract (CEE) and fractions from branches of Protium spruceanum were subjected to antibacterial and cytotoxicity assays. Compounds of the most active fraction were identified by GC-MS and LC-MS. CEE was active against 19 bacteria and the ethyl acetate fraction (EAF) showed the lowest minimum bactericidal concentration (MBC 0.3-80.0 mg/mL). Through time-kill assay was observed that EAF induced rapid bactericidal effect against Staphylococcus saprophyticus. The cytotoxicity tests against L929 fibroblasts showed great potential of EAF on the treatment of infections caused by five bacteria (MBC < IC50). The results provide in vitro scientific support to the possible application of branches of P. spruceanum as antimicrobial agent that may contribute for treatment of infections.

  16. PTCH1 +/- dermal fibroblasts isolated from healthy skin of Gorlin syndrome patients exhibit features of carcinoma associated fibroblasts.

    Directory of Open Access Journals (Sweden)

    Alexandre Valin

    Full Text Available Gorlin's or nevoid basal cell carcinoma syndrome (NBCCS causes predisposition to basal cell carcinoma (BCC, the commonest cancer in adult human. Mutations in the tumor suppressor gene PTCH1 are responsible for this autosomal dominant syndrome. In NBCCS patients, as in the general population, ultraviolet exposure is a major risk factor for BCC development. However these patients also develop BCCs in sun-protected areas of the skin, suggesting the existence of other mechanisms for BCC predisposition in NBCCS patients. As increasing evidence supports the idea that the stroma influences carcinoma development, we hypothesized that NBCCS fibroblasts could facilitate BCC occurence of the patients. WT (n = 3 and NBCCS fibroblasts bearing either nonsense (n = 3 or missense (n = 3 PTCH1 mutations were cultured in dermal equivalents made of a collagen matrix and their transcriptomes were compared by whole genome microarray analyses. Strikingly, NBCCS fibroblasts over-expressed mRNAs encoding pro-tumoral factors such as Matrix Metalloproteinases 1 and 3 and tenascin C. They also over-expressed mRNA of pro-proliferative diffusible factors such as fibroblast growth factor 7 and the stromal cell-derived factor 1 alpha, known for its expression in carcinoma associated fibroblasts. These data indicate that the PTCH1(+/- genotype of healthy NBCCS fibroblasts results in phenotypic traits highly reminiscent of those of BCC associated fibroblasts, a clue to the yet mysterious proneness to non photo-exposed BCCs in NBCCS patients.

  17. Egg drop syndrome virus enters duck embryonic fibroblast cells via clathrin-mediated endocytosis.

    Science.gov (United States)

    Huang, Jingjing; Tan, Dan; Wang, Yang; Liu, Caihong; Xu, Jiamin; Wang, Jingyu

    2015-12-02

    Previous studies of egg drop syndrome virus (EDSV) is restricted to serological surveys, disease diagnostics, and complete viral genome analysis. Consequently, the infection characteristics and entry routes of EDSV are poorly understood. Therefore, we aimed to explore the entry pathway of EDSV into duck embryonic fibroblast (DEF) cells as well as the infection characteristics and proliferation of EDSV in primary DEF and primary chicken embryo liver (CEL) cells. Transmission electron microscopy revealed that the virus triggered DEF cell membrane invagination as early as 10 min post-infection and that integrated endocytic vesicles formed at 20 min post-infection. The virus yield in EDSV-infected DEF cells treated with chlorpromazine (CPZ), sucrose, methyl-β-cyclodextrin (MβCD), or NH4Cl was measured by quantitative real-time PCR. Compared with the mock treatment, CPZ and sucrose greatly inhibited the production of viral progeny in a dose-dependent manner, while MβCD treatment did not result in a significant difference. Furthermore, NH4Cl had a strong inhibitory effect on the production of EDSV progeny. In addition, indirect immunofluorescence demonstrated that virus particles clustered on the surface of DEF cells treated with CPZ or sucrose. These results indicate that EDSV enters DEF cells through clathrin-mediated endocytosis followed by a pH-dependent step, which is similar to the mechanism of entry of human adenovirus types 2 and 5. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. Transient expression of acidic fibroblast growth factor in pea (Pisum ...

    African Journals Online (AJOL)

    Yomi

    2012-03-15

    Mar 15, 2012 ... Key words: Pea plants, acidic fibroblast growth factor, leave injection, plant ... It plays important roles in various stages of development and morphogenesis and also in angiogenesis and wound healing processes (Basilico and.

  19. Inhibitive Effects of Quercetin on Rabbit Tenon Capsule Fibroblasts Proliferation

    Institute of Scientific and Technical Information of China (English)

    Su Liu; Lin Chen

    2005-01-01

    Purpose:To study the inhibitive effects of quercetin (QU) on the fibroblasts proliferation of rabbit Tenon's capsule and its mechanism.Methods: Cultured fibroblasts were exposed to different concentrations of QU solution and investigated by microculture tetrazolium (MTT) assay. The effect of QU was obser ved on cells cycle using the flow cytometer. Besults: QU can suppress the proliferation of rabbit Tenon's capsule fibroblasts in vitro and show a dose-time dependent tendency.Flow cytometer results showed 26.92% cell increase in G1 phase, 23.50% decrease in S phase and 3.42% decrease in G2 phase.Conclusions: QU can suppress the proliferation of rabbit Tenon's capsule fibroblasts in vitro and show a dose-time dependent tendency. QU may effect all phase of cell cycle and inhibit cell proliferation by inhibiting G1 phase transitting to S phase and G2 phase.

  20. Growth of fibroblasts and endothelial cells on wettability gradient surfaces

    NARCIS (Netherlands)

    Ruardy, TG; Moorlag, HE; Schakenraad, JM; VanderMei, HC; Busscher, HJ

    1997-01-01

    The growth, spreading, and shape of human skin fibroblasts (PK 84) and human umbilical cord endothelial cells on dichlorodimethylsilane (DDS) and dimethyloctadecylchlorosilane (DOGS) gradient surfaces were investigated in the presence of serum proteins. Gradient surfaces were prepared on glass using

  1. Overexpression of CDKN1B inhibits fibroblast proliferation in a rabbit model of experimental glaucoma filtration surgery.

    Science.gov (United States)

    Yang, Jian-gang; Deng, Ying; Zhou, Ling-xiao; Li, Xiao-yan; Sun, Peng-rui; Sun, Nai-xue

    2013-01-14

    To investigate the potential antiproliferative effect of cyclin-dependent kinase inhibitor 1B (CDKN1B) overexpression in a rabbit model of glaucoma filtration surgery (GFS). The recombinant adenovector expressing exogenous CDKN1B was delivered to Tenon's capsule by subconjunctival injection during unilateral filtration surgery. The time course of CDKN1B expression was monitored by immunohistochemistry and Western blot analysis. Evaluation of proliferating activity was performed by proliferating cell nuclear antigen (PCNA), argyrophilic nucleolar organizing region (AgNOR) staining, and fibroblast-specific protein 1 (FSP-1). Cyclin-dependent kinase 2 (Cdk2) and Cdk4 expression were detected with immunohistochemical analysis. The overexpression of CDKN1B in Tenon's capsule was monitored throughout the experimental period. Immunoreactivity to CDKN1B was mainly observed in the nucleus of fibroblasts. The increased expression of CDKN1B in sclera was detected up to 21 days after viral infection, whereas the level of CDKN1B protein in corneal stroma was not significantly increased. The overexpression of CDKN1B induced a significant decrease in AgNOR number/nucleus and area/nucleus, PCNA staining, FSP-1 positive cells, and the decreased expressions of Cdk2 and Cdk4, as evidenced by nuclear and cytoplasmic immunoreactivity to Cdk2 and Cdk4 antibodies in positive fibroblasts. The persistent overexpression of CDKN1B mediated by the recombinant adenovector expressing exogenous CDKN1B in Tenon's fibroblasts after GFS may lead to the inhibition of fibroblast proliferation and the downregulation of Cdk2 and Cdk4 activity, thereby reducing the severity of scar formation and the surgical outcome.

  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. Tumor-secreted LOXL2 Activates Fibroblasts Through FAK Signaling

    OpenAIRE

    Barker, Holly E.; Bird, Demelza; Lang, Georgina; Janine T. Erler

    2013-01-01

    Cancer-associated fibroblasts enhance cancer progression when activated by tumor cells through mechanisms not yet fully understood. Blocking mammary tumor cell-derived lysyl oxidase-like 2 (LOXL2) significantly inhibited mammary tumor cell invasion and metastasis in transgenic and orthotopic mouse models. Here we discovered that tumor-derived LOXL2 directly activated stromal fibroblasts in the tumor microenvironment. Genetic manipulation or antibody inhibition of LOXL2 in orthotopically grown...

  4. Understanding the role of stromal fibroblasts in cancer progression

    OpenAIRE

    2012-01-01

    The major cellular components of tumor microenvironment, referred to as the cancer stroma, are composed of cancer-associated fibroblasts that support tumor epithelial growth, invasion and therapeutic resistance. Thus when we speak of developing therapies that address tumor heterogeneity it is not only a matter of different mutations within the tumor epithelia. While individual mutations in the stromal compartment are controversial, the heterogeneity in fibroblastic population in a single tumo...

  5. Preclinical safety studies on autologous cultured human skin fibroblast transplantation.

    Science.gov (United States)

    Zeng, Wei; Zhang, Shuying; Liu, Dai; Chai, Mi; Wang, Jiaqi; Zhao, Yuming

    2014-01-01

    Recently, FDA approved the clinical use of autologous fibroblasts (LAVIV™) for the improvement of nasolabial fold wrinkles in adults. The use of autologous fibroblasts for the augmentation of dermal and subcutaneous defects represents a potentially exciting natural alternative to the use of other filler materials for its long-term corrective ability and absence of allergic adverse effects proved by clinical application. However, compared to the clinical evidence, preclinical studies are far from enough. In this study, human skin-derived fibroblasts were cultured and expanded for both in vitro and in vivo observations. In vitro, the subcultured fibroblasts were divided into two groups. One set of cells underwent cell cycle and karyotype analysis at passages 5 and 10. The second group of cells was cocultured in medium with different concentrations of human skin extract D for the measurement of collagen concentration and cell count. In vivo, the subcultured fibroblasts were injected into nude mice subcutaneously. Biopsies were taken for morphology observation and specific collagen staining at 1, 2, and 3 months after injection. The results in vitro showed no significant differences in cell cycle distribution between passages 5 and 10. Cell proliferation and secretion were inhibited as the concentration of extract D increased. In vivo, the fibroblasts were remarkably denser on the experimental side with no dysplastic cells. Mitotic cells were easily observed at the end of the first month but were rare at the end of the third month. Type III collagen was detected at the end of the first month, while collagen type I was positive at the end of the second month. The content of both collagens increased as time passed. The above results indicated that the use of the autologous fibroblasts was safe, providing a basic support for clinical use of fibroblasts.

  6. Fibroblasts as Efficient Antigen-Presenting Cells in Lymphoid Organs

    Science.gov (United States)

    Kundig, Thomas M.; Bachmann, Martin F.; Dipaolo, Claudio; Simard, John J. L.; Battegay, Manuel; Lother, Heinz; Gessner, Andre; Kuhlcke, Klaus; Ohashi, Pamela S.; Hengartner, Hans; Zinkernagel, Rolf M.

    1995-06-01

    Only so-called "professional" antigen-presenting cells (APCs) of hematopoietic origin are believed capable of inducing T lymphocyte responses. However, fibroblasts transfected with viral proteins directly induced antiviral cytotoxic T lymphocyte responses in vivo, without involvement of host APCs. Fibroblasts induced T cells only in the milieu of lymphoid organs. Thus, antigen localization affects self-nonself discrimination and cell-based vaccine strategies.

  7. Localization of ZO-1 in the nucleolus of corneal fibroblasts.

    Science.gov (United States)

    Benezra, Miriam; Greenberg, Roseanne S; Masur, Sandra K

    2007-05-01

    Within the multidomain structure of ZO-1 are motifs responsible for ZO-1's localization to intercellular junctions and its newly demonstrated localization to the leading edge of lamellipodia in corneal fibroblasts. Since ZO-1 also has two nuclear localization signals, this study was undertaken to determine whether stimuli associated with wounding would induce nuclear translocation of ZO-1 Immunocytochemistry and immunoblot analysis were used to localize endogenous and exogenous ZO-1 in nuclear and cytoplasmic sites in corneal fibroblasts and 293T fibroblasts, with and without myc-ZO-1 transfection. Cells were serum starved by growth for 48 hours in DMEM/F12 with 0.2% FBS and subsequently were either scrape wounded or treated with 10% FBS, PDGF, or FGF-2 for 6 hours. For immunoblot analysis, after lysis, the nuclear and cytosolic fractions were separated and analyzed by SDS-PAGE. Cells on companion coverslips were fixed with 3% p-formaldehyde and permeabilized with 1% Triton before immunocytochemical detection of ZO-1 and nuclear proteins. ZO-1 was rarely detected in the nucleus of serum-starved corneal fibroblasts. In contrast, it colocalized with nucleolin in the nucleoli of corneal fibroblasts after serum-starved cells were treated with 10% FBS, PDGF, or FGF-2. Immunoblot analysis confirmed the immunocytochemical results: Little ZO-1 was detected in the nuclear fraction of lysates of serum-starved cells, but ZO-1 was found in the nuclear fractions of rabbit corneal and 293T fibroblasts treated with 10% FBS, PDGF, or FGF-2. Furthermore in scrape-wounded corneal fibroblasts, ZO-1 was localized to nucleoli of both serum-starved and serum-treated cells. Localization of ZO-1 to nucleoli of corneal and 293T fibroblasts under proliferative and promigratory conditions suggests a physiologically significant interaction of ZO-1 with proteins in nucleoli during the healing process.

  8. Fibroblast behavior after titanium surfaces exposure.

    Science.gov (United States)

    Danza, Matteo; Zollino, Ilaria; Candotto, Valentina; Cura, Francesca; Carinci, Francesco

    2012-12-01

    The main requirements for a good material are its ability to promote attraction and adhesion of bone precursor cells and their proliferation and differentiation. Different biocompatible materials are currently employed as scaffold. Among these, titanium is considered a gold standard because of its biocompatibility and good corrosion resistance. The aim of this work was to compare two different AoN titanium layers (GR4 and GR5) to investigate which one had a greater osteoconductive power using human fibroblasts (HFb) culture at two different time-points. The expression levels of some adhesion and traction-resistance related genes (COL11A1, COL2A1, COL9A1, DSP, ELN, HAS1, and TFRC) were analyzed using real time reverse transcription-polymerase chain reaction. After 7 days of treatment with TiA 4GR, the only two up-regulated genes were COL2A1 and DSP. After 15 days of treatment, none of genes over expressed. Our preliminary results suggest that neither AoN 4GR nor AoN 5GR are able to promote the production of protein involved in cell-cell and cell-matrix adhesion and in stress-resistance, required for a good outcome in dental implantology.

  9. Investigation of CNTs interaction with fibroblast cells.

    Science.gov (United States)

    Pensabene, V; Vittorio, O; Raffa, V; Menciassi, A; Dario, P

    2007-01-01

    The need for toxicological studies on carbon nanotubes (CNTs) has arisen from the rapidly emerging applications of CNTs well beyond material science and engineering. In order to provide a method to collect data about toxicology, we characterized by Scanning Electron Microscopy (SEM), by Energy Dispersive X-ray Spectrometry (EDS) analysis and by Focused Ion Beam (FIB) microscopy different kinds of treated CNTs. The bio-interaction was investigated seeding Crandell feline kidney fibroblasts with CNT-modified medium; a dedicated sample preparation by FIB has been defined to fix cells. In the present study, the cytotoxic effects of CNTs with 91% and 97% of purity were compared and changes in the growth behaviour of cells after 3 days in culture with modified medium have been recorded, considering also the distribution of CNTs within cells. While lower purified CNTs induced a slight cytotoxic effect, homogeneously suspended CNTs with high purity were less cytotoxic, and the rate of cell growth remained constant. CNTs aggregated in bundles, showed high adhesion on cell membrane. Interestingly, CNTs bundles were observed inside cells, underneath the cell membrane, and despite of that, cells were extended, in good vitality conditions and no cell-degeneration was observed.

  10. Isoforms of receptors of fibroblast growth factors.

    Science.gov (United States)

    Gong, Siew-Ging

    2014-12-01

    The breadth and scope of Fibroblast Growth Factor signaling is immense, with documentation of its role in almost every organism and system studied so far. FGF ligands signal through a family of four distinct tyrosine kinase receptors, the FGF receptors (FGFRs). One contribution to the diversity of function and signaling of FGFs and their receptors arises from the numerous alternative splicing variants that have been documented in the FGFR literature. The present review discusses the types and roles of alternatively spliced variants of the FGFR family members and the significant impact of alternative splicing on the physiological functions of five broad classes of FGFR isoforms. Some characterized known regulatory mechanisms of alternative splicing and future directions in studies of FGFR alternative splicing are also discussed. Presence, absence, and/or the combination of specific exons within each FGFR protein impart upon each individual isoform its unique function and expression pattern during normal function and in diseased states (e.g., in cancers and birth defects). A better understanding of the diversity of FGF signaling in different developmental contexts and diseased states can be achieved through increased knowledge of the presence of specific FGFR isoforms and their impact on downstream signaling and functions. Modern high-throughput techniques afford an opportunity to explore the distribution and function of isoforms of FGFR during development and in diseases.

  11. Ozone oil promotes wound healing by increasing the migration of fibroblasts via PI3K/Akt/mTOR signaling pathway.

    Science.gov (United States)

    Xiao, Weirong; Tang, Hua; Wu, Meng; Liao, Yangying; Li, Ke; Li, Lan; Xu, Xiaopeng

    2017-09-01

    Background: Skin injury affects millions of people via the uncontrolled inflammation and infection. Many cellular components including fibroblasts and signaling pathways such as TGF-β were activated to facilitate the wound healing to repair injured tissues. Methods and Results: C57BL/6 female mice were divided into control and ozone oil treated groups. Excisional wounds were created on the dorsal skin and the fibroblasts were isolated from granulation tissues. The skin injured mouse model revealed that ozone oil could significantly decrease the wound area and accelerate wound healing compared with control group. QPCR and western blotting assays showed that ozone oil upregulated collagen I, α-SMA and TGF-β1 mRNA and protein levels in fibroblasts. Wound healing assay demonstrated that ozone oil could increase the migration of fibroblasts. Western blotting assay demonstrated that ozone oil increased the EMT process of fibroblasts via upregulating fibronectin, vimentin, N-cadherin, MMP-2, MMP-9, IGFBP-3, IGFBP5 and IGFBP6 and decreasing epithelial protein E-cadherin and cellular senescence marker p16 expression. Mechanistically, western blotting assay revealed that ozone oil increased the phosphorylation of PI3K, Akt and mTOR to regulate the EMT process, while inhibition of PI3K reversed this effect of ozone oil. At last, the results from Cytometric Bead Array demonstrated ozone oil significantly decreased the inflammation in fibroblasts. Conclusion: Our results demonstrated ozone oil facilitated the wound healing via increasing fibroblast migration and EMT process via PI3K/Akt/mTOR signaling pathway in vivo and vitro. The cellular and molecular mechanisms we found here may provide new therapeutic targets for the treatment of skin injury. ©2017 The Author(s).

  12. Fibroblast nemosis induces angiogenic responses of endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Enzerink, Anna, E-mail: anna.enzerink@helsinki.fi [Haartman Institute, University of Helsinki, P.O. BOX 21, FIN-00014 Helsinki (Finland); Rantanen, Ville, E-mail: ville.rantanen@helsinki.fi [Computational Systems Biology Laboratory, Institute of Biomedicine and Genome-Scale Biology Research Program, University of Helsinki, P.O. BOX 63, 00014 Helsinki (Finland); Vaheri, Antti, E-mail: antti.vaheri@helsinki.fi [Haartman Institute, University of Helsinki, P.O. BOX 21, FIN-00014 Helsinki (Finland)

    2010-03-10

    Increasing evidence points to a central link between inflammation and activation of the stroma, especially of fibroblasts therein. However, the mechanisms leading to such activation mostly remain undescribed. We have previously characterized a novel type of fibroblast activation (nemosis) where clustered fibroblasts upregulated the production of cyclooxygenase-2, secretion of prostaglandins, proteinases, chemotactic cytokines, and hepatocyte growth factor (HGF), and displayed activated nuclear factor-{kappa}B. Now we show that nemosis drives angiogenic responses of endothelial cells. In addition to HGF, nemotic fibroblasts secreted vascular endothelial growth factor (VEGF), and conditioned medium from spheroids promoted sprouting and networking of human umbilical venous endothelial cells (HUVEC). The response was partly inhibited by function-blocking antibodies against HGF and VEGF. Conditioned nemotic fibroblast medium promoted closure of HUVEC and human dermal microvascular endothelial cell monolayer wounds, by increasing the motility of the endothelial cells. Wound closure in HUVEC cells was partly inhibited by the antibodies against HGF. The stromal microenvironment regulates wound healing responses and often promotes tumorigenesis. Nemosis offers clues to the activation process of stromal fibroblasts and provides a model to study the part they play in angiogenesis-related conditions, as well as possibilities for therapeutical approaches desiring angiogenesis in tissue.

  13. Conversion of Fibroblasts to Neural Cells by p53 Depletion

    Directory of Open Access Journals (Sweden)

    Di Zhou

    2014-12-01

    Full Text Available Conversion from fibroblasts to neurons has recently been successfully induced. However, the underlying mechanisms are poorly understood. Here, we find that depletion of p53 alone converts fibroblasts into all three major neural lineages. The induced neuronal cells express multiple neuron-specific proteins and generate action potentials and transmitter-receptor-mediated currents. Surprisingly, depletion does not affect the well-known tumorigenic p53 target, p21. Instead, knockdown of p53 upregulates neurogenic transcription factors, which in turn boosts fibroblast-neuron conversion. p53 binds the promoter of the neurogenic transcription factor Neurod2 and regulates its expression during fibroblast-neuron conversion. Furthermore, our method provides a high efficiency of conversion in late-passage fibroblasts. Genome-wide transcriptional analysis shows that the p53-deficiency-induced neurons exhibit an expression profile different from parental fibroblasts and similar to control-induced neurons. The results may help to understand and improve neural conversion mechanisms to develop robust neuron-replacement therapy strategies.

  14. Dogs cloned from fetal fibroblasts by nuclear transfer.

    Science.gov (United States)

    Hong, So Gun; Jang, Goo; Kim, Min Kyu; Oh, Hyun Ju; Park, Jung Eun; Kang, Jung Taek; Koo, Ok Jae; Kim, Dae Yong; Lee, Byeong Chun

    2009-10-01

    Fetal fibroblasts have been considered as the prime candidate donor cells for the canine reproductive cloning by somatic cell nuclear transfer (SCNT) in regard to the future production of transgenic dogs, mainly due to their higher developmental competence and handling advantage in gene targeting. In this study, the cloning efficiency with canine fetal fibroblasts as donor cells was determined. A total of 50 presumptive cloned embryos were reconstructed, activated and transferred into the oviducts of naturally synchronous recipient bitches. While the fusion rate (76.9%) was similar to those of our earlier studies with adult fibroblasts as donor cells (73.9-77.1%), a high cloning efficiency (4.0%; 2 births/50 embryos transferred) was found compared to the previous success rate with adult fibroblasts (0.2-1.8%). The cloned beagles were healthy and genotypically identical to the donor fibroblast cells. This study shows that a fetal fibroblast cell would be an excellent donor for future production of transgenic dogs via gene targeting in this cell followed cloning using SCNT technology.

  15. Autophagy is required for IL-2-mediated fibroblast growth

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Rui [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Tang, Daolin, E-mail: tangd2@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Lotze, Michael T., E-mail: lotzemt@upcm.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States); Zeh III, Herbert J., E-mail: zehh@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15219 (United States)

    2013-02-15

    Autophagy is an evolutionarily conserved pathway responsible for delivery of cytoplasmic material into the lysosomal degradation pathway to enable vesicular exocytosis. Interleukin (IL)-2 is produced by T-cells and its activity is important for immunoregulation. Fibroblasts are an immune competent cell type, playing a critical role in wound healing, chronic inflammation, and tumor development. Although autophagy plays an important role in each of these processes, whether it regulates IL-2 activity in fibroblasts is unknown. Here, we show that autophagy is required for IL-2-induced cell growth in fibroblasts. IL-2 significantly induced autophagy in mouse embryonic fibroblasts (MEFs) and primary lung fibroblasts. Autophagy inhibitors (e.g., 3-methylamphetamine and bafilomycin A1) or knockdown of ATG5 and beclin 1 blocked clinical grade IL-2-induced autophagy. Moreover, IL-2 induced HMGB1 cytoplasmic translocation in MEFs and promoted interaction between HMGB1 and beclin1, which is required for autophagy induction. Pharmacological and genetic inhibition of autophagy inhibited IL-2-induced cell proliferation and enhanced IL-2-induced apoptosis. These findings suggest that autophagy is an important pro-survival regulator for IL-2-induced cell growth in fibroblasts.

  16. Periostin induces fibroblast proliferation and myofibroblast persistence in hypertrophic scarring.

    Science.gov (United States)

    Crawford, Justin; Nygard, Karen; Gan, Bing Siang; O'Gorman, David Brian

    2015-02-01

    Hypertrophic scarring is characterized by the excessive development and persistence of myofibroblasts. These cells contract the surrounding extracellular matrix resulting in the increased tissue density characteristic of scar tissue. Periostin is a matricellular protein that is abnormally abundant in fibrotic dermis, however, its roles in hypertrophic scarring are largely unknown. In this report, we assessed the ability of matrix-associated periostin to promote the proliferation and myofibroblast differentiation of dermal fibroblasts isolated from the dermis of hypertrophic scars or healthy skin. Supplementation of a thin type-I collagen cell culture substrate with recombinant periostin induced a significant increase in the proliferation of hypertrophic scar fibroblasts but not normal dermal fibroblasts. Periostin induced significant increases in supermature focal adhesion formation, α smooth muscle actin levels and collagen contraction in fibroblasts cultured from hypertrophic scars under conditions of increased matrix tension in three-dimensional type-I collagen lattices. Inhibition of Rho-associated protein kinase activity significantly attenuated the effects of matrix-associated periostin on hypertrophic scar fibroblasts and myofibroblasts. Depletion of endogenous periostin expression in hypertrophic scar myofibroblasts resulted in a sustained decrease in α smooth muscle actin levels under conditions of reducing matrix tension, while matrix-associated periostin levels caused the cells to retain high levels of a smooth muscle actin under these conditions. These findings indicate that periostin promotes Rho-associated protein kinase-dependent proliferation and myofibroblast persistence of hypertrophic scar fibroblasts and implicate periostin as a potential therapeutic target to enhance the resolution of scars.

  17. Fibroblast-specific upregulation of Flightless I impairs wound healing.

    Science.gov (United States)

    Turner, Christopher T; Waters, James M; Jackson, Jessica E; Arkell, Ruth M; Cowin, Allison J

    2015-09-01

    The cytoskeletal protein Flightless (Flii) is a negative regulator of wound healing. Upregulation of Flii is associated with impaired migration, proliferation and adhesion of both fibroblasts and keratinocytes. Importantly, Flii translocates from the cytoplasm to the nucleus in response to wounding in fibroblasts but not keratinocytes. This cell-specific nuclear translocation of Flii suggests that Flii may directly regulate gene expression in fibroblasts, providing one potential mechanism of action for Flii in the wound healing response. To determine whether the tissue-specific upregulation of Flii in fibroblasts was important for the observed inhibitory effects of Flii on wound healing, an inducible fibroblast-specific Flii overexpressing mouse model was generated. The inducible ROSA26 system allowed the overexpression of Flii in a temporal and tissue-specific manner in response to tamoxifen treatment. Wound healing in the inducible mice was impaired, with wounds at day 7 postwounding significantly larger than those from non-inducible controls. There was also reduced collagen maturation, increased myofibroblast infiltration and elevated inflammation. The impaired healing response was similar in magnitude to that observed in mice with non-tissue-specific upregulation of Flii suggesting that fibroblast-derived Flii may have an important role in the wound healing response. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. Ceramide-induced apoptosis in rabbit corneal fibroblasts.

    Science.gov (United States)

    Kim, Tae-im; Pak, Jhang Ho; Tchah, Hungwon; Lee, Seung-ah; Kook, Michael S

    2005-01-01

    To evaluate the effect of various ceramides on the apoptosis of corneal fibroblasts and to determine the pathway on which they act. Corneal fibroblasts isolated and cultured from New Zealand white rabbits were exposed to various concentrations of ceramide types II and VI and phytoceramide types II and VI, and their apoptotic response was evaluated using an LDH assay and Hoechst and Annexin V staining. Corneal fibroblasts were preincubated with various concentrations of the CPP32-like protease inhibitor Z-VAD-FMK, the caspase-8 inhibitor IETD-CHO, and the caspase-9 inhibitor Z-LEHD-FMK before treatment with ceramide, and apoptotic response was assayed by LDH assay. In addition, cells treated with ceramide or phytoceramide were stained with an antibody to cytochrome c. At concentrations of 20 microM and higher, all 4 ceramides increased fibroblast apoptotic response significantly after 12 hours. Hoechst staining showed shrinkage of the cytoplasm, formation of apoptotic bodies, and nuclear fragmentation after ceramide exposure, and Annexin V staining showed small vesicles around the cell membrane. The CPP32-like protease inhibitor reduced the apoptotic response to all 4 ceramides. The specific caspase-8 inhibitor reduced the apoptotic response to ceramide type VI and phytoceramide types II and VI, whereas the specific caspase-9 inhibitor significantly reduced the apoptotic response to phytoceramide types II and VI. Following exposure to ceramides, corneal fibroblasts stained positively with antibody to cytochrome c. Ceramide induced apoptosis in cultured corneal fibroblasts. This apoptosis involved the caspase cascade and the mitochondrial pathway.

  19. Ciprofloxacin Decreases Collagen in Mouse Tympanic Membrane Fibroblasts.

    Science.gov (United States)

    Orobello, Nicklas C; Dirain, Carolyn O; Schultz, Gregory; Milne-Davies, Bailey A; Ng, Maria R A; Antonelli, Patrick J

    2016-07-01

    To determine how collagen production by tympanic membrane fibroblasts is affected by ciprofloxacin at levels found in eardrops. Prospective, controlled, and blinded cell culture study. Academic tertiary medical center. Cell culture of mouse fibroblasts. A primary fibroblast culture was established from mouse tympanic membranes. Fibroblasts were cultured until they were 75% confluent, then treated with dilute hydrochloric acid (control) or ciprofloxacin (0.01% or 0.3%) for 24 or 72 hours for Western blotting and for 24 or 48 hours for cytotoxicity assay. Cells were observed with phase-contrast microscope. Western blotting was performed for collagen type 1 α1 (collagen 1A1) and α-tubulin. Fibroblasts treated with 0.01% and 0.3% ciprofloxacin for 24 hours had lower levels of collagen 1A1 (P = .0005 and P ciprofloxacin (P = .02 and P = .014). After 72 hours, 0.3% ciprofloxacin completely eliminated collagen 1A1 and α-tubulin (P ciprofloxacin for 72 hours also had lower collagen 1A1 (P ciprofloxacin resulted in lower levels of collagen 1A1 (P = .009 and P ciprofloxacin, as found in eardrops, reduces fibroblast viability and collagen and α-tubulin protein levels. These findings could explain tympanic membrane healing problems associated with quinolone eardrops. © American Academy of Otolaryngology—Head and Neck Surgery Foundation 2016.

  20. Roles of dental pulp fibroblasts in the recognition of bacterium-related factors and subsequent development of pulpitis

    Directory of Open Access Journals (Sweden)

    Tadashi Nakanishi

    2011-08-01

    Full Text Available As caries-related bacteria invade deeply into dentin and come into close proximity to the pulp, inflammatory cells (such as lymphocytes, macrophages and neutrophils infiltrate into the bacterium-invaded area and consequently pulpitis develops. Many types of cytokines and adhesion molecules are responsible for the initiation and progression of pulpitis. Dental pulp fibroblasts, a major cell type in the dental pulp, also have capacity to produce pro-inflammatory cytokines and express adhesion molecules in response to pathogen-associated molecular patterns (PAMPs, including lipopolysaccharide. The innate immune system senses microbial infection using pattern recognition receptors, such as Toll-like receptors (TLRs and nucleotide-binding oligomerization domain (NOD, for PAMPs. In this review, we summarize the roles of dental pulp fibroblasts in the recognition of invaded bacterium-related factors via TLR and NOD pathways, and the subsequent pulpal immune responses, leading to progressive pulpitis.

  1. Comparative analysis of the expression of surface markers on fibroblasts and fibroblast-like cells isolated from different human tissues.

    Science.gov (United States)

    Lupatov, A Yu; Vdovin, A S; Vakhrushev, I V; Poltavtseva, R A; Yarygin, K N

    2015-02-01

    Expression of 20 surface markers was analyzed in cultures of mesenchymal stromal cells of the umbilical cord, fibroblasts from adult and fetal human skin, and fibroblast-like cells of fetal liver was analyzed by fl ow cytometry. The studied cultures did not express hemopoietic cells markers, but were positive for CD73, CD90, and CD105 markers recommended by the International Society of Cell Therapy for the identification of the multipotent mesenchymal stromal cells. Fetal liver fibroblast-like cells were positive for CD54; this marker was absent in skin fibroblast cultures, but was expressed by umbilical cord mesenchymal stromal cells. Further study of these cells revealed a minor subpopulation of cells co-expressing CD24 and CD90 or CD24 and CD54. We hypothesized that these cells probably participate in epithelial mesenchymal transition.

  2. Chitosan/TiO2 composite membrane improves proliferation and survival of L929 fibroblast cells: Application in wound dressing and skin regeneration.

    Science.gov (United States)

    Behera, Sudhanshu Shekhar; Das, Urmimala; Kumar, Awanish; Bissoyi, Akalabya; Singh, Abhishek Kumar

    2017-05-01

    The microbial infection and growth of fibroblasts are the critical factors for the effective wound healing. The natural polymer-based dressing membranes may mimic extracellular matrix to support the survival, proliferation and differentiation of fibroblasts. The present study deals with the preparation of chitosan/titanium dioxide (CS/TiO2) composite membranes with different degree of TiO2 incorporation, and their characterization in terms of morphology, ultrastructure, thermal behavior and mechanical properties with SEM, FTIR, XRD and tensile strength analyses. The data demonstrated the formation of strong O-Ti-O bonding between TiO2 and CS resulting in superior porosity, mechanical strength, crystallinity and flexibility of the composite membranes. Further, the cyto-compatibility, proliferation, oxidative stress, cell cycle and apoptosis analyses of fibroblast L929 cells demonstrated the enhanced proliferation and survival, and decreased oxidative stress and apoptosis in L929 cells grown on CS/TiO2 membrane incorporated with 025% TiO2. Next, we measured the significant up-regulation in the expression of fibroblast-markers in L929 cells cultured on CS/TiO2 (0.25%) membrane. Furthermore, the CS/TiO2 composite membranes exhibited a superior antibacterial activity against Staphylococcus aureus. Taken together, the data confirmed that CS/TiO2 (0.25%) membrane improved the growth, survival and functional integrity of fibroblasts, and exerted antibacterial activity which may be utilized as potential dressing materials.

  3. Quiescent and proliferative fibroblasts exhibit differential p300 HAT activation through control of 5-methoxytryptophan production.

    Directory of Open Access Journals (Sweden)

    Huei-Hsuan Cheng

    Full Text Available Quiescent fibroblasts possess unique genetic program and exhibit high metabolic activity distinct from proliferative fibroblasts. In response to inflammatory stimulation, quiescent fibroblasts are more active in expressing cyclooxygenase-2 and other proinflammatory genes than proliferative fibroblasts. The underlying transcriptional mechanism is unclear. Here we show that phorbol 12-myristate 13-acetate (PMA and cytokines increased p300 histone acetyltransferase activity to a higher magnitude (> 2 fold in quiescent fibroblasts than in proliferative fibroblasts. Binding of p300 to cyclooxygenase-2 promoter was reduced in proliferative fibroblasts. By ultrahigh-performance liquid chromatography coupled with a quadrupole time of flight mass spectrometer and enzyme-immunoassay, we found that production of 5-methoxytryptophan was 2-3 folds higher in proliferative fibroblasts than that in quiescent fibroblasts. Addition of 5-methoxytryptophan and its metabolic precursor, 5-hydroxytryptophan, to quiescent fibroblasts suppressed PMA-induced p300 histone acetyltransferase activity and cyclooxygenase-2 expression to the level of proliferative fibroblasts. Silencing of tryptophan hydroxylase-1 or hydroxyindole O-methyltransferase in proliferative fibroblasts with siRNA resulted in elevation of PMA-induced p300 histone acetyltransferase activity to the level of that in quiescent fibroblasts, which was rescued by addition of 5-hydroxytryptophan or 5-methoxytryptophan. Our findings indicate that robust inflammatory gene expression in quiescent fibroblasts vs. proliferative fibroblasts is attributed to uncontrolled p300 histone acetyltransferase activation due to deficiency of 5-methoxytryptophan production. 5-methoxytryptophan thus is a potential valuable lead compound for new anti-inflammatory drug development.

  4. Agglutination of Trypanosoma cruzi in infected cells treated with serum from chronically infected mice.

    Science.gov (United States)

    Wendelken, Jennifer L; Rowland, Edwin C

    2009-04-01

    The protozoan parasite Trypanosoma cruzi is the causative agent of Chagas disease. The chronic stage of infection is characterized by a production of neutralizing antibodies in the vertebrate host. A polyclonal antibody, anti-egressin, has been found to inhibit egress of parasites from the host cell late in the intracellular cycle, after the parasites have transformed from the replicative amastigote into the trypomastigote. It has also been found that BALB/c mouse fibroblasts in the late stages of parasite infection become permeable to molecules as large as antibodies, leading to the possibility that anti-egressin affects the intracellular parasites. This project addresses the fate of the intracellular trypomastigotes that have been inhibited from egressing the host cell. Extended cultures of infected fibroblasts treated with chronic mouse serum reduced parasite egress at all time points measured. Parasites released from infected fibroblasts treated with chronic serum had a reduced ability to infect fibroblasts in culture, yet did not lose infectivity entirely. Absorption of chronic serum with living trypomastigotes removed the anti-egressin effect. The possibility that the target of anti-egressin is a parasite surface component is further indicated by the agglutination of extracellular trypomastigotes by chronic serum. The possibility that cross-linking by antibody occurs intracellularly, thus inhibiting egress, was reinforced by cleaving purified IgG into Fab fragments, which did not inhibit egress when added to infected cultures. From this work, it is proposed that the current, best explanation of the mechanism of egress inhibition by anti-egressin is intracellular agglutination, preventing normal parasite-driven egress.

  5. Super-telomeres in transformed human fibroblasts.

    Science.gov (United States)

    Chiodi, Ilaria; Belgiovine, Cristina; Zongaro, Samantha; Ricotti, Roberta; Horard, Beatrice; Lossani, Andrea; Focher, Federico; Gilson, Eric; Giulotto, Elena; Mondello, Chiara

    2013-08-01

    Telomere length maintenance is critical for organisms' long-term survival and cancer cell proliferation. Telomeres are kept within species-specific length ranges by the interplay between telomerase activity and telomeric chromatin organization. In this paper, we exploited telomerase immortalized human fibroblasts (cen3tel) that gradually underwent neoplastic transformation during culture propagation to study telomere composition and length regulation during the transformation process. Just after telomerase catalytic subunit (hTERT) expression, cen3tel telomeres shortened despite the presence of telomerase activity. At a later stage and concomitantly with transformation, cells started elongating telomeres, which reached a mean length greater than 100kb in about 900 population doublings. Super-telomeres were stable and compatible with cell growth and tumorigenesis. Telomere extension was associated with increasing levels of telomerase activity that were linked to the deregulation of endogenous telomerase RNA (hTERC) and exogenous telomerase reverse transcriptase (hTERT) expression. Notably, the increase in hTERC levels paralleled the increase in telomerase activity, suggesting that this subunit plays a role in regulating enzyme activity. Telomeres ranging in length between 10 and more than 100kb were maintained in an extendible state although TRF1 and TRF2 binding increased with telomere length. Super-telomeres neither influenced subtelomeric region global methylation nor the expression of the subtelomeric gene FRG1, attesting the lack of a clear-cut relationship between telomere length, subtelomeric DNA methylation and expression in human cells. The cellular levels of the telomeric proteins hTERT, TRF1, TRF2 and Hsp90 rose with transformation and were independent of telomere length, pointing to a role of these proteins in tumorigenesis.

  6. 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

  7. Fibroblast growth factor signaling in metabolic regulation

    Directory of Open Access Journals (Sweden)

    Vera eNies

    2016-01-01

    Full Text Available The prevalence of obesity is a growing health problem. Obesity is strongly associated with several comorbidities, such as non-alcoholic fatty liver disease, certain cancers, insulin resistance and type 2 diabetes, which all reduce life expectancy and life quality. Several drugs have been put forward in order to treat these diseases, but many of them have detrimental side effects. The unexpected role of the family of fibroblast growth factors in the regulation of energy metabolism provides new approaches to the treatment of metabolic diseases, and offers a valuable tool to gain more insight into metabolic regulation. The known beneficial effects of FGF19 and FGF21 on metabolism, together with recently discovered similar effects of FGF1 suggest that FGFs and their derivatives carry great potential as novel therapeutics to treat metabolic conditions. To facilitate the development of new therapies with improved targeting and minimal side effects, a better understanding of the molecular mechanism of action of FGFs is needed.In this review we will discuss what is currently known about the physiological roles of FGF signaling in tissues important for metabolic homeostasis. In addition, we will discuss current concepts regarding their pharmacological properties and effector tissues in the context of metabolic disease. Also the recent progress in the development of FGF variants will be reviewed. Our goal is to provide a comprehensive overview of the current concepts and consensuses regarding FGF signaling in metabolic health and disease, and to provide starting points for the development of FGF-based therapies against metabolic conditions.

  8. Cellular signaling by fibroblast growth factor receptors.

    Science.gov (United States)

    Eswarakumar, V P; Lax, I; Schlessinger, J

    2005-04-01

    The 22 members of the fibroblast growth factor (FGF) family of growth factors mediate their cellular responses by binding to and activating the different isoforms encoded by the four receptor tyrosine kinases (RTKs) designated FGFR1, FGFR2, FGFR3 and FGFR4. Unlike other growth factors, FGFs act in concert with heparin or heparan sulfate proteoglycan (HSPG) to activate FGFRs and to induce the pleiotropic responses that lead to the variety of cellular responses induced by this large family of growth factors. A variety of human skeletal dysplasias have been linked to specific point mutations in FGFR1, FGFR2 and FGFR3 leading to severe impairment in cranial, digital and skeletal development. Gain of function mutations in FGFRs were also identified in a variety of human cancers such as myeloproliferative syndromes, lymphomas, prostate and breast cancers as well as other malignant diseases. The binding of FGF and HSPG to the extracellular ligand domain of FGFR induces receptor dimerization, activation and autophosphorylation of multiple tyrosine residues in the cytoplasmic domain of the receptor molecule. A variety of signaling proteins are phosphorylated in response to FGF stimulation including Shc, phospholipase-Cgamma, STAT1, Gab1 and FRS2alpha leading to stimulation of intracellular signaling pathways that control cell proliferation, cell differentiation, cell migration, cell survival and cell shape. The docking proteins FRS2alpha and FRS2beta are major mediators of the Ras/MAPK and PI-3 kinase/Akt signaling pathways as well as negative feedback mechanisms that fine-tune the signal that is initiated at the cell surface following FGFR stimulation.

  9. Autophagy in periodontitis patients and gingival fibroblasts: unraveling the link between chronic diseases and inflammation

    Directory of Open Access Journals (Sweden)

    Bullon Pedro

    2012-10-01

    Full Text Available Abstract Background Periodontitis, the most prevalent chronic inflammatory disease, has been related to cardiovascular diseases. Autophagy provides a mechanism for the turnover of cellular organelles and proteins through a lysosome-dependent degradation pathway. The aim of this research was to study the role of autophagy in peripheral blood mononuclear cells from patients with periodontitis and gingival fibroblasts treated with a lipopolysaccharide of Porphyromonas gingivalis. Autophagy-dependent mechanisms have been proposed in the pathogenesis of inflammatory disorders and in other diseases related to periodontitis, such as cardiovascular disease and diabetes. Thus it is important to study the role of autophagy in the pathophysiology of periodontitis. Methods Peripheral blood mononuclear cells from patients with periodontitis (n = 38 and without periodontitis (n = 20 were used to study autophagy. To investigate the mechanism of autophagy, we evaluated the influence of a lipopolysaccharide from P. gingivalis in human gingival fibroblasts, and autophagy was monitored morphologically and biochemically. Autophagosomes were observed by immunofluorescence and electron microscopy. Results We found increased levels of autophagy gene expression and high levels of mitochondrial reactive oxygen species production in peripheral blood mononuclear cells from patients with periodontitis compared with controls. A significantly positive correlation between both was observed. In human gingival fibroblasts treated with lipopolysaccharide from P. gingivalis, there was an increase of protein and transcript of autophagy-related protein 12 (ATG12 and microtubule-associated protein 1 light chain 3 alpha LC3. A reduction of mitochondrial reactive oxygen species induced a decrease in autophagy whereas inhibition of autophagy in infected cells increased apoptosis, showing the protective role of autophagy. Conclusion Results from the present study suggest that autophagy

  10. Central line infections - hospitals

    Science.gov (United States)

    ... infection; Central venous catheter - infection; CVC - infection; Central venous device - infection; Infection control - central line infection; Nosocomial infection - central line infection; Hospital acquired ...

  11. Pirfenidone suppresses keloid fibroblast-embedded collagen gel contraction.

    Science.gov (United States)

    Saito, Masuyoshi; Yamazaki, Masashi; Maeda, Tatsuo; Matsumura, Hajime; Setoguchi, Yasuhiro; Tsuboi, Ryoji

    2012-04-01

    Keloid is a clinically intractable disease that causes disfigurement, itching, and pain due to abnormal proliferation of fibroblasts and production of collagen. Pirfenidone is a novel anti-fibrotic agent that inhibits the progression of fibrosis occurring in the keloid lesions of the lung and kidney. In order to examine whether pirfenidone has a therapeutic effect on keloid lesions, we prepared an in vitro wound contraction model with keloid fibroblasts. The gel contractility of a mixture of keloid fibroblasts and an acid-soluble collagen solution was examined with/without transforming growth factor (TGF)-β1 in the presence or absence of pirfenidone. Real time RT-PCR was performed to detect mRNA expression of TGFB1, CTGF, aSMA, and Col1A1 quantitatively in keloid fibroblasts incubated with/without TGF-β1 in the presence or absence of pirfenidone. The contractility of keloid fibroblast-embedded collagen gel was increased after the addition of TGF-β1. Pirfenidone suppressed gel contraction with TGF-β1 dose dependently. TGF-β1 stimulated mRNA expression of TGFB1, CTGF, aSMA, and Col1A1 in keloid fibroblasts, while pirfenidone significantly inhibited mRNA expression of CTGF and aSMA in the identical cells. These findings suggest that pirfenidone suppresses the contraction of keloid-derived fibroblasts by inhibiting the down-stream pathway of TGF-β1, thus demonstrating its therapeutic utility for the treatment of keloid lesions.

  12. Adiponectin Promotes Monocyte-to-Fibroblast Transition in Renal Fibrosis

    Science.gov (United States)

    Yang, Jun; Lin, Song-Chang; Chen, Gang; He, Liqun; Hu, Zhaoyong; Chan, Lawrence; Trial, JoAnn; Entman, Mark L.

    2013-01-01

    Bone marrow–derived fibroblasts may contribute substantially to the pathogenesis of renal fibrosis through the excessive production and deposition of extracellular matrix. However, the mechanisms underlying the accumulation and activation of these fibroblasts are not understood. Here, we used a mouse model of tubulointerstitial fibrosis to determine whether adiponectin, which is elevated in CKD and is associated with disease progression, regulates monocyte-to-fibroblast transition and fibroblast activation in injured kidneys. In wild-type mice, the expression of adiponectin and the number of bone marrow–derived fibroblasts in the kidney increased after renal obstruction. In contrast, the obstructed kidneys of adiponectin-knockout mice had fewer bone marrow–derived fibroblasts. Adiponectin deficiency also led to a reduction in the number of myofibroblasts, the expression of profibrotic chemokines and cytokines, and the number of procollagen-expressing M2 macrophages in injured kidneys. Consistent with these findings, adiponectin-deficiency reduced the expression of collagen I and fibronectin. Similar results were observed in wild-type and adiponectin-knockout mice after ischemia-reperfusion injury. In cultured bone marrow–derived monocytes, adiponectin stimulated the expression of α-smooth muscle actin (SMA) and extracellular matrix proteins and activated AMP-activated protein kinase (AMPK) in a time- and dose-dependent manner. Furthermore, specific activation of AMPK increased the expression of α-SMA and extracellular matrix proteins, while inhibition of AMPK attenuated these responses. Taken together, these findings identify adiponectin as a critical regulator of monocyte-to-fibroblast transition and renal fibrosis, suggesting that inhibition of adiponectin/AMPK signaling may represent a novel therapeutic target for fibrotic kidney disease. PMID:23833260

  13. Binding, uptake, and release of nicotine by human gingival fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Hanes, P.J.; Schuster, G.S.; Lubas, S. (Medical College of Georgia, Augusta (USA))

    1991-02-01

    Previous studies of the effects of nicotine on fibroblasts have reported an altered morphology and attachment of fibroblasts to substrates and disturbances in protein synthesis and secretion. This altered functional and attachment response may be associated with changes in the cell membrane resulting from binding of the nicotine, or to disturbances in cell metabolism as a result of high intracellular levels of nicotine. The purpose of the present study, therefore, was to (1) determine whether gingival fibroblasts bound nicotine and if any binding observed was specific or non-specific in nature; (2) determine whether gingival fibroblasts internalized nicotine, and if so, at what rate; (3) determine whether gingival fibroblasts also released nicotine back into the extracellular environment; and (4) if gingival fibroblasts release nicotine intact or as a metabolite. Cultures of gingival fibroblasts were prepared from gingival connective tissue biopsies. Binding was evaluated at 4{degree}C using a mixture of {sup 3}H-nicotine and unlabeled nicotine. Specific binding was calculated as the difference between {sup 3}H-nicotine bound in the presence and absence of unlabeled nicotine. The cells bound 1.44 (+/- 0.42) pmols/10(6) cells in the presence of unlabeled nicotine and 1.66 (+/- 0.55) pmols/10(6) cells in the absence of unlabeled nicotine. The difference was not significant. Uptake of nicotine was measured at 37{degree}C after treating cells with {sup 3}H-nicotine for time periods up to 4 hours. Uptake in pmols/10(6) cells was 4.90 (+/- 0.34) at 15 minutes, 8.30 (+/- 0.75) at 30 minutes, 12.28 (+/- 2.62) at 1 hour and 26.31 (+/- 1.15) at 4 hours.

  14. Functional screen of paracrine signals in breast carcinoma fibroblasts.

    Directory of Open Access Journals (Sweden)

    Gui Su

    Full Text Available Stromal fibroblasts actively participate in normal mammary gland homeostasis and in breast carcinoma growth and progression by secreting paracrine factors; however, little is known about the identity of paracrine mediators in individual patients. The purpose of this study was to characterize paracrine signaling pathways between breast carcinoma cells and breast carcinoma-associated fibroblasts (CAF or normal mammary fibroblasts (NF, respectively. CAF and NF were isolated from breast carcinoma tissue samples and adjacent normal mammary gland tissue of 28 patients. The fibroblasts were grown in 3D collagen gel co-culture with T47D human breast carcinoma cells and T47D cell growth was measured. CAF stimulated T47D cell growth to a significantly greater degree than NF. We detected a considerable inter-individual heterogeneity of paracrine interactions but identified FGF2, HB-EGF, heparanase-1 and SDF1 as factors that were consistently responsible for the activity of carcinoma-associated fibroblasts. CAF from low-grade but not high-grade carcinomas required insulin-like growth factor 1 and transforming growth factor beta 1 to stimulate carcinoma growth. Paradoxically, blocking of membrane-type 1 matrix metalloprotease stimulated T47D cell growth in co-culture with NF. The results were largely mirrored by treating the fibroblasts with siRNA oligonucleotides prior to co-culture, implicating the fibroblasts as principal production site for the secreted mediators. In summary, we identify a paracrine signaling network with inter-individual commonalities and differences. These findings have significant implications for the design of stroma-targeted therapies.

  15. Homogeneous Inflammatory Gene Profiles Induced in Human Dermal Fibroblasts in Response to the Three Main Species of Borrelia burgdorferi sensu lato

    Science.gov (United States)

    Meddeb, Mariam; Carpentier, Wassila; Cagnard, Nicolas; Nadaud, Sophie; Grillon, Antoine; Barthel, Cathy; De Martino, Sylvie Josiane; Jaulhac, Benoît; Boulanger, Nathalie

    2016-01-01

    In Lyme borreliosis, the skin is the key site for bacterial inoculation by the infected tick and for cutaneous manifestations. We previously showed that different strains of Borrelia burgdorferi sensu stricto isolated from tick and from different clinical stages of the Lyme borreliosis (erythema migrans, and acrodermatitis chronica atrophicans) elicited a very similar transcriptional response in normal human dermal fibroblasts. In this study, using whole transcriptome microarray chips, we aimed to compare the transcriptional response of normal human dermal fibroblasts stimulated by 3 Borrelia burgdorferi sensu lato strains belonging to 3 main pathogenic species (B. afzelii, B. garinii and B. burgdorferi sensu stricto) in order to determine whether “species-related” inflammatory pathways could be identified. The three Borrelia strains tested exhibited similar transcriptional profiles, and no species-specific fingerprint of transcriptional changes in fibroblasts was observed. Conversely, a common core of chemokines/cytokines (CCL2, CXCL1, CXCL2, CXCL6, CXCL10, IL-6, IL-8) and interferon-related genes was stimulated by all the 3 strains. Dermal fibroblasts appear to play a key role in the cutaneous infection with Borrelia, inducing a homogeneous inflammatory response, whichever Borrelia species was involved. PMID:27706261

  16. Inhibition of growth of Toxoplasma gondii in cultured fibroblasts by human recombinant gamma interferon.

    Science.gov (United States)

    Pfefferkorn, E R; Guyre, P M

    1984-01-01

    The growth of Toxoplasma gondii in cultured human fibroblasts was inhibited by recombinant human gamma interferon at concentrations of 8 to 16 U/ml. The interferon was titrated by observing a total inhibition of parasite plaque formation 7 days after infection. Inhibition of the growth of T. gondii in the early days after infection was measured by marked reductions in the incorporation of radioactive uracil, a precursor that can only be used by the parasites. This assay showed that when cells were pretreated with gamma interferon for 1 day and then infected, inhibition of T. gondii growth could be readily detected 1 or 2 days after infection. When the pretreatment was omitted and parasites and gamma interferon were added at the same time, no inhibition of parasite growth could be detected 1 day later, although it was apparent after 2 days. Cultures from which the gamma interferon had been removed by washing after a 1-day treatment showed inhibition of T. gondii growth. Gamma interferon had no effect on the viability of extracellular parasites, but it did inhibit the synthesis of host cell RNA and protein by ca. 50% 3 days after treatment. This degree of inhibition is unlikely, of itself, to compromise the growth of T. gondii. Recombinant alpha and beta interferons had no effect on the growth of T. gondii. Images PMID:6425215

  17. Hookworm infection

    Science.gov (United States)

    Hookworm disease; Ground itch; Ancylostoma duodenale infection; Necator americanus infection; Parasitic infection - hookworm ... The infection is caused by infestation with any of the following ... Ancylostoma duodenale Ancylostoma ceylanicum Ancylostoma ...

  18. Kidney Infection

    Science.gov (United States)

    ... X-ray called a voiding cystourethrogram. Antibiotics for kidney infections Antibiotics are the first line of treatment ... the infection is completely eliminated. Hospitalization for severe kidney infections For a severe kidney infection, your doctor ...

  19. Fibroblastic reticular cells from lymph nodes attenuate T cell expansion by producing nitric oxide.

    Directory of Open Access Journals (Sweden)

    Stefanie Siegert

    Full Text Available Adaptive immune responses are initiated when T cells encounter antigen on dendritic cells (DC in T zones of secondary lymphoid organs. T zones contain a 3-dimensional scaffold of fibroblastic reticular cells (FRC but currently it is unclear how FRC influence T cell activation. Here we report that FRC lines and ex vivo FRC inhibit T cell proliferation but not differentiation. FRC share this feature with fibroblasts from non-lymphoid tissues as well as mesenchymal stromal cells. We identified FRC as strong source of nitric oxide (NO thereby directly dampening T cell expansion as well as reducing the T cell priming capacity of DC. The expression of inducible nitric oxide synthase (iNOS was up-regulated in a subset of FRC by both DC-signals as well as interferon-γ produced by primed CD8+ T cells. Importantly, iNOS expression was induced during viral infection in vivo in both LN FRC and DC. As a consequence, the primary T cell response was found to be exaggerated in Inos(-/- mice. Our findings highlight that in addition to their established positive roles in T cell responses FRC and DC cooperate in a negative feedback loop to attenuate T cell expansion during acute inflammation.

  20. Generation of Integration-free and Region-Specific Neural Progenitors from Primate Fibroblasts

    Directory of Open Access Journals (Sweden)

    Jianfeng Lu

    2013-05-01

    Full Text Available Postnatal and adult human and monkey fibroblasts were infected with Sendai virus containing the Yamanaka factors for 24 hr, then they were cultured in a chemically defined medium containing leukemia inhibitory factor (LIF, transforming growth factor (TGF-β inhibitor SB431542, and glycogen synthase kinase (GSK-3β inhibitor CHIR99021 at 39°C for inactivation of the virus. Induced neural progenitor (iNP colonies appeared as early as day 13 and can be expanded for >20 passages. Under the same defined condition, no induced pluripotent stem cell (iPSC colonies formed at either 37°C or 39°C. The iNPs predominantly express hindbrain genes and differentiate into hindbrain neurons, and when caudalized, they produced an enriched population of spinal motor neurons. Following transplantation into the forebrain, the iNP-derived cells retained the hindbrain identity. The ability to generate defined, integration-free iNPs from adult primate fibroblasts under a defined condition with predictable fate choices will facilitate disease modeling and therapeutic development.

  1. The role of calcineurin in the lung fibroblasts proliferation and collagen synthesis induced by basic fibroblast growth factor

    Institute of Scientific and Technical Information of China (English)

    陈亚红; 赵鸣武; 符民桂; 姚婉贞; 唐朝枢

    2003-01-01

    Objective To investigate the role of calcineurin (CaN) in the lung fibroblast proliferation and collagen synthesis induced by basic fibroblast growth factor (bFGF).Methods We used Western blot and immunohistochemical methods for investigating the content and distribution of calcineurin in the lung tissue. Calcineurin activity in different tissues was measured using 32P-labelled substrate. In the primary culture of lung fibroblasts, 3H-thymidine (3H-TdR) and 3H-proline incorporation methods were used to study the effect of cyclosporin A(CsA), an inhibitor of calcineurin, on the lung fibroblast DNA and collagen synthesis stimulated by bFGF. Results We found that calcineurin was expressed in lung tissue and has phosphatase activity (7.1±2.0 pmol Pi/mg pr/min). CsA(10-8-10-6mol/L) inhibited lung fibroblast,3H-TdR incorporation induced by bFGF in a dose-dependent manner, with the inhibitory rates by20%, 46% and 66%(P<0.01). CsA(10-7-10-6mol/L) inhibited 3H-proline incorporation in lung fibroblasts stimulated by bFGF, with the inhibitory rates by 21% and 37%(P<0.01). In a culture medium, CsA (10-8-10-6mol/L) inhibited 3H-proline secretion induced by bFGF in a dose-dependent manner, with the inhibitory rates by 19%,29%(P<0.05) and 56% (P<0.01). CsA (10-7mol/L) could inhibit calcineurin activity by 44% in lung fibroblasts(P<0.01). Conclusions Calcineurin is expressed in lung tissue and has phosphatase activity. It is involved in the bFGF stimulated lung fibroblast DNA and collagen synthesis.

  2. Modeled Microgravity Affects Fibroblast Functions Related to Wound Healing

    Science.gov (United States)

    Cialdai, Francesca; Vignali, Leonardo; Morbidelli, Lucia; Colciago, Alessandra; Celotti, Fabio; Santi, Alice; Caselli, Anna; Cirri, Paolo; Monici, Monica

    2017-02-01

    Wound healing is crucial for the survival of an organism. Therefore, in the perspective of space exploration missions, it is important to understand if and how microgravity conditions affect the behavior of the cell populations involved in wound healing and the evolution of the process. Since fibroblasts are the major players in tissue repair, this study was focused on the behavior of fibroblasts in microgravity conditions, modeled by a RCCS. Cell cytoskeleton was studied by immunofluorescence microscopy, the ability to migrate was assessed by microchemotaxis and scratch assay, and the expression of markers of fibroblast activation, angiogenesis, and inflammation was assessed by western blot. Results revealed that after cell exposure to modeled microgravity conditions, a thorough rearrangement of microtubules occurred and α-SMA bundles were replaced by a tight network of faulty and disorganized filaments. Exposure to modeled microgravity induced a decrease in α-SMA and E-CAD expressions. Also, the expression of the pro-angiogenic protein VEGF decreased, while that of the inflammatory signal COX-2 increased. Fibroblast ability to adhere, migrate, and respond to chemoattractants (PRP), closely related to cytoskeleton integrity and membrane junctions, was significantly impaired. Nevertheless, PRP was able to partially restore fibroblast migration.

  3. Cannabinoids inhibit fibrogenesis in diffuse systemic sclerosis fibroblasts.

    Science.gov (United States)

    Garcia-Gonzalez, Estrella; Selvi, Enrico; Balistreri, Epifania; Lorenzini, Sauro; Maggio, Roberta; Natale, Maria-Rita; Capecchi, Pier-Leopoldo; Lazzerini, Pietro-Enea; Bardelli, Marco; Laghi-Pasini, Franco; Galeazzi, Mauro

    2009-09-01

    It has been demonstrated that the endocannabinoid system is up-regulated in pathologic fibrosis and that modulation of the cannabinoid receptors might limit the progression of uncontrolled fibrogenesis. The aim of this study was to investigate whether the synthetic cannabinoid receptor agonist WIN55,212-2 could modulate fibrogenesis in an in vitro model of dcSSc. The expression of cannabinoid receptors CB1 and CB2 was assessed in dcSSc fibroblasts and healthy control fibroblasts. To investigate the effect of WIN55,212-2 on dcSSc fibrogenesis, we studied type I collagen, profibrotic cytokines, fibroblast transdifferentiation into myofibroblasts, apoptotic processes and activation of the extracellular signal-related kinase 1/2 pathway prior to and after the treatment with the synthetic cannabinoid at increasing concentrations. Both CB1 and CB2 receptors were over-expressed in dcSSc fibroblasts compared with healthy controls. WIN55,212-2 caused a reduction in extracellular matrix deposition and counteracted several behavioural abnormalities of scleroderma fibroblasts including transdifferentiation into myofibroblasts and resistance to apoptosis. The anti-fibrogenic effect of WIN55,212-2 was not reverted by selective cannabinoid antagonists. Our preliminary findings suggest that cannabinoids are provided with an anti-fibrotic activity, thereby possibly representing a new class of agents targeting fibrosis diseases.

  4. Fibroblasts and myofibroblasts in wound healing: force generation and measurement.

    Science.gov (United States)

    Li, Bin; Wang, James H-C

    2011-11-01

    Fibroblasts are one of the most abundant cell types in connective tissues. These cells are responsible for tissue homeostasis under normal physiological conditions. When tissues are injured, fibroblasts become activated and differentiate into myofibroblasts, which generate large contractions and actively produce extracellular matrix (ECM) proteins to facilitate wound closure. Both fibroblasts and myofibroblasts play a critical role in wound healing by generating traction and contractile forces, respectively, to enhance wound contraction. This review focuses on the mechanisms of force generation in fibroblasts and myofibroblasts and techniques for measuring such cellular forces. Such a topic was chosen specifically because of the dual effects that fibroblasts/myofibroblasts have in wound healing process- a suitable amount of force generation and matrix deposition is beneficial for wound healing; excessive force and matrix production, however, result in tissue scarring and even malfunction of repaired tissues. Therefore, understanding how forces are generated in these cells and knowing exactly how much force they produce may guide the development of optimal protocols for more effective treatment of tissue wounds in clinical settings. Copyright © 2009 Tissue Viability Society. Published by Elsevier Ltd. All rights reserved.

  5. Helium-neon laser treatment transforms fibroblasts into myofibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Pourreau-Schneider, N.; Ahmed, A.; Soudry, M.; Jacquemier, J.; Kopp, F.; Franquin, J.C.; Martin, P.M. (CNRS Unite 1175, Marseille (France))

    1990-07-01

    The differentiation of myofibroblastic cells from normal human gingival fibroblasts in vitro has been established by transmission electron microscopy and quantitated by immunohistochemistry, using antigelsolin monoclonal antibodies. Untreated control cultures were compared to cultures exposed to Helium-Neon (He-Ne) laser irradiation. A direct and massive transformation of the cultured fibroblasts into myofibroblasts was observed as early as 24 hours after laser treatment, whereas control cultures were comprised of only resting fibroblasts and active fibroblasts. This in vitro induction of myofibroblasts may be analogous to that which occurs in vivo. Therefore we undertook a similar study using biopsies from gingival tissues after wisdom tooth extraction. Myofibroblasts were present in the connective tissue of laser-treated gums 48 hours after irradiation, but not in untreated contralateral control tissues. These data provide evidence that the primary biologic effect of the Helium-Neon laser on connective tissue is the rapid generation of myofibroblasts from fibroblasts. The induction of a phenotype with contractile properties may have clinical significance in the acceleration of the wound-healing process.

  6. Effect of captopril on collagen metabolisms in keloid fibroblast cells.

    Science.gov (United States)

    Chen, Junjie; Zhao, Sha; Liu, Yong; Cen, Ying; Nicolas, Crook

    2016-12-01

    Keloid is a proliferative disease of fibrous tissues. The mechanism and consistently effective treatments of keloid remained unknown. Although there was a report about treating keloid with topical captopril, the further investigation about captopril affecting keloid has not been performed so far. The aim of this study was to analyse the effect of captopril on collagen metabolisms in keloid fibroblast cells, and to provide information for the mechanism and therapy of keloid. To investigate the effects and relative mechanism of captopril on keloid fibroblast cells, we examined the changes of collagen metabolism, expression of angiotensin, transforming growth factor (TGF)-β1, platelet-derived growth factor (PDGF)-BB and heat shock protein 47 (HSP47), and cellular proliferation in keloid fibroblast cells. We found that all collagen metabolisms, expression of TGF-β1, PDGF-BB and HSP47, and cellular proliferation decreased significantly with effective captopril concentrations in keloid fibroblast cells. With a comprehensive analysis of test results, we proposed that captopril may decrease the expression of angiotensin, PDGF-BB, TGF-β1 and HSP47, and further inhibit proliferation and collagen synthesis of keloid fibroblast cells, which were the key in keloid formation. © 2014 Royal Australasian College of Surgeons.

  7. Fibroblasts from long-lived rodent species exclude cadmium.

    Science.gov (United States)

    Dostál, Lubomír; Kohler, William M; Penner-Hahn, James E; Miller, Richard A; Fierke, Carol A

    2015-01-01

    Resistance to the lethal effects of cellular stressors, including the toxic heavy metal cadmium (Cd), is characteristic of fibroblast cell lines derived from long-lived bird and rodent species, as well as cell lines from several varieties of long-lived mutant mice. To explore the mechanism of resistance to Cd, we used inductively coupled plasma mass spectroscopy to measure the rate of Cd uptake into primary fibroblasts of 15 rodent species. These data indicate that fibroblasts from long-lived rodent species have slower rates of Cd uptake from the extracellular medium than those from short-lived species. In addition, fibroblasts from short-lived species export more zinc after exposure to extracellular Cd than cells from long-lived species. Lastly, fibroblasts from long-lived rodent species have lower baseline concentrations of two redox-active metals, iron and copper. Our results suggest that evolution of longevity among rodents required adjustment of cellular properties to alter metal homeostasis and to reduce the toxic effects of heavy metals that accumulate over the course of a longer life span.

  8. Modeled Microgravity Affects Fibroblast Functions Related to Wound Healing

    Science.gov (United States)

    Cialdai, Francesca; Vignali, Leonardo; Morbidelli, Lucia; Colciago, Alessandra; Celotti, Fabio; Santi, Alice; Caselli, Anna; Cirri, Paolo; Monici, Monica

    2017-01-01

    Wound healing is crucial for the survival of an organism. Therefore, in the perspective of space exploration missions, it is important to understand if and how microgravity conditions affect the behavior of the cell populations involved in wound healing and the evolution of the process. Since fibroblasts are the major players in tissue repair, this study was focused on the behavior of fibroblasts in microgravity conditions, modeled by a RCCS. Cell cytoskeleton was studied by immunofluorescence microscopy, the ability to migrate was assessed by microchemotaxis and scratch assay, and the expression of markers of fibroblast activation, angiogenesis, and inflammation was assessed by western blot. Results revealed that after cell exposure to modeled microgravity conditions, a thorough rearrangement of microtubules occurred and α-SMA bundles were replaced by a tight network of faulty and disorganized filaments. Exposure to modeled microgravity induced a decrease in α-SMA and E-CAD expressions. Also, the expression of the pro-angiogenic protein VEGF decreased, while that of the inflammatory signal COX-2 increased. Fibroblast ability to adhere, migrate, and respond to chemoattractants (PRP), closely related to cytoskeleton integrity and membrane junctions, was significantly impaired. Nevertheless, PRP was able to partially restore fibroblast migration.

  9. Fibroblast Cell-Based Therapy for Experimental Autoimmune Diabetes.

    Directory of Open Access Journals (Sweden)

    Reza B Jalili

    Full Text Available Type 1 diabetes (T1D results from autoimmune destruction of insulin producing β cells of the pancreatic islets. Curbing autoimmunity at the initiation of T1D can result in recovery of residual β cells and consequently remission of diabetes. Here we report a cell-based therapy for autoimmune diabetes in non-obese diabetic (NOD mice using dermal fibroblasts. This was achieved by a single injection of fibroblasts, expressing the immunoregulatory molecule indoleamine 2,3 dioxygenase (IDO, into peritoneal cavity of NOD mice shortly after the onset of overt hyperglycemia. Mice were then monitored for reversal of hyperglycemia and changes in inflammatory/regulatory T cell profiles. Blood glucose levels dropped into the normal range in 82% of NOD mice after receiving IDO-expressing fibroblasts while all control mice remained diabetic. We found significantly reduced islet inflammation, increased regulatory T cells, and decreased T helper 17 cells and β cell specific autoreactive CD8+ T cells following IDO cell therapy. We further showed that some of intraperitoneal injected fibroblasts migrated to local lymph nodes and expressed co-inhibitory molecules. These findings suggest that IDO fibroblasts therapy can reinstate self-tolerance and alleviate β cell autoreactivity in NOD mice, resulting in remission of autoimmune diabetes.

  10. Fibroblast Cell-Based Therapy for Experimental Autoimmune Diabetes.

    Science.gov (United States)

    Jalili, Reza B; Zhang, Yun; Hosseini-Tabatabaei, Azadeh; Kilani, Ruhangiz T; Khosravi Maharlooei, Mohsen; Li, Yunyuan; Salimi Elizei, Sanam; Warnock, Garth L; Ghahary, Aziz

    2016-01-01

    Type 1 diabetes (T1D) results from autoimmune destruction of insulin producing β cells of the pancreatic islets. Curbing autoimmunity at the initiation of T1D can result in recovery of residual β cells and consequently remission of diabetes. Here we report a cell-based therapy for autoimmune diabetes in non-obese diabetic (NOD) mice using dermal fibroblasts. This was achieved by a single injection of fibroblasts, expressing the immunoregulatory molecule indoleamine 2,3 dioxygenase (IDO), into peritoneal cavity of NOD mice shortly after the onset of overt hyperglycemia. Mice were then monitored for reversal of hyperglycemia and changes in inflammatory/regulatory T cell profiles. Blood glucose levels dropped into the normal range in 82% of NOD mice after receiving IDO-expressing fibroblasts while all control mice remained diabetic. We found significantly reduced islet inflammation, increased regulatory T cells, and decreased T helper 17 cells and β cell specific autoreactive CD8+ T cells following IDO cell therapy. We further showed that some of intraperitoneal injected fibroblasts migrated to local lymph nodes and expressed co-inhibitory molecules. These findings suggest that IDO fibroblasts therapy can reinstate self-tolerance and alleviate β cell autoreactivity in NOD mice, resulting in remission of autoimmune diabetes.

  11. Fibroblast cytoskeletal remodeling contributes to connective tissue tension.

    Science.gov (United States)

    Langevin, Helene M; Bouffard, Nicole A; Fox, James R; Palmer, Bradley M; Wu, Junru; Iatridis, James C; Barnes, William D; Badger, Gary J; Howe, Alan K

    2011-05-01

    The visco-elastic behavior of connective tissue is generally attributed to the material properties of the extracellular matrix rather than cellular activity. We have previously shown that fibroblasts within areolar connective tissue exhibit dynamic cytoskeletal remodeling within minutes in response to tissue stretch ex vivo and in vivo. Here, we tested the hypothesis that fibroblasts, through this cytoskeletal remodeling, actively contribute to the visco-elastic behavior of the whole tissue. We measured significantly increased tissue tension when cellular function was broadly inhibited by sodium azide and when cytoskeletal dynamics were compromised by disrupting microtubules (with colchicine) or actomyosin contractility (via Rho kinase inhibition). These treatments led to a decrease in cell body cross-sectional area and cell field perimeter (obtained by joining the end of all of a fibroblast's processes). Suppressing lamellipodia formation by inhibiting Rac-1 decreased cell body cross-sectional area but did not affect cell field perimeter or tissue tension. Thus, by changing shape, fibroblasts can dynamically modulate the visco-elastic behavior of areolar connective tissue through Rho-dependent cytoskeletal mechanisms. These results have broad implications for our understanding of the dynamic interplay of forces between fibroblasts and their surrounding matrix, as well as for the neural, vascular, and immune cell populations residing within connective tissue.

  12. Reprogramming fibroblasts into induced pluripotent stem cells with Bmi

    Institute of Scientific and Technical Information of China (English)

    Jai-Hee Moon; June Seok Heo; Jun Sung Kim; Eun Kyoung Jun; Jung Han Lee; Aeree Kim; Jonggun Kim

    2011-01-01

    Somatic cells can be reprogrammed into induced pluripotent stem (iPS) cells by the transcription factors Oct4,Sox2,and Klf4 in combination with c-Myc.Recently,Sox2 plus Oct4 was shown to reprogram fibroblasts and Oct4 alone was able to reprogram mouse and human neural stem cells (NSCs) into iPS cells.Here,we report that Bmi1 leads to the transdifferentiation of mouse fibroblasts into NSC-like cells,and,in combination with Oct4,can replace Sox2,Klf4 and c-Myc during the reprogramming of fibroblasts into iPS cells.Furthermore,activation of sonic hedgehog signaling (by Shh,purmorphamine,or oxysterol) compensates for the effects of Bmil,and,in combination with Oct4,reprograms mouse embryonic and adult fibroblasts into iPS cells.One- and two-factor iPS cells are similar to mouse embryonic stem cells in their global gene expression profile,epigenetic status,and in vitro and in vivo differentiation into all three germ layers,as well as teratoma formation and germline transmission in vivo.These data support that converting fibroblasts with Bmi1 or activation of the sonic hedgehog pathway to an intermediate cell type that expresses Sox2,KIf4,and N-Myc allows iPS generation via the addition of Oct4.

  13. Immunopathology of Brucella infection.

    Science.gov (United States)

    Baldi, Pablo C; Giambartolomei, Guillermo H

    2013-04-01

    In spite of the protean nature of the disease, inflammation is a hallmark of brucellosis and affected tissues usually exhibit inflammatory infiltrates. As Brucella lacks exotoxins, exoproteases or cytolysins, pathological findings in brucellosis probably arise from inflammation-driven processes. The cellular and molecular bases of immunopathological phenomena probably involved in Brucella pathogenesis have been unraveled in the last few years. Brucella-infected osteoblasts, either alone or in synergy with infected macrophages, produce cytokines, chemokines and matrixmetalloproteinases (MMPs), and similar phenomena are mounted by fibroblast-like synoviocytes. The released cytokines promote the secretion of MMPs and induce osteoclastogenesis. Altogether, these phenomena may contribute to the bone loss and cartilage degradation usually observed in brucellar arthritis and osteomyelitis. Proinflammatory cytokines may be also involved in the pathogenesis of neurobrucellosis. B. abortus and its lipoproteins elicit an inflammatory response in the CNS of mice, leading to astrogliosis, a characteristic feature of neurobrucellosis. Heat-killed bacteria (HKBA) and the L-Omp19 lipoprotein elicit astrocyte apoptosis and proliferation (two features of astrogliosis), and apoptosis depends on TNF-α signaling. Brucella also infects and replicates in human endothelial cells, inducing the production of chemokines and IL-6, and an increased expression of adhesion molecules. The sustained inflammatory process derived from the longlasting infection of the endothelium may be important for the development of endocarditis. Therefore, while Brucella induces a low grade inflammation as compared to other pathogens, its prolonged intracellular persistence in infected tissues supports a long-lasting inflammatory response that mediates different pathways of tissue damage. In this context, approaches to avoid the invasion of host cells or limit the intracellular survival of the bacterium may be

  14. A fibroblast-associated antigen: Characterization in fibroblasts and immunoreactivity in smooth muscle differentiated stromal cells

    DEFF Research Database (Denmark)

    Rønnov-Jessen, Lone; Celis, Julio E.; van Deurs, Bo

    1992-01-01

    from vascular smooth muscle cells. The antigen was detected on the cell surface and in cathepsin D-positive and acridine orange-accumulating vesicular compartments of fibroblasts. Ultrastructurally, the antigen was revealed in coated pits and in endosomal and lysosomal structures. 1B10 recognized three...... major brands migrating at apparent Mr of 38,000, 45,000, and 80,000, in addition to many minor bands between Mr 45,000 and 97,000, including Mr 52,000. The Mr 45,000 and 38,000 were associated with the cell membrane and Mr 52,000 as well as Mr 38,000 were associated with the lysosomes. The 1B10...

  15. Keratinocyte growth factor mRNA expression in periodontal ligament fibroblasts

    DEFF Research Database (Denmark)

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

    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 expres......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 m...

  16. Feedback Activation of Basic Fibroblast Growth Factor Signaling via the Wnt/β-Catenin Pathway in Skin Fibroblasts.

    Science.gov (United States)

    Wang, Xu; Zhu, Yuting; Sun, Congcong; Wang, Tao; Shen, Yingjie; Cai, Wanhui; Sun, Jia; Chi, Lisha; Wang, Haijun; Song, Na; Niu, Chao; Shen, Jiayi; Cong, Weitao; Zhu, Zhongxin; Xuan, Yuanhu; Li, Xiaokun; Jin, Litai

    2017-01-01

    Skin wound healing is a complex process requiring the coordinated behavior of many cell types, especially in the proliferation and migration of fibroblasts. Basic fibroblast growth factor (bFGF) is a member of the FGF family that promotes fibroblast migration, but the underlying molecular mechanism remains elusive. The present RNA sequencing study showed that the expression levels of several canonical Wnt pathway genes, including Wnt2b, Wnt3, Wnt11, T-cell factor 7 (TCF7), and Frizzled 8 (FZD8) were modified by bFGF stimulation in fibroblasts. Enzyme-linked immunosorbent assay (ELISA) analysis also showed that Wnt pathway was activated under bFGF treatment. Furthermore, treatment of fibroblasts with lithium chloride or IWR-1, an inducer and inhibitor of the Wnt signaling pathway, respectively, promoted and inhibited cell migration. Also, levels of cytosolic glycogen synthase kinase 3 beta phosphorylated at serine(9) (pGSK3β Ser(9)) and nuclear β-catenin were increased upon exposure to bFGF. Molecular and biochemical assays indicated that phosphoinositide 3-kinase (PI3K) signaling activated the GSK3β/β-catenin/Wnt signaling pathway via activation of c-Jun N-terminal kinase (JNK), suggesting that PI3K and JNK act at the upstream of β-catenin. In contrast, knock-down of β-catenin delayed fibroblast cell migration even under bFGF stimulation. RNA sequencing analysis of β-catenin knock-down fibroblasts demonstrated that β-catenin positively regulated the transcription of bFGF and FGF21. Moreover, FGF21 treatment activated AKT and JNK, and accelerated fibroblast migration to a similar extent as bFGF does. In addition, ELISA analysis demonstrated that both of bFGF and FGF21 were auto secretion factor and be regulated by Wnt pathway stimulators. Taken together, our analyses define a feedback regulatory loop between bFGF (FGF21) and Wnt signaling acting through β-catenin in skin fibroblasts.

  17. Fibroblasts from phenotypically normal palmar fascia exhibit molecular profiles highly similar to fibroblasts from active disease in Dupuytren's Contracture

    Directory of Open Access Journals (Sweden)

    Satish Latha

    2012-05-01

    Full Text Available Abstract Background Dupuytren's contracture (DC is a fibroproliferative disorder characterized by the progressive development of a scar-like collagen-rich cord that affects the palmar fascia of the hand and leads to digital flexion contractures. DC is most commonly treated by surgical resection of the diseased tissue, but has a high reported recurrence rate ranging from 27% to 80%. We sought to determine if the transcriptomic profiles of fibroblasts derived from DC-affected palmar fascia, adjacent phenotypically normal palmar fascia, and non-DC palmar fascial tissues might provide mechanistic clues to understanding the puzzle of disease predisposition and recurrence in DC. Methods To achieve this, total RNA was obtained from fibroblasts derived from primary DC-affected palmar fascia, patient-matched unaffected palmar fascia, and palmar fascia from non-DC patients undergoing carpal tunnel release (6 patients in each group. These cells were grown on a type-1 collagen substrate (to better mimic their in vivo environments. Microarray analyses were subsequently performed using Illumina BeadChip arrays to compare the transcriptomic profiles of these three cell populations. Data were analyzed using Significance Analysis of Microarrays (SAM v3.02, hierarchical clustering, concordance mapping and Venn diagram. Results We found that the transcriptomic profiles of DC-disease fibroblasts and fibroblasts from unaffected fascia of DC patients exhibited a much greater overlap than fibroblasts derived from the palmar fascia of patients undergoing carpal tunnel release. Quantitative real time RT-PCR confirmed the differential expression of select genes validating the microarray data analyses. These data are consistent with the hypothesis that predisposition and recurrence in DC may stem, at least in part, from intrinsic similarities in the basal gene expression of diseased and phenotypically unaffected palmar fascia fibroblasts. These data also demonstrate that

  18. Fibroblasts from phenotypically normal palmar fascia exhibit molecular profiles highly similar to fibroblasts from active disease in Dupuytren's Contracture

    Science.gov (United States)

    2012-01-01

    Background Dupuytren's contracture (DC) is a fibroproliferative disorder characterized by the progressive development of a scar-like collagen-rich cord that affects the palmar fascia of the hand and leads to digital flexion contractures. DC is most commonly treated by surgical resection of the diseased tissue, but has a high reported recurrence rate ranging from 27% to 80%. We sought to determine if the transcriptomic profiles of fibroblasts derived from DC-affected palmar fascia, adjacent phenotypically normal palmar fascia, and non-DC palmar fascial tissues might provide mechanistic clues to understanding the puzzle of disease predisposition and recurrence in DC. Methods To achieve this, total RNA was obtained from fibroblasts derived from primary DC-affected palmar fascia, patient-matched unaffected palmar fascia, and palmar fascia from non-DC patients undergoing carpal tunnel release (6 patients in each group). These cells were grown on a type-1 collagen substrate (to better mimic their in vivo environments). Microarray analyses were subsequently performed using Illumina BeadChip arrays to compare the transcriptomic profiles of these three cell populations. Data were analyzed using Significance Analysis of Microarrays (SAM v3.02), hierarchical clustering, concordance mapping and Venn diagram. Results We found that the transcriptomic profiles of DC-disease fibroblasts and fibroblasts from unaffected fascia of DC patients exhibited a much greater overlap than fibroblasts derived from the palmar fascia of patients undergoing carpal tunnel release. Quantitative real time RT-PCR confirmed the differential expression of select genes validating the microarray data analyses. These data are consistent with the hypothesis that predisposition and recurrence in DC may stem, at least in part, from intrinsic similarities in the basal gene expression of diseased and phenotypically unaffected palmar fascia fibroblasts. These data also demonstrate that a collagen

  19. Effects of trichostatin A on human Tenon capsule fibroblast

    Directory of Open Access Journals (Sweden)

    Xiao-Yan Li

    2014-11-01

    Full Text Available AIM:To investigate the effect of trichostatin A(TSAon cell proliferation and the expressions of histone deacetylase 1(HDAC1and HDAC2 in cultured human Tenon capsule fibroblast(HTF.METHODS: Human Tenon capsule fibroblasts were cultured in vitro after glaucoma filtration surgery. The third to sixth passage of cell were treated by 600nmol/L TSA or none. Cell viability measured by MTT assay after 1, 2 and 3d respectively. The expressions of HDAC1 and HDAC2 were analyzed by Western blot 2d after TSA treatment.RESULTS: Compared to the control, cell viability decreased significantly after treatment with TSA at 1d(PCONCLUSION: TSA inhibits the proliferation of Tenon capsule fibroblast by inhibiting the expression of HDAC1 and HDAC2, and reduces subconjunctival scar formation.

  20. Substratum topography modulates corneal fibroblast to myofibroblast transformation.

    Science.gov (United States)

    Myrna, Kathern E; Mendonsa, Rima; Russell, Paul; Pot, Simon A; Liliensiek, Sara J; Jester, James V; Nealey, Paul F; Brown, Donald; Murphy, Christopher J

    2012-02-01

    The transition of corneal fibroblasts to the myofibroblast phenotype is known to be important in wound healing. The purpose of this study was to determine the effect of topographic cues on TGFβ-induced myofibroblast transformation of corneal cells. Rabbit corneal fibroblasts were cultured on nanopatterned surfaces having topographic features of varying sizes. Cells were cultured in media containing TGFβ at concentrations ranging from 0 to 10 ng/mL. RNA and protein were collected from cells cultured on topographically patterned and planar substrates and analyzed for the myofibroblast marker α-smooth muscle actin (αSMA) and Smad7 expression by quantitative real time PCR. Western blot and immunocytochemistry analysis for αSMA were also performed. Cells grown on patterned surfaces demonstrated significantly reduced levels of αSMA (P fibroblast phenotype while pathologic microenvironmental alterations may be permissive for increased myofibroblast differentiation and the development of fibrosis and corneal haze.

  1. Chemical Conversion of Human Fibroblasts into Functional Schwann Cells

    Directory of Open Access Journals (Sweden)

    Eva C. Thoma

    2014-10-01

    Full Text Available Direct transdifferentiation of somatic cells is a promising approach to obtain patient-specific cells for numerous applications. However, conversion across germ-layer borders often requires ectopic gene expression with unpredictable side effects. Here, we present a gene-free approach that allows efficient conversion of human fibroblasts via a transient progenitor stage into Schwann cells, the major glial cell type of peripheral nerves. Using a multikinase inhibitor, we transdifferentiated fibroblasts into transient neural precursors that were subsequently further differentiated into Schwann cells. The resulting induced Schwann cells (iSCs expressed numerous Schwann cell-specific proteins and displayed neurosupportive and myelination capacity in vitro. Thus, we established a strategy to obtain mature Schwann cells from human postnatal fibroblasts under chemically defined conditions without the introduction of ectopic genes.

  2. Expression of fibroblast growth factor-21 in muscle is associated with lipodystrophy, insulin resistance and lipid disturbances in patients with HIV

    DEFF Research Database (Denmark)

    Lindegaard, Birgitte; Hvid, Thine; Grøndahl, Thomas;

    2013-01-01

    Fibroblast growth factor (FGF)-21 is a novel regulator of glucose and lipid metabolism. Recently, increased FGF-21 mRNA expression in muscle was found in patients with type 2 diabetes, but the role for FGF-21 in muscle is not well understood. Patients with HIV-infection and lipodystrophy are char......Fibroblast growth factor (FGF)-21 is a novel regulator of glucose and lipid metabolism. Recently, increased FGF-21 mRNA expression in muscle was found in patients with type 2 diabetes, but the role for FGF-21 in muscle is not well understood. Patients with HIV-infection and lipodystrophy...... are characterised by various degree of lipid-driven insulin resistance. We hypothesized that muscle FGF-21 mRNA would be altered in HIV patients with lipodystrophy....

  3. Overexpression of N-acetylgalactosamine-4-sulphatase induces a multiple sulphatase deficiency in mucopolysaccharidosis-type-VI fibroblasts.

    Science.gov (United States)

    Anson, D S; Muller, V; Bielicki, J; Harper, G S; Hopwood, J J

    1993-09-15

    High-titre stocks of an amphotropic retrovirus, constructed so as to express a full-length cDNA encoding the human lysosomal enzyme N-acetylgalactosamine-4-sulphatase (4-sulphatase) from the cytomegalovirus immediate early promoter, were used to infect skin fibroblasts from a clinically severe mucopolysaccharidosis type VI (MPS VI) patient. The infected MPS VI cells showed correction of the enzymic defect with the enzyme being expressed at high levels and in the correct subcellular compartment. Surprisingly this did not result in correction of glycosaminoglycan turnover as measured by accumulation of 35S in metabolically labelled cells. We demonstrate that this is apparently caused by an induced reduction of the activities of other lysosomal sulphatases, presumably due to competition for a sulphatase-specific processing mechanism by the over-expressed 4-sulphatase. The level of steroid sulphatase, which is a microsomal sulphatase, was also reduced. Infection of skin fibroblasts from a second, clinically mildly affected, MPS VI patient with the same virus also resulted in no significant change in the level of glycosaminoglycan storage. However, in this case the cause of the observed phenomenon was less clear. These results are of obvious practical importance when considering gene therapy for a sulphatase deficiency such as MPS VI and also provide possible new avenues for exploration of the processes involved in sulphatase synthesis and genetically determined multiple sulphatase deficiency.

  4. Clinical Experience with Chitosan Matrix and Cultured Fibroblasts for Burns

    Directory of Open Access Journals (Sweden)

    Gaziza Danlybayeva

    2014-12-01

    Full Text Available Introduction. Burns are an important public health challenge due to the frequency of getting burns in day-to-day life, occupational hazards, and catastrophes. Treatment of burns is complex and is associated with high morbidity and mortality. Duration and complexity of burn treatment require finding new ways of curing and rehabilitating burns. The result of burn treatment plays a significant role in post-traumatic status of a patient and his or her consequent adaptation in society. Chitosan is a natural safe non-toxic product compatible with human tissues, characterized by hydrosorbid, anticoagulant, antibacterial, and wound healing features. The study aims to  show a clinical application of chitosan-pectin scaffold with cultured human skin fibroblasts in the treatment of deep burns.Methods. The substrate was prepared by dissolving 3% chitosan in 0.5N acetic acid, which was then mixed with 3% solution of pectin dissolved in distillated water. Chitosan film was formed in a Petri dish for 20-24 hours at 20-25 °C. After drying the film, cultured allogeneic fibroblasts (patent number RK-25091 were seeded on its surface.Results. The results from an in vitro culture study showed that human allogeneic fibroblasts could adhere well and grow on the selected scaffold with a typical morphology. During autodermoplasty surgery, cultured allogeneic fibroblasts were applied on granulating wounds of 9 patients with IIIA to IVB degree burns and limited donor resources. Wounds treated with the fibroblast-seeded scaffold among all patients provided the highest level of re-epithelialization (day 5, in comparison to cell-free scaffold (day 7 and untreated surface of wounds (day 10.Conclusion. Our results indicate the potential use of chitosan for wound healing due to its allogenic fibroblast adherence to scaffolding as well as high epithelization. This warrants further studies on chitosan for use in wounds resulting from third and fourth degree burns.

  5. Expression of the endocannabinoid system in fibroblasts and myofascial tissues.

    Science.gov (United States)

    McPartland, John M

    2008-04-01

    The endocannabinoid (eCB) system, like the better-known endorphin system, consists of cell membrane receptors, endogenous ligands and ligand-metabolizing enzymes. Two cannabinoid receptors are known: CB(1) is principally located in the nervous system, whereas CB(2) is primarily associated with the immune system. Two eCB ligands, anandamide (AEA) and 2-arachidonoylglycerol (2-AG), are mimicked by cannabis plant compounds. The first purpose of this paper was to review the eCB system in detail, highlighting aspects of interest to bodyworkers, especially eCB modulation of pain and inflammation. Evidence suggests the eCB system may help resolve myofascial trigger points and relieve symptoms of fibromyalgia. However, expression of the eCB system in myofascial tissues has not been established. The second purpose of this paper was to investigate the eCB system in fibroblasts and other fascia-related cells. The investigation used a bioinformatics approach, obtaining microarray data via the GEO database (www.ncbi.nlm.nih.gov/geo/). GEO data mining revealed that fibroblasts, myofibroblasts, chondrocytes and synoviocytes expressed CB(1), CB(2) and eCB ligand-metabolizing enzymes. Fibroblast CB(1) levels nearly equalled levels expressed by adipocytes. CB(1) levels upregulated after exposure to inflammatory cytokines and equiaxial stretching of fibroblasts. The eCB system affects fibroblast remodeling through lipid rafts associated with focal adhesions and dampens cartilage destruction by decreasing fibroblast-secreted metalloproteinase enzymes. In conclusion, the eCB system helps shape biodynamic embryological development, diminishes nociception and pain, reduces inflammation in myofascial tissues and plays a role in fascial reorganization. Practitioners wield several tools that upregulate eCB activity, including myofascial manipulation, diet and lifestyle modifications, and pharmaceutical approaches.

  6. Association of spirochetal infection with Morgellons disease

    OpenAIRE

    Middelveen, Marianne J; Divya Burugu; Akhila Poruri; Jennie Burke; Mayne, Peter J; Eva Sapi; Kahn, Douglas G; Stricker, Raphael B

    2013-01-01

    Morgellons disease (MD) is an emerging multisystem illness characterized by skin lesions with unusual filaments embedded in or projecting from epithelial tissue. Filament formation results from abnormal keratin and collagen expression by epithelial-based keratinocytes and fibroblasts. Recent research comparing MD to bovine digital dermatitis, an animal infectious disease with similar skin features, provided clues that spirochetal infection could play an important role in the human disease as ...

  7. Inhibition and recovery of the replication of depurinated parvovirus DNA in mouse fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Vos, J.M.; Avalosse, B.; Su, Z.Z.; Rommelaere, J.

    1984-01-01

    Apurinic sites were introduced in the single-stranded DNA of parvovirus minute-virus-of-mice (MVM) and their effect on viral DNA synthesis was measured in mouse fibroblasts. Approximately one apurinic site per viral genome, is sufficient to block its replication in untreated cells. The exposure of host cells to a sublethal dose of UV-light 15 hours prior to virus infection, enhances their ability to support the replication of depurinated MVM. Cell preirradiation induces the apparent overcome of 10-15% of viral DNA replication blocks. These results indicate that apurinic sites prevent mammalian cells from replicating single-stranded DNA unless a recovery process is activated by cell UV-irradiation.

  8. The role of hexon in egg drop syndrome virus (EDSV) inducing apoptosis in duck embryo fibroblast cells.

    Science.gov (United States)

    Qi, Xuefeng; Xu, Jiamin; Wang, Zugui; Wang, Xueping; Wang, Jingyu

    2017-07-17

    Although extensive efforts have been made to understand adenovirus infection in human cells, little is known for egg drop syndrome virus (EDSV) infection in the avian-derived cells. In this study, the effects of EDSV infection as well as the possible role hexon protein, the main building block of the EDSV capsid, on apoptosis induction in duck embryo fibroblast (DEF) cells was examined. Flow cytometry analysis and TUNEL assay revealed that EDSV infection induced significant apoptosis in DEF cells compared with mock infected cells. Interestingly, the increase of the apoptosis rate detected in EDSV infected DEF cells were accompanied by an increased virus load in cells in a time-dependent manner. Furthermore, a time-dependent decrease in hexon protein expression levels in hexon transfected DEF cells in parallel with a gradual decrease in TUNEL-labeling cells was also observed in the current study. In addition, caspase activity detection and western blot analysis indicates that either EDSV infection or EDSV hexon transfection both induced apoptosis of DEF cells via activating both the exogenous and the mitochondrial pathway. Copyright © 2017. Published by Elsevier Ltd.

  9. Dipeptides Increase Functional Activity of Human Skin Fibroblasts.

    Science.gov (United States)

    Malinin, V V; Durnova, A O; Polyakova, V O; Kvetnoi, I M

    2015-05-01

    We analyzed the effect of dipeptide Glu-Trp and isovaleroyl-Glu-Trp in concentrations of 0.2, 2 and 20 μg/ml and Actovegin preparation on functional activity of human skin fibroblasts. Dipeptides, especially Glu-Trp, produce a stimulating effect on human skin fibroblasts and their effect is equivalent to that of Actovegin. Dipeptides stimulate cell renewal processes by activating synthesis of Ki-67 and reducing expression of caspase-9 and enhance antioxidant function of the cells by stimulating the expression of Hsp-90 and inducible NO-synthase. These findings suggest that dipeptides are promising candidates for preparations stimulating reparative processes.

  10. Attachment of epithelial cells and fibroblasts to ceramic materials.

    Science.gov (United States)

    Niederauer, G G; McGee, T D; Keller, J C; Zaharias, R S

    1994-04-01

    This study examined in vitro gingival epithelial and fibroblast cell attachment to ceramic materials made of tricalcium phosphate and/or magnesium aluminate spinel. The composite made of tricalcium phosphate and spinel is called 'osteoceramic'. These ceramics had various compositions and surface structures, which were initially characterized. Cell attachment assays were performed using both cell types to compare cellular response to the ceramic materials. Specimens were also prepared for scanning electron microscopy to investigate cellular morphology. The highest levels of cell attachment for gingival epithelial cells were observed on the rough osteoceramic surface, whereas gingival fibroblasts attached least to the rough osteoceramic surface.

  11. Agminated Fibroblastic Conective Tissue Nevus: A New Clinical Presentation.

    Science.gov (United States)

    Downey, Camila; Requena, Luis; Bagué, Silvia; Sánchez Martínez, Miquel Ángel; Lloreta, Josep; Baselga, Eulalia

    2016-07-01

    Connective tissue nevi are benign hamartomatous lesions in which one or several of the components of the dermis (collagen, elastin, glicosaminoglycans) show predominance or depletion. Recently, de Feraudy et al broadened the spectrum of connective tissue nevus, describing fibroblastic connective tissue nevus (FCTN), which is characterized by proliferation of CD34(+) cells of fibroblastic and myofibroblastic lineage. Only solitary papules and nodules have been described. We present the first case of FCTN with multiple agminated lesions on the leg of an infant and the difficulties encountered in the differential diagnosis with dermatofibrosarcoma protuberans.

  12. Effects of the novel compound DK223 ([1E,2E-1,2-Bis(6-methoxy-2H-chromen-3-yl)methylene]hydrazine) on migration and proliferation of human keratinocytes and primary dermal fibroblasts.

    Science.gov (United States)

    Ho, Manh Tin; Kang, Hyun Sik; Huh, Jung Sik; Kim, Young Mee; Lim, Yoongho; Cho, Moonjae

    2014-07-23

    Wound healing plays an important role in protecting the human body from external infection. Cell migration and proliferation of keratinocytes and dermal fibroblasts are essential for proper wound healing. Recently, several studies have demonstrated that secondary compounds produced in plants could affect skin cells migration and proliferation. In this study, we identified a novel compound DK223 ([1E,2E-1,2-bis(6-methoxy-2H-chromen-3-yl)methylene]hydrazine) that concomitantly induced human keratinocyte migration and dermal fibroblast proliferation. We evaluated the regulation of epithelial and mesenchymal protein markers, such as E-cadherin and Vimentin, in human keratinocytes, as well as extracellular matrix (ECM) secretion and metalloproteinase families in dermal fibroblasts. DK223 upregulated keratinocyte migration and significantly increased the epithelial marker E-cadherin in a time-dependent manner. We also found that reactive oxygen species (ROS) increased significantly in keratinocytes after 2 h of DK223 exposure, returning to normal levels after 24 h, which indicated that DK223 had an early shock effect on ROS production. DK223 also stimulated fibroblast proliferation, and induced significant secretion of ECM proteins, such as collagen I, III, and fibronectin. In dermal fibroblasts, DK223 treatment induced TGF-β1, which is involved in a signaling pathway that mediates proliferation. In conclusion, DK223 simultaneously induced both keratinocyte migration via ROS production and fibroblast proliferation via TGF-β1 induction.

  13. Effects of the Novel Compound DK223 ([1E,2E-1,2-Bis(6-methoxy-2H-chromen-3-ylmethylene]hydrazine on Migration and Proliferation of Human Keratinocytes and Primary Dermal Fibroblasts

    Directory of Open Access Journals (Sweden)

    Manh Tin Ho

    2014-07-01

    Full Text Available Wound healing plays an important role in protecting the human body from external infection. Cell migration and proliferation of keratinocytes and dermal fibroblasts are essential for proper wound healing. Recently, several studies have demonstrated that secondary compounds produced in plants could affect skin cells migration and proliferation. In this study, we identified a novel compound DK223 ([1E,2E-1,2-bis(6-methoxy-2H-chromen-3-ylmethylene]hydrazine that concomitantly induced human keratinocyte migration and dermal fibroblast proliferation. We evaluated the regulation of epithelial and mesenchymal protein markers, such as E-cadherin and Vimentin, in human keratinocytes, as well as extracellular matrix (ECM secretion and metalloproteinase families in dermal fibroblasts. DK223 upregulated keratinocyte migration and significantly increased the epithelial marker E-cadherin in a time-dependent manner. We also found that reactive oxygen species (ROS increased significantly in keratinocytes after 2 h of DK223 exposure, returning to normal levels after 24 h, which indicated that DK223 had an early shock effect on ROS production. DK223 also stimulated fibroblast proliferation, and induced significant secretion of ECM proteins, such as collagen I, III, and fibronectin. In dermal fibroblasts, DK223 treatment induced TGF-β1, which is involved in a signaling pathway that mediates proliferation. In conclusion, DK223 simultaneously induced both keratinocyte migration via ROS production and fibroblast proliferation via TGF-β1 induction.

  14. Expression of fibroblast growth factor-21 in muscle is associated with lipodystrophy, insulin resistance and lipid disturbances in patients with HIV

    DEFF Research Database (Denmark)

    Lindegaard, Birgitte; Hvid, Thine; Grøndahl, Thomas;

    2013-01-01

    Fibroblast growth factor (FGF)-21 is a novel regulator of glucose and lipid metabolism. Recently, increased FGF-21 mRNA expression in muscle was found in patients with type 2 diabetes, but the role for FGF-21 in muscle is not well understood. Patients with HIV-infection and lipodystrophy are char...... are characterised by various degree of lipid-driven insulin resistance. We hypothesized that muscle FGF-21 mRNA would be altered in HIV patients with lipodystrophy....

  15. Expression of fibroblast growth factor-21 in muscle is associated with lipodystrophy, insulin resistance and lipid disturbances in patients with HIV

    DEFF Research Database (Denmark)

    Lindegaard, Birgitte; Hvid, Thine; Grøndahl, Thomas

    2013-01-01

    Fibroblast growth factor (FGF)-21 is a novel regulator of glucose and lipid metabolism. Recently, increased FGF-21 mRNA expression in muscle was found in patients with type 2 diabetes, but the role for FGF-21 in muscle is not well understood. Patients with HIV-infection and lipodystrophy...... are characterised by various degree of lipid-driven insulin resistance. We hypothesized that muscle FGF-21 mRNA would be altered in HIV patients with lipodystrophy....

  16. Dynamics of the cellular metabolome during human cytomegalovirus infection.

    Directory of Open Access Journals (Sweden)

    Joshua Munger

    2006-12-01

    Full Text Available Viral replication requires energy and macromolecular precursors derived from the metabolic network of the host cell. Despite this reliance, the effect of viral infection on host cell metabolic composition remains poorly understood. Here we applied liquid chromatography-tandem mass spectrometry to measure the levels of 63 different intracellular metabolites at multiple times after human cytomegalovirus (HCMV infection of human fibroblasts. Parallel microarray analysis provided complementary data on transcriptional regulation of metabolic pathways. As the infection progressed, the levels of metabolites involved in glycolysis, the citric acid cycle, and pyrimidine nucleotide biosynthesis markedly increased. HCMV-induced transcriptional upregulation of specific glycolytic and citric acid cycle enzymes mirrored the increases in metabolite levels. The peak levels of numerous metabolites during infection far exceeded those observed during normal fibroblast growth or quiescence, demonstrating that HCMV markedly disrupts cellular metabolic homeostasis and institutes its own specific metabolic program.

  17. Inhibition by rebamipide of cytokine-induced or lipopolysaccharide-induced chemokine synthesis in human corneal fibroblasts.

    Science.gov (United States)

    Fukuda, Ken; Ishida, Waka; Tanaka, Hiroshi; Harada, Yosuke; Fukushima, Atsuki

    2014-12-01

    The dry-eye drug rebamipide has mucin secretagogue activity in and anti-inflammatory effects on corneal epithelial cells. Corneal stromal fibroblasts (transdifferentiated keratocytes) function as immune modulators in the pathogenesis of chronic ocular allergic inflammation and in innate immune responses at the ocular surface. The possible anti-inflammatory effects of rebamipide on human corneal stromal fibroblasts were examined. Serum-deprived cells were incubated for 1 h with rebamipide and then for various times in the additional absence or presence of cytokines or bacterial lipopolysaccharide (LPS). The release of chemokines into culture supernatants was determined with ELISAs. The intracellular abundance of chemokine mRNAs was quantitated by reverse transcription and real-time PCR analysis. Degradation of the nuclear factor κB (NFκB) inhibitor IκBα was detected by immunoblot analysis. Rebamipide suppressed the release of interleukin (IL)-8 and the upregulation of IL-8 mRNA induced by tumour necrosis factor α (TNF-α) or LPS in corneal fibroblasts. It also inhibited eotaxin-1 (CCL-11) expression at the protein and mRNA levels induced by the combination of TNF-α and IL-4. In addition, rebamipide attenuated the degradation of IκBα induced by TNF-α or LPS. Rebamipide inhibited the synthesis of chemokines by corneal fibroblasts in association with suppression of NFκB signalling. Rebamipide may therefore prove effective for the treatment of corneal stromal inflammation associated with allergy or bacterial infection. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  18. Gingival and periodontal ligament fibroblasts differ in their inflammatory response to viable Porphyromonas gingivalis

    NARCIS (Netherlands)

    Scheres, N; Laine, M L; de Vries, T J; Everts, V; van Winkelhoff, A J

    2010-01-01

    BACKGROUND AND OBJECTIVE: Porphyromonas gingivalis is an oral pathogen strongly associated with destruction of the tooth-supporting tissues in human periodontitis. Gingival fibroblasts (GF) and periodontal ligament fibroblasts (PDLF) are functionally different cell types in the periodontium that can

  19. Gingival fibroblast responsiveness is differentially affected by Porphyromonas gingivalis: implications for the pathogenesis of periodontitis

    NARCIS (Netherlands)

    Scheres, N.; Crielaard, W.

    2013-01-01

    In periodontitis, tissue damage results mainly from aberrant host responses to oral microorganisms. Fibroblasts can play an important role in this. Gingival fibroblasts do not develop tolerance against the lipopolysaccharide of Porphyromonas gingivalis, a keystone pathogen in periodontitis, which ma

  20. 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.

  1. Insights into the role of immunosenescence during varicella zoster virus infection (shingles) in the aging cell model.

    Science.gov (United States)

    Kim, Ji-Ae; Park, Seul-Ki; Kumar, Mukesh; Lee, Chan-Hee; Shin, Ok Sarah

    2015-11-03

    Varicella zoster virus (VZV) is the etiological agent of shingles, a painful skin rash that affects a significant proportion of the elderly population. In the present study, we used two aging cell models, Hutchinson-Gilford progeria syndrome (HGPS) fibroblasts and stress or replicative senescence-induced normal human dermal fibroblasts (NHDFs), to investigate age-associated susceptibility to VZV infection. VZV infectivity titers were significantly associated with donor age in HGPS fibroblasts and senescence induction in NHDFs. High throughput RNA-sequencing (RNA-seq) analysis was performed to assess global and dynamic changes in the host transcriptomes of VZV-infected aging cells. Analysis of differentially expressed genes (DEGs) indicated that VZV infection in aged HGPS fibroblasts resembled that in senescent NHDFs, particularly in terms of genes associated with pattern recognition receptors in virus sensing network, providing novel insights into the mechanisms of senescence-associated susceptibility to VZV infection. Additionally, we identified stimulator of interferon genes (STING) as a potential VZV sensing receptor. Knockdown of STING expression resulted in increased viral replication in primary fibroblasts, whereas STING overexpression led to suppression of VZV plaque formation. In conclusion, our findings highlight the important role of immunosenescence following VZV infection and provide significant insights into the mechanisms underlying cellular sensing of VZV infection and the induction of immune responses in aged skin cells.

  2. Water-filtered near-infrared influences collagen synthesis of keloid-fibroblasts in contrast to normal foreskin fibroblasts.

    Science.gov (United States)

    Zöller, Nadja; König, Anke; Butting, Manuel; Kaufmann, Roland; Bernd, August; Valesky, Eva; Kippenberger, Stefan

    2016-10-01

    Hypertrophic scar development is associated to impaired wound healing, imbalanced fibroblast proliferation and extracellular matrix synthesis. Stigmatization, physical restrictions and high recurrence rates are only some aspects that illustrate the severe influence impaired wound healing can have on patients' life. The treatment of hypertrophic scars especially keloids is still a challenge. In recent years water-filtered near-infrared irradiation (wIRA) composed of near-infrared (NIR) and a thermal component is applied for an increasing penal of clinical purposes. It is described to beneficially influence e.g. wound healing. But discrimination between the thermal and the NIR dependent components of these effects has not been conclusively elucidated. Aim of our study was therefore to investigate the influence of the light fraction on the thermal impact of wIRA irradiation in dermal cells. We concentrated our analysis on morphological properties and collagen synthesis. Foreskin fibroblasts and the keloid fibroblast cell line KF111 were exposed to temperatures between 37°C and 46°C with or without additional irradiation with 360J/cm(2) NIR. Our results show that viability was not influenced by irradiation. Independent of the analysed fibroblast species temperature dependent occurrence of spheric cells could be observed. These morphological changes were clearly counteracted by additional light exposure. Convective heat reduced collagen type I synthesis in both cell species depending on the applied temperature. Co-treatment with NIR significantly reversed this effect in keloid fibroblast cultures treated at 46°C whereas no difference could be observed in the foreskin fibroblasts. The observed influence on collagen type I synthesis was associated to a temperature dependent TGF-β1 secretion reduction. Co-stimulation of keloid cultures with NIR at 46°C completely abolished the temperature dependent TGF-β1 secretion reduction. In foreskin fibroblast cultures co

  3. Rate of sister chromatid exchanges in Bloom syndrome fibroblasts reduced by co-cultivation with normal fibroblasts.

    OpenAIRE

    1980-01-01

    Six strains of Bloom syndrome (BlS) fibroblasts responded to co-cultivation with normal fibroblasts at a 1:2 ratio by a reduced rate of sister chromatid exchanges (SCE's) from a mean of 67.5 (range = 59--78) to 28.4 (range = 21--35). The response was dose-dependent in one strain tested at 1:2, 1:1, and 2:1 ratios. In addition, quadriradial exchange figures and other signs of increased chromosomal instability were not found in BlS cells following co-cultivation with control cells. Control cell...

  4. Growth in agarose of human cells infected with cytomegalovirus.

    Science.gov (United States)

    Lang, D J; Montagnier, L; Latarjet, R

    1974-08-01

    After infection by human cytomegalovirus (CMV), human diploid fibroblasts could grow in agarose medium for several generations. Clones of infected cells grew for weeks, although in every case they ultimately underwent lysis owing to the cytopathic effect of the virus. Virus was inoculated at high dilution and after UV irradiation in an effort to derive cells infected with noninfectious defective particles still capable of inducing cell stimulation. Dilute or irradiated virus occasionally yielded large colonies of replicating cells, although permanent transformation was not observed. One clone derived from UV-CMV-infected cells was passaged four times before undergoing lysis. During these passages the cells exhibited alterations in morphology and orientation.

  5. Cancer-Associated Fibroblasts: Perspectives in Cancer Therapy

    NARCIS (Netherlands)

    Prakash, Jai

    2016-01-01

    The interplay between cancer cells and stromal cells is increasingly recognized as a main driver of tumor progression and metastasis. This Forum article highlights the role of cancer-associated stromal fibroblasts (CAFs) in tumorigenesis and discusses the potential for developing specific stromal ca

  6. Fibroblast growth factor 23 and dietary factors in renal disease

    NARCIS (Netherlands)

    da Cunha Baia, Leandro

    2015-01-01

    Omega-3 poly-unsaturated fatty acids and mineral metabolism: novel therapy for cardiovascular disease in renal patients? Deregulations in mineral metabolism, particularly related to phosphate and its regulating hormone fibroblast growth factor 23 (FGF23), are common in patients with chronic kidney d

  7. Airway smooth muscle and fibroblasts in the pathogenesis of asthma

    NARCIS (Netherlands)

    Johnson, Peter R A; Burgess, Janette K

    2004-01-01

    Asthma is a disease characterized by marked structural changes within the airway wall. These changes include deposition of extracellular matrix proteins and an increase in the numbers of airway smooth muscle cells and subepithelial fibroblasts. Both these cell types possess properties that would ena

  8. Fibroblast Growth Factor-2 Alters the Nature of Extinction

    Science.gov (United States)

    Graham, Bronwyn M.; Richardson, Rick

    2011-01-01

    These experiments examined the effects of the NMDA-receptor (NMDAr) antagonist MK801 on reacquisition and re-extinction of a conditioned fear that had been previously extinguished before injection of fibroblast growth factor-2 (FGF2) or vehicle. Recent findings have shown that relearning and re-extinction, unlike initial learning and extinction,…

  9. DETACHMENT OF HUMAN FIBROBLASTS FROM FEP-TEFLON SURFACES

    NARCIS (Netherlands)

    VANKOOTEN, TG; SCHAKENRAAD, JM; VANDERMEI, HC; BUSSCHER, HJ

    1991-01-01

    In this study a comparison is made between the detachment behavior of human fibroblasts adhered to hydrophobic FEP-Teflon (water contact angle 109 degrees) and to hydrophilic glass (water contact angle smaller than 15 degrees) during exposure to a laminar, incrementally loaded flow. Detachment from

  10. Primary skin fibroblasts as a model of Parkinson's disease

    NARCIS (Netherlands)

    Auburger, G.; Klinkenberg, M.; Droste, J.A.H.; Marcus, K.; Morales-Gordo, B.; Kunz, W.S.; Brandt, U.; Broccoli, V.; Reichmann, H.; Gispert, S.; Jendrach, M.

    2012-01-01

    Parkinson's disease is the second most frequent neurodegenerative disorder. While most cases occur sporadic mutations in a growing number of genes including Parkin (PARK2) and PINK1 (PARK6) have been associated with the disease. Different animal models and cell models like patient skin fibroblasts

  11. Regulation of NAMPT in Human Gingival Fibroblasts and Biopsies

    Directory of Open Access Journals (Sweden)

    Anna Damanaki

    2014-01-01

    Full Text Available Adipokines, such as nicotinamide phosphoribosyltransferase (NAMPT, are molecules, which are produced in adipose tissue. Recent studies suggest that NAMPT might also be produced in the tooth-supporting tissues, that is, periodontium, which also includes the gingiva. The aim of this study was to examine if and under what conditions NAMPT is produced in gingival fibroblasts and biopsies from healthy and inflamed gingiva. Gingival fibroblasts produced constitutively NAMPT, and this synthesis was significantly increased by interleukin-1β and the oral bacteria P. gingivalis and F. nucleatum. Inhibition of the MEK1/2 and NFκB pathways abrogated the stimulatory effects of F. nucleatum on NAMPT. Furthermore, the expression and protein levels of NAMPT were significantly enhanced in gingival biopsies from patients with periodontitis, a chronic inflammatory infectious disease of the periodontium, as compared to gingiva from periodontally healthy individuals. In summary, the present study provides original evidence that gingival fibroblasts produce NAMPT and that this synthesis is increased under inflammatory and infectious conditions. Local synthesis of NAMPT in the inflamed gingiva may contribute to the enhanced gingival and serum levels of NAMPT, as observed in periodontitis patients. Moreover, local production of NAMPT by gingival fibroblasts may represent a possible mechanism whereby periodontitis may impact on systemic diseases.

  12. High molecular weight plant heteropolysaccharides stimulate fibroblasts but inhibit keratinocytes.

    Science.gov (United States)

    Shahbuddin, Munira; Shahbuddin, Dahlia; Bullock, Anthony J; Ibrahim, Halijah; Rimmer, Stephen; MacNeil, Sheila

    2013-06-28

    Konjac glucomannan (KGM) is a natural polysaccharide of β(1-4)-D-glucomannopyranosyl backbone of D-mannose and D-glucose derived from the tuber of Amorphophallus konjac C. Koch. KGM has been reported to have a wide range of activities including wound healing. In this study we examined KGM extracts prepared from five plant species, (Amorphophallus konjac Koch, Amorphophallus oncophyllus, Amorphophallus prainii, Amorphophallus paeoniifolius and Amorphophallus elegans) for their effects on cultured human keratinocytes and fibroblasts. Extracts from A. konjac Koch, A. oncophyllus and A. prainii (but not from A. paeoniifolius or A. elegans) stimulated fibroblast proliferation both in the absence and presence of serum. However, these materials inhibited keratinocyte proliferation. The fibroblast stimulatory activity was associated with high molecular weight fractions of KGM and was lost following ethanol extraction or enzyme digestion with β-mannanase. It was also reduced by the addition of concanavalin A but not mannose suggesting that these heteropolysaccharides are acting on lectins but not via receptors specific to mannose. The most dramatic effect of KGM was seen in its ability to support fibroblasts for 3weeks under conditions of deliberate media starvation. This effect did not extend to supporting keratinocytes under conditions of media starvation but KGM did significantly help support adipose derived stem cells under media starvation conditions. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Modeling action potential generation and propagation in NRK fibroblasts

    NARCIS (Netherlands)

    Torres, J.J.; Cornelisse, L.N.; Harks, E.G.; Meerwijk, W.P. van; Theuvenet, A.P.R.; Ypey, D.L.

    2004-01-01

    Normal rat kidney (NRK) fibroblasts change their excitability properties through the various stages of cell proliferation. The present mathematical model has been developed to explain excitability of quiescent (serum deprived) NRK cells. It includes as cell membrane components, on the basis of patch

  14. Cloned goats (Gapra hircus) from foetal fibroblast cell lines

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Mammalian cloning has been one of the most active research topics in the world.Cloning with in vitro culured foetal fibroblast cells,in comparison with embryonic cells,can be used not only to theoretically study the embryonic or cellular development and differentiation in mammals,but also to utilize the unlimited fibroblast cells to produce large numbers of clonings.The preliminary results are as follows:(i) The division and development of the cloned embryos with embryonic donor cells and goat foetal fibroblast donor cells were 55%,77% and 35%,31%,respectively.There is no significant statistical difference between them.(ii) These studies result in the birth of two cloned goats derived from two 30-day foetal fibroblast cell lines,which are the first cloned mammals from somatic cells in China.This project has established a technological data base for the furture research on adult mammalian somatic cloning and nucleocytoplasmic interactions in animal development,and a novel technique for the cloning of animals with a high-level expression of transgene(s).

  15. Fibroblast-like synoviocyte-chondrocyte interaction in cartilage degradation

    NARCIS (Netherlands)

    Steenvoorden, M.M.C.; Bank, R.A.; Ronday, H.K.; Toes, R.E.M.; Huizinga, T.W.J.; Groot, J. de

    2007-01-01

    Objective: In vitro models for joint diseases often focus on a single cell type, such as chondrocytes in osteoarthritis (OA) or fibroblast-like synoviocytes (synoviocytes) in rheumatoid arthritis (RA). However, these joint diseases affect the whole joint and interaction between chondrocytes and syno

  16. FIBROBLAST CYTOSKELETAL REMODELING CONTRIBUTES TO CONNECTIVE TISSUE TENSION

    Science.gov (United States)

    Langevin, Helene M.; Bouffard, Nicole A.; Fox, James R.; Palmer, Bradley M.; Wu, Junru; Iatridis, James C.; Barnes, William D.; Badger, Gary J.; Howe, Alan K.

    2011-01-01

    The viscoelastic behavior of connective tissue is generally attributed to the material properties of the extracellular matrix rather than cellular activity. We have previously shown that fibroblasts within areolar connective tissue exhibit dynamic cytoskeletal remodeling within minutes in response to tissue stretch ex vivo and in vivo. Here, we tested the hypothesis that fibroblasts, through this cytoskeletal remodeling, actively contribute to the viscoelastic behavior of the whole tissue. We measured significantly increased tissue tension when cellular function was broadly inhibited by sodium azide and when cytoskeletal dynamics were compromised by disrupting microtubules (with colchicine) or actomyosin contractility (via Rho kinase inhibition). These treatments led to a decrease in cell body cross-sectional area and cell field perimeter (obtained by joining the end of all of a fibroblast’s processes). Suppressing lamellipodia formation by inhibiting Rac-1 decreased cell body cross-sectional area but did not affect cell field perimeter or tissue tension. Thus, by changing shape, fibroblasts can dynamically modulate the viscoelastic behavior of areolar connective tissue through Rho-dependent cytoskeletal mechanisms. These results have broad implications for our understanding of the dynamic interplay of forces between fibroblasts and their surrounding matrix, as well as for the neural, vascular and immune cell populations residing within connective tissue. PMID:20945345

  17. Involvement of the mitochondrial compartment in human NCL fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Pezzini, Francesco; Gismondi, Floriana [Department of Neurological, Psychological, Morphological and Motor Sciences, Divisions of Neurology (Child Neurology) and Neuropathology, University of Verona Medical School, Verona (Italy); Tessa, Alessandra [IRCCS Fondazione Stella Maris-Molecular Medicine Unit, Pisa (Italy); Tonin, Paola [Department of Neurological, Psychological, Morphological and Motor Sciences, Divisions of Neurology (Child Neurology) and Neuropathology, University of Verona Medical School, Verona (Italy); Carrozzo, Rosalba [IRCCS Bambino Gesu Hospital-Molecular Medicine Unit, Roma (Italy); Mole, Sara E. [MRC Laboratory for Molecular Cell Biology, Molecular Medicines Unit, UCL Institute of Child Health and Department of Genetics, Evolution and Environment, University College London (United Kingdom); Santorelli, Filippo M. [IRCCS Fondazione Stella Maris-Molecular Medicine Unit, Pisa (Italy); Simonati, Alessandro, E-mail: alessandro.simonati@univr.it [Department of Neurological, Psychological, Morphological and Motor Sciences, Divisions of Neurology (Child Neurology) and Neuropathology, University of Verona Medical School, Verona (Italy)

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer Mitochondrial reticulum fragmentation occurs in human CLN1 and CLN6 fibroblasts. Black-Right-Pointing-Pointer Likewise mitochondrial shift-to periphery and decreased mitochondrial density are seen. Black-Right-Pointing-Pointer Enhanced caspase-mediated apoptosis occurs following STS treatment in CLN1 fibroblasts. -- Abstract: Neuronal ceroid lipofuscinosis (NCL) are a group of progressive neurodegenerative disorders of childhood, characterized by the endo-lysosomal storage of autofluorescent material. Impaired mitochondrial function is often associated with neurodegeneration, possibly related to the apoptotic cascade. In this study we investigated the possible effects of lysosomal accumulation on the mitochondrial compartment in the fibroblasts of two NCL forms, CLN1 and CLN6. Fragmented mitochondrial reticulum was observed in all cells by using the intravital fluorescent marker Mitotracker, mainly in the perinuclear region. This was also associated with intense signal from the lysosomal markers Lysotracker and LAMP2. Likewise, mitochondria appeared to be reduced in number and shifted to the cell periphery by electron microscopy; moreover the mitochondrial markers VDCA and COX IV were reduced following quantitative Western blot analysis. Whilst there was no evidence of increased cell death under basal condition, we observed a significant increase in apoptotic nuclei following Staurosporine treatment in CLN1 cells only. In conclusion, the mitochondrial compartment is affected in NCL fibroblasts invitro, and CLN1 cells seem to be more vulnerable to the negative effects of stressed mitochondrial membrane than CLN6 cells.

  18. Cancer-Associated Fibroblasts: Perspectives in Cancer Therapy

    NARCIS (Netherlands)

    Prakash, Jai

    2016-01-01

    The interplay between cancer cells and stromal cells is increasingly recognized as a main driver of tumor progression and metastasis. This Forum article highlights the role of cancer-associated stromal fibroblasts (CAFs) in tumorigenesis and discusses the potential for developing specific stromal ca

  19. fibroblast growth factor, MTDH/Astrocyte elevated gene-1 ...

    African Journals Online (AJOL)

    2012-12-05

    Dec 5, 2012 ... Basic fibroblast growth factor (bFGF), also known as. FGF2 is a ..... lung metastases, but does not affect the tumor growth rate and decreases in .... Knox JD, Wolf C, McDaniel K, Clark V, Loriot M, Bowden GT, et al. Matrilysin.

  20. Primary skin fibroblasts as a model of Parkinson's disease

    NARCIS (Netherlands)

    Auburger, G.; Klinkenberg, M.; Droste, J.A.H.; Marcus, K.; Morales-Gordo, B.; Kunz, W.S.; Brandt, U.; Broccoli, V.; Reichmann, H.; Gispert, S.; Jendrach, M.

    2012-01-01

    Parkinson's disease is the second most frequent neurodegenerative disorder. While most cases occur sporadic mutations in a growing number of genes including Parkin (PARK2) and PINK1 (PARK6) have been associated with the disease. Different animal models and cell models like patient skin fibroblasts a

  1. Beryllium nitrate inhibits fibroblast migration to disrupt epimorphic regeneration.

    Science.gov (United States)

    Cook, Adam B; Seifert, Ashley W

    2016-10-01

    Epimorphic regeneration proceeds with or without formation of a blastema, as observed for the limb and skin, respectively. Inhibition of epimorphic regeneration provides a means to interrogate the cellular and molecular mechanisms that regulate it. In this study, we show that exposing amputated limbs to beryllium nitrate disrupts blastema formation and causes severe patterning defects in limb regeneration. In contrast, exposing full-thickness skin wounds to beryllium only causes a delay in skin regeneration. By transplanting full-thickness skin from ubiquitous GFP-expressing axolotls to wild-type hosts, we demonstrate that beryllium inhibits fibroblast migration during limb and skin regeneration in vivo Moreover, we show that beryllium also inhibits cell migration in vitro using axolotl and human fibroblasts. Interestingly, beryllium did not act as an immunostimulatory agent as it does in Anurans and mammals, nor did it affect keratinocyte migration, proliferation or re-epithelialization, suggesting that the effect of beryllium is cell type-specific. While we did not detect an increase in cell death during regeneration in response to beryllium, it did disrupt cell proliferation in mesenchymal cells. Taken together, our data show that normal blastema organogenesis cannot occur without timely infiltration of local fibroblasts and highlights the importance of positional information to instruct pattern formation during regeneration. In contrast, non-blastemal-based skin regeneration can occur despite early inhibition of fibroblast migration and cell proliferation.

  2. Adhesion, growth, and matrix production by fibroblasts on laminin substrates

    DEFF Research Database (Denmark)

    Couchman, J R; Höök, M; Rees, D A;

    1983-01-01

    Human embryonic skin fibroblasts have been shown to attach and spread on laminin substrates in the absence of protein synthesis and presence of fibronectin-depleted serum and anti-fibronectin antibodies. Rates of attachment and the type of spreading are virtually identical on fibronectin and lami......Human embryonic skin fibroblasts have been shown to attach and spread on laminin substrates in the absence of protein synthesis and presence of fibronectin-depleted serum and anti-fibronectin antibodies. Rates of attachment and the type of spreading are virtually identical on fibronectin...... and laminin-coated substrates with the development of microfilament bundles and focal adhesions. Antibodies to laminin, but not fibronectin, will prevent or reverse fibroblast adhesion to laminin, whereas antibodies to fibronectin but not laminin will give similar results on fibronectin-coated substrates....... These and other results indicate that fibroblasts possess distinct receptors for laminin and fibronectin which on contact with suitable substrates promote adhesion through interaction with common intermediates. This type of adhesion is compatible with subsequent growth and extracellular matrix production....

  3. Cancer-Associated Fibroblasts: Perspectives in Cancer Therapy

    NARCIS (Netherlands)

    Prakash, Jai

    2016-01-01

    The interplay between cancer cells and stromal cells is increasingly recognized as a main driver of tumor progression and metastasis. This Forum article highlights the role of cancer-associated stromal fibroblasts (CAFs) in tumorigenesis and discusses the potential for developing specific stromal

  4. Exposure to biomass smoke extract enhances fibronectin release from fibroblasts

    NARCIS (Netherlands)

    Krimmer, David; Ichimaru, Yukikazu; Burgess, Janette; Black, Judith; Oliver, Brian

    2013-01-01

    COPD induced following biomass smoke exposure has been reported to be associated with a more fibrotic phenotype than cigarette smoke induced COPD. This study aimed to investigate if biomass smoke induced extracellular matrix (ECM) protein production from primary human lung fibroblasts in vitro. Prim

  5. Proteoglycan synthesis in normal and Lowe syndrome fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Harper, G.S.; Hascall, V.C.; Yanagishita, M.; Gahl, W.A.

    1987-04-25

    Lowe (oculocerebrorenal) syndrome (LS) is an X-linked disorder characterized by congenital cataracts, generalized hypotonia, mental retardation, and renal Fanconi syndrome. The basic defect remains unknown, but the possibility that fibroblasts express reduced sulfation of glycosaminoglycans has been studied in several laboratories. A mechanism involving overproduction of an enzyme (nucleotide pyrophosphatase) active against adenosine 3'-phosphate, 5'-phosphosulfate (PAPS) has been postulated. Decreased synthesis of normally sulfated glycosaminoglycans was also reported. We measured the synthesis of proteoglycans and glycosaminoglycans by incorporation of (/sup 3/H)glucosamine and Na/sub 2/(/sup 35/)SO/sub 4/ into cultured fibroblasts from four LS patients and related it directly to the synthesis in six normal fibroblast cultures. We found that the rate of synthesis varied greatly among the normal cultures (cv, 30%), but not significantly between LS and the normal. The LS fibroblasts' ability to sulfate glycosaminoglycans was assayed as the amount of /sup 3/H-glycosaminoglycan eluting at low ionic strength on anion exchange chromatography, the amount of non-sulfated disaccharide present in chondroitinase digests of labeled proteoglycans, and the ratio of /sup 35/S to 3H incorporation into proteoglycans. Each parameter suggested that the LS cells were synthesizing normally sulfated glycosaminoglycans (e.g. % delta Di-0S, 21 +/- 6 in normal; 27 +/- 6 in LS). The cells' ability to sulfate glycosaminoglycans was tested under conditions of markedly stimulated glycosaminoglycan synthesis, by treating the cultures with a beta-D-xyloside.

  6. The Living Scar – Cardiac Fibroblasts and the Injured Heart

    Science.gov (United States)

    Rog-Zielinska, Eva A; Norris, Russell A; Kohl, Peter; Markwald, Roger

    2015-01-01

    Cardiac scars, often perceived as “dead” tissue, are very much alive, with heterocellular activity ensuring the maintenance of structural and mechanical integrity following heart injury. To form a scar, non-myocytes such as fibroblasts, proliferate and are recruited from intra- and extra-cardiac sources. Fibroblasts perform important autocrine and paracrine signalling functions. They also establish mechanical and, as is increasingly evident, electrical junctions with other cells. While fibroblasts were previously thought to act simply as electrical insulators, they may be electrically connected among themselves and, under certain circumstances, to other cells, including cardiomyocytes. A better understanding of these interactions will help target scar structure and function and facilitate the development of novel therapies aimed at modifying scar properties for patient benefit. This review explores available insight and recent concepts on fibroblast integration in the heart, and highlights potential avenues for harnessing their roles to optimise scar function following heart injury such as infarction, and therapeutic interventions such as ablation. PMID:26776094

  7. File list: InP.Epd.10.AllAg.Dermal_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.Epd.10.AllAg.Dermal_fibroblast hg19 Input control Epidermis Dermal fibroblast S...,SRX447386,SRX666594,SRX447389,SRX200051,SRX864096,SRX200053 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Epd.10.AllAg.Dermal_fibroblast.bed ...

  8. File list: NoD.Epd.05.AllAg.Dermal_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available NoD.Epd.05.AllAg.Dermal_fibroblasts mm9 No description Epidermis Dermal fibroblasts... http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/NoD.Epd.05.AllAg.Dermal_fibroblasts.bed ...

  9. File list: Oth.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Oth.05.AllAg.Fibroblasts hg19 TFs and others Others Fibroblasts SRX396266,SRX48...396251,SRX396255,SRX396257,SRX396268,SRX396260,SRX396253,SRX396256,SRX396265 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Oth.05.AllAg.Fibroblasts.bed ...

  10. File list: Oth.Oth.10.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Oth.10.AllAg.Fibroblasts hg19 TFs and others Others Fibroblasts SRX396266,SRX39...396255,SRX396268,SRX396252,SRX396257,SRX396260,SRX396256,SRX396253,SRX396265 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Oth.10.AllAg.Fibroblasts.bed ...

  11. File list: His.Oth.10.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.10.AllAg.Tail_fibroblast mm9 Histone Others Tail fibroblast SRX335673,SRX33...5668,SRX335667,SRX335674,SRX335665 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.10.AllAg.Tail_fibroblast.bed ...

  12. File list: Pol.Oth.10.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Oth.10.AllAg.Tail_fibroblast mm9 RNA polymerase Others Tail fibroblast SRX33566...6 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Pol.Oth.10.AllAg.Tail_fibroblast.bed ...

  13. File list: ALL.Oth.20.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.20.AllAg.Tail_fibroblast mm9 All antigens Others Tail fibroblast SRX306689,...5 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.20.AllAg.Tail_fibroblast.bed ...

  14. File list: Unc.Oth.10.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Oth.10.AllAg.Tail_fibroblast mm9 Unclassified Others Tail fibroblast SRX306689,...SRX306688,SRX306691,SRX306690,SRX306687,SRX306686 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Oth.10.AllAg.Tail_fibroblast.bed ...

  15. File list: Oth.Oth.50.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Oth.50.AllAg.Fibroblasts hg19 TFs and others Others Fibroblasts SRX338979,SRX48...396255,SRX481483,SRX396260,SRX396266,SRX396257,SRX396265,SRX396258,SRX396256 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Oth.50.AllAg.Fibroblasts.bed ...

  16. File list: His.Oth.05.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.05.AllAg.Fibroblast mm9 Histone Others Fibroblast SRX024353,SRX024354,SRX02...4352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.05.AllAg.Fibroblast.bed ...

  17. File list: His.Oth.10.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.10.AllAg.Fibroblasts hg19 Histone Others Fibroblasts SRX651503,SRX651504,SR...X651505,SRX651506,SRX480793,SRX480792 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.Oth.10.AllAg.Fibroblasts.bed ...

  18. File list: Unc.Oth.05.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Oth.05.AllAg.Tail_fibroblast mm9 Unclassified Others Tail fibroblast SRX306690,...SRX306689,SRX306688,SRX306691,SRX306687,SRX306686 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Oth.05.AllAg.Tail_fibroblast.bed ...

  19. File list: DNS.Oth.50.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.Oth.50.AllAg.Fibroblasts hg19 DNase-seq Others Fibroblasts SRX135564,SRX089279,...SRX100966,SRX089280,SRX100965,SRX480645,SRX480647,SRX480646,SRX480648 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Oth.50.AllAg.Fibroblasts.bed ...

  20. File list: His.Oth.05.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.05.AllAg.Tail_fibroblast mm9 Histone Others Tail fibroblast SRX335673,SRX33...5668,SRX335667,SRX335674,SRX335665 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.05.AllAg.Tail_fibroblast.bed ...

  1. File list: ALL.Oth.50.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.50.AllAg.Fibroblast mm9 All antigens Others Fibroblast SRX024353,SRX191057,...SRX024354,SRX024355,SRX024352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.50.AllAg.Fibroblast.bed ...

  2. File list: ALL.Oth.05.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.05.AllAg.Tail_fibroblast mm9 All antigens Others Tail fibroblast SRX306690,...5 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.05.AllAg.Tail_fibroblast.bed ...

  3. File list: ALL.Oth.10.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.10.AllAg.Tail_fibroblast mm9 All antigens Others Tail fibroblast SRX335673,...5 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.10.AllAg.Tail_fibroblast.bed ...

  4. File list: His.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.05.AllAg.Fibroblasts hg19 Histone Others Fibroblasts SRX651503,SRX651504,SR...X651505,SRX651506,SRX480793,SRX480792 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.Oth.05.AllAg.Fibroblasts.bed ...

  5. File list: ALL.Oth.10.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.10.AllAg.Fibroblast mm9 All antigens Others Fibroblast SRX024353,SRX191057,...SRX024355,SRX024354,SRX024352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.10.AllAg.Fibroblast.bed ...

  6. File list: His.Oth.20.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.20.AllAg.Fibroblasts hg19 Histone Others Fibroblasts SRX480793,SRX480792,SR...X651506,SRX651504,SRX651505,SRX651503 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.Oth.20.AllAg.Fibroblasts.bed ...

  7. File list: DNS.Oth.10.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.Oth.10.AllAg.Fibroblasts hg19 DNase-seq Others Fibroblasts SRX100966,SRX135564,...SRX480646,SRX480648,SRX089279,SRX480647,SRX480645,SRX089280,SRX100965 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Oth.10.AllAg.Fibroblasts.bed ...

  8. File list: ALL.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.05.AllAg.Fibroblasts hg19 All antigens Others Fibroblasts SRX396266,SRX4814...X651506,SRX480793,SRX480792,SRX396262 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.Oth.05.AllAg.Fibroblasts.bed ...

  9. File list: His.Oth.20.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.20.AllAg.Fibroblast mm9 Histone Others Fibroblast SRX024353,SRX024354,SRX02...4352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.20.AllAg.Fibroblast.bed ...

  10. File list: ALL.Oth.20.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.20.AllAg.Fibroblasts hg19 All antigens Others Fibroblasts SRX396266,SRX3389...X396267,SRX651503,SRX396255,SRX396269 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.Oth.20.AllAg.Fibroblasts.bed ...

  11. File list: ALL.Oth.10.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.10.AllAg.Fibroblasts hg19 All antigens Others Fibroblasts SRX396266,SRX3962...X396262,SRX396269,SRX396264,SRX396265 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.Oth.10.AllAg.Fibroblasts.bed ...

  12. File list: ALL.Oth.50.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.50.AllAg.Fibroblasts hg19 All antigens Others Fibroblasts SRX338979,SRX4814...X396257,SRX396265,SRX396258,SRX396256 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.Oth.50.AllAg.Fibroblasts.bed ...

  13. File list: His.Oth.50.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.50.AllAg.Fibroblasts hg19 Histone Others Fibroblasts SRX480793,SRX480792,SR...X651506,SRX651504,SRX651505,SRX651503 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/His.Oth.50.AllAg.Fibroblasts.bed ...

  14. File list: Oth.Oth.20.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Oth.20.AllAg.Fibroblasts hg19 TFs and others Others Fibroblasts SRX396266,SRX33...396252,SRX396260,SRX396268,SRX481483,SRX396256,SRX396265,SRX396253,SRX396255 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.Oth.20.AllAg.Fibroblasts.bed ...

  15. File list: His.Oth.10.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.10.AllAg.Fibroblast mm9 Histone Others Fibroblast SRX024353,SRX024354,SRX02...4352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.10.AllAg.Fibroblast.bed ...

  16. File list: ALL.Oth.20.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.20.AllAg.Fibroblast mm9 All antigens Others Fibroblast SRX191057,SRX024353,...SRX024354,SRX024355,SRX024352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.20.AllAg.Fibroblast.bed ...

  17. File list: DNS.Oth.20.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.Oth.20.AllAg.Fibroblasts hg19 DNase-seq Others Fibroblasts SRX135564,SRX089279,...SRX480648,SRX100966,SRX480645,SRX480646,SRX089280,SRX100965,SRX480647 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Oth.20.AllAg.Fibroblasts.bed ...

  18. File list: DNS.Oth.05.AllAg.Fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.Oth.05.AllAg.Fibroblasts hg19 DNase-seq Others Fibroblasts SRX100966,SRX480646,...SRX480648,SRX135564,SRX089279,SRX480647,SRX089280,SRX480645,SRX100965 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.Oth.05.AllAg.Fibroblasts.bed ...

  19. File list: His.Oth.50.AllAg.Fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.50.AllAg.Fibroblast mm9 Histone Others Fibroblast SRX024353,SRX024354,SRX02...4352 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.50.AllAg.Fibroblast.bed ...

  20. File list: Unc.Oth.50.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Oth.50.AllAg.Tail_fibroblast mm9 Unclassified Others Tail fibroblast SRX306689,...SRX306688,SRX306687,SRX306686,SRX306691,SRX306690 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Oth.50.AllAg.Tail_fibroblast.bed ...

  1. File list: Pol.Oth.50.AllAg.Tail_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Oth.50.AllAg.Tail_fibroblast mm9 RNA polymerase Others Tail fibroblast SRX33566...6 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Pol.Oth.50.AllAg.Tail_fibroblast.bed ...

  2. File list: ALL.Epd.20.AllAg.Dermal_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Epd.20.AllAg.Dermal_fibroblasts mm9 All antigens Epidermis Dermal fibroblasts S...26,SRX310225,SRX310224,SRX388187 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Epd.20.AllAg.Dermal_fibroblasts.bed ...

  3. File list: InP.Epd.50.AllAg.Dermal_fibroblast [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.Epd.50.AllAg.Dermal_fibroblast hg19 Input control Epidermis Dermal fibroblast S...,SRX200045,SRX864096,SRX200053,SRX200051,SRX447389,SRX447386 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Epd.50.AllAg.Dermal_fibroblast.bed ...

  4. File list: Oth.Epd.20.AllAg.Dermal_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Epd.20.AllAg.Dermal_fibroblasts mm9 TFs and others Epidermis Dermal fibroblasts... SRX247302,SRX382120,SRX247301,SRX247303,SRX382119,SRX310229,SRX310227,SRX310225 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.Epd.20.AllAg.Dermal_fibroblasts.bed ...

  5. Pengaruh Cairan Cultur Filtrate Fibroblast (CFF Terhadap Penyembuhan Luka; Penelitian eksperimental pada Rattus Norvegicus Galur Wistar

    Directory of Open Access Journals (Sweden)

    Oky Masir

    2012-11-01

    component that plays an important role in the healing process of fibroplasia. Culture filtrat Fibroblast (CFF is a result of fibroblast culture to be proven effect on the acceleration of wound healing in this study.Methods. This study used an experimental design method post test only control group design and randomized block design (RBD by using wistar mice. Experimental animals were divided into 4 groups, the two groups of treatment given to the area of excision wound CFF and the control group were given 0.9% NaCl solution to the excision wound area. Data processed with SPSS 16.0. Data were analyzed with categories Chi squared and numerical data by the Independent T-test.Result. From degree of wound healing is not found differences in both groups, but the changes in the network restriction greater in the treatment group. The score formation of collagen, the degree of epithelialization and the amount of neovascularisation formation at 3rd day there was no difference between the two groups. However, the observation of 7th day shows the formation of collagen, the degree of epithelialization and the amount of neovascularisation formation more in the treatment group. On the 3rd day fibrosis and 7th day showed more fibrosis in the treatment group compared to controls. In observation of the 3rd day infection showed fewer infections in the treatment group and the same infection between the two group at 7th day.Conclusion. CFF give wound healing better than NaCl.Keywords:CFF, NaCl 0,9 %, degree of wound healing.

  6. RT-qPCR-based microneutralization assay for human cytomegalovirus using fibroblasts and epithelial cells.

    Science.gov (United States)

    Wang, Xiao; Peden, Keith; Murata, Haruhiko

    2015-12-16

    Human cytomegalovirus (HCMV) is a leading cause of congenital infection that can result in serious disabilities in affected children. To facilitate HCMV vaccine development, a microscale neutralization assay based on reverse transcription quantitative PCR (RT-qPCR) was developed to quantify HCMV-neutralizing antibodies. Our approach relies on the generation of crude lysates from virus-infected cells that are amenable to direct analysis by RT-qPCR, thereby circumventing rate-limiting procedures associated with sample RNA extraction and purification. By serial passaging of the laboratory HCMV strain AD169 in epithelial cells (ARPE-19), a revertant virus with restored epithelial cell tropism, designated AD169(wt131), was obtained. AD169 and AD169(wt131) were evaluated in both epithelial cells (ARPE-19) and fibroblasts (MRC-5) by one-step RT-qPCR targeting the immediate-early gene IE1 transcript of HCMV. Expression kinetics indicated that RT-qPCR assessment could be conducted as early as 6h post-infection. Human serum samples (n=30) from healthy donors were tested for HCMV-specific IgG using a commercially available ELISA and for HCMV-neutralizing activity using our RT-qPCR-based neutralization assay. In agreement with the ELISA results, higher neutralizing activity was observed in the HCMV IgG seropositive group when compared with the HCMV IgG seronegative group. In addition, HCMV IgG seropositive human sera exhibited higher neutralizing titers using epithelial cells compared with using fibroblasts (geometric mean titers of 344 and 8 in ARPE-19 cells and MRC-5 cells, respectively). Our assay was robust to variation in input virus dose. In addition, a simple lysis buffer containing a non-ionic detergent was successfully demonstrated to be a less costly alternative to commercial reagents for cell-lysate preparation. Thus, our rapid HCMV neutralization assay may be a straightforward and flexible high-throughput tool for measuring antibody responses induced by vaccination

  7. Shigella Infections

    Science.gov (United States)

    ... Old Feeding Your 1- to 2-Year-Old Shigella Infections KidsHealth > For Parents > Shigella Infections Print A ... the Doctor en español Infecciones por Shigella About Shigella Shigella are bacteria that can infect the digestive ...

  8. Spinal Infections

    Science.gov (United States)

    ... infections may occur following surgery or spontaneously in patients with certain risk factors. Risk factors for spinal infections include poor nutrition, immune suppression, human immunodeficiency virus (HIV) infection, cancer, diabetes and obesity. Surgical risk factors ...

  9. Anti-proliferative effect of olmesartan on Tenon's capsule fibroblasts

    Directory of Open Access Journals (Sweden)

    Xuan Wang

    2016-05-01

    Full Text Available AIM: To evaluate the inhibitive effect of olmesartan to fibroblast proliferation and the anti-scarring effect in Tenon’s capsule, both in vitro and in vivo. METHODS: Human primary Tenon’s capsule fibroblasts were cultured in vitro, treated with up titrating concentrations of olmesartan. The rate of inhibition was tested with methyl thiazol tetrazolium (MTT method. Real-time PCR was performed to analyze changes in mRNA expressions of the fibrosis-related factors: matrix metalloproteinase-2 (MMP-2, tissue inhibitor of metalloproteinase (TIMP-1,2 and proliferating cell nuclear antigen (PCNA. Thirty rabbits were divided into 5 groups (3, 7, 14, 21, and 28d. A rabbit conjunctiva flap model was created in each eye. Olmesartan solution was injected subconjunctivally and then evaluated its anti-proliferation and anti-fibrosis effects through the histological morphology and immunohistochemistry of MMP-2 and PCNA in each group. Only the 7d group was treated with Masson’s trichrome to compare the neovascularization in the subconjunctiva area. RESULTS: In vitro, cultured Tenon's capsule human fibroblasts showed a dose dependent inhibition by olmesartan in MTT. Olmesartan reduced mRNA expressions of MMP-2 and PCNA but increased mRNA expressions of TIMP-1 and TIMP-2. In vivo, the rabbit eyes treated with olmesartan at 3rd, 7th, 14th and 21st days demonstrated a significant reduced expressions of MMP-2 and PCNA compared with control eye, no significant difference observed in 28th day group. The cellular proliferation and neovascularization was suppressed by olmesartan in Masson’s trichrome observation. CONCLUSION: By inhibiting fibroblasts in vitro and in vivo, olmesartan prevents the proliferation and activity of fibroblasts in scar tissue formation, which might benefit glaucoma filtering surgery.

  10. Anti-proliferative effect of olmesartan on Tenon's capsule fibroblasts.

    Science.gov (United States)

    Wang, Xuan; Fan, Ya-Zhi; Yao, Liang; Wang, Jian-Ming

    2016-01-01

    To evaluate the inhibitive effect of olmesartan to fibroblast proliferation and the anti-scarring effect in Tenon's capsule, both in vitro and in vivo. Human primary Tenon's capsule fibroblasts were cultured in vitro, treated with up titrating concentrations of olmesartan. The rate of inhibition was tested with methyl thiazol tetrazolium (MTT) method. Real-time PCR was performed to analyze changes in mRNA expressions of the fibrosis-related factors: matrix metalloproteinase-2 (MMP-2), tissue inhibitor of metalloproteinase (TIMP-1,2) and proliferating cell nuclear antigen (PCNA). Thirty rabbits were divided into 5 groups (3, 7, 14, 21, and 28d). A rabbit conjunctiva flap model was created in each eye. Olmesartan solution was injected subconjunctivally and then evaluated its anti-proliferation and anti-fibrosis effects through the histological morphology and immunohistochemistry of MMP-2 and PCNA in each group. Only the 7d group was treated with Masson's trichrome to compare the neovascularization in the subconjunctiva area. In vitro, cultured Tenon's capsule human fibroblasts showed a dose dependent inhibition by olmesartan in MTT. Olmesartan reduced mRNA expressions of MMP-2 and PCNA but increased mRNA expressions of TIMP-1 and TIMP-2. In vivo, the rabbit eyes treated with olmesartan at 3(rd), 7(th), 14(th) and 21(st) days demonstrated a significant reduced expressions of MMP-2 and PCNA compared with control eye, no significant difference observed in 28(th) day group. The cellular proliferation and neovascularization was suppressed by olmesartan in Masson's trichrome observation. By inhibiting fibroblasts in vitro and in vivo, olmesartan prevents the proliferation and activity of fibroblasts in scar tissue formation, which might benefit glaucoma filtering surgery.

  11. In vitro biomechanical strain regulation of fibroblast wound healing.

    Science.gov (United States)

    Cao, Thanh V; Hicks, Michael R; Standley, Paul R

    2013-11-01

    Strain-directed therapy such as vacuum compression and manual manipulative therapies are clinically effective, but their cellular and molecular mechanisms are not well understood. To determine the effects of modeled myofascial release (MFR) on fibroblast wound healing and to investigate the potential role of nitric oxide (NO) in mediating these responses. Using an in vitro scratch wound strain model, the authors investigated human fibroblast wound healing characteristics in response to injurious repetitive motion strain (RMS) and MFR. Secretion of NO was induced with interleukin-1β and sodium nitroprusside and inhibited with NO synthase inhibitor L-N(G)-monomethyl arginine citrate (L-NMMA) to determine the effects of NO on wound healing. Protein microarray was also performed to evaluate the expression of intracellular protein and activation of protein kinase G (PKG), extracellular signal-regulated kinase (ERK1/2), protein kinase C (PKC), and phosphoinositide 3-kinase (PI3K), the downstream effectors in the NO pathway. Fibroblasts that received RMS resulted in reduced wound closure rates (vs nonstrain, Pfibroblast wound healing. However, strained fibroblasts appeared to express increased sensitivity to NO. The authors also observed a 12.2% increase in NO secretion, an increase in PKG activation, and a downregulation of PKC and PI3K inhibitory domain in the combined strain group. If clinically translatable, these data suggest that mechanical strain such as vacuum compression therapy and manual manipulative therapy may modify PKC and PI3K to sensitize fibroblasts to NO and improve wound healing by promoting cell proliferation and migration by means of PKC and PKG signaling.

  12. Infective Endocarditis

    Science.gov (United States)

    ... Artery Disease Venous Thromboembolism Aortic Aneurysm More Infective Endocarditis Updated:Oct 10,2016 View an illustration of endocarditis Infective endocarditis (IE), also called bacterial endocarditis (BE), ...

  13. The impact of vascular endothelial growth factor and basic fibroblast growth factor on cardiac fibroblasts grown under altered gravity conditions

    DEFF Research Database (Denmark)

    Ulbrich, Claudia; Leder, Annekatrin; Pietsch, Jessica

    2010-01-01

    Myocardium is very sensitive to gravitational changes. During a spaceflight cardiovascular atrophy paired with rhythm problems and orthostatic intolerance can occur. The aim of this study was to investigate the impact of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor...

  14. Cdc42 promotes host defenses against fatal infection

    DEFF Research Database (Denmark)

    Lee, Keunwook; Boyd, Kelli L; Parekh, Diptiben V;

    2013-01-01

    The small Rho GTPase, Cdc42, regulates key signaling pathways required for multiple cell functions including maintenance of shape, polarity, proliferation, invasion, migration, differentiation and morphogenesis. As the role of Cdc42-dependent signaling in fibroblasts in vivo is unknown, we...... showed that in addition to fibroblasts, the FSP-1 cre deleted Cdc42 very efficiently in all leukocytes. Thus, by using this non-specific cre mouse we inadvertently demonstrated the importance of Cdc42 in host protection from lethal infections and suggest a critical role for this small GTPase in innate...

  15. 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.

  16. Differential Expression of Matrix Metalloproteases in Human Fibroblasts with Different Origins

    Directory of Open Access Journals (Sweden)

    Diana Lindner

    2012-01-01

    Full Text Available Fibroblasts are widely distributed cells and are responsible for the deposition of extracellular matrix (ECM components but also secrete ECM-degrading matrix metalloproteases. A finely balanced equilibrium between deposition and degradation of ECM is essential for structural integrity of tissues. In the past, fibroblasts have typically been understood as a uniform cell population with comparable functions regardless of their origin. Here, we determined growth curves of fibroblasts derived from heart, skin, and lung and clearly show the lowest proliferation rate for cardiac fibroblasts. Furthermore, we examined basal expression levels of collagen and different MMPs in these three types of fibroblasts and compared these concerning their site of origin. Interestingly, we found major differences in basal mRNA expression especially for MMP1 and MMP3. Moreover, we treated fibroblasts with TNF-α and observed different alterations under these proinflammatory conditions. In conclusion, fibroblasts show different properties in proliferation and MMP expression regarding their originated tissue.

  17. 8种硫酸化多糖对新城疫病毒感染鸡胚成纤维细胞能力的影响%Effects of eight sulfated polysaccharides on infecting chicken embryo fibroblast of Newcastle disease virus

    Institute of Scientific and Technical Information of China (English)

    王君敏; 胡元亮; 张帆; 王德云; 赵晓娜; 张靖; 刘静; 赵彪; 郝丽华

    2011-01-01

    Chinese angelica polysaccharide (CAPS)and lycium barbarum polysaccharide (LBPS)were modified by chlrulfonicpyridine method to obtain four sulfated CAPS, sCAPS0.6, sCAPS1.1, sCAPS1.9 and sCAPS2.2 , and four sulfated LBPS, sLBPS0.7,sLBPS1.1 ,sLBPS1.5 and sLBPS1.9. The effects of eight sulfated polysaccharides on cellular infectivity of Newcastle disease virus (NDV) were compared by MTT method taking the non-modified CAPS and LBPS as controls. The results showed that sulfated modification could significantly enhance the anti-virus activity of CAPS and LBPS,which was correlated with the degree of sulfation of sulfated polysaccharide. sCAPS2.2, sLBPS1.5, sCAPS1.9 and sLBPS1.9 possessed better activity and would be as the materials for further research.%用氯磺酸一吡啶法修饰当归多糖(CAPS)和枸杞多糖(LBPS),得到4种硫酸化当归多糖sCAPS06、sCAPS11、sCAPS1.9、sCAPS2.2和4种硫酸化枸杞多糖sLBPS07、sLBPS11、sLBPS15和sLBPS19.分别以CAPS和LBPS为埘照,采用MTT法比较了这8种硫酸化多糖对新城疫病毒感染鸡胚成纤维细胞能力的影响.结果表明:硫酸化修饰能显著提高当归多糖和枸杞多糖的抗病毒活性,且与硫酸基取代度有一定的相关性.sCAPS2.2、sLBPS1.5、sCAPS1.9和sLBPS1.9的活性较好,可以作为进一步研究的材料.

  18. 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.

  19. Multiple Directional Differentiation Difference of Neonatal Rat Fibroblasts from Six Organs

    Directory of Open Access Journals (Sweden)

    Yuqiao Chang

    2016-06-01

    Full Text Available Background/Aims: Fibroblasts are abundantly distributed throughout connective tissues in the body and are very important in maintaining the structural and functional integrity. Recent reports have proved that fibroblasts and mesenchymal stem cells share much more in common than previously recognized. The aim of this study was to investigate comparative studies in fibroblasts on the differences in the expression of molecular markers and differentiation capacity from different organs. Methods: Combined trypsin/collagenase enzymes digestion method was used to isolate and culture the fibroblasts derived from heart, liver, spleen, lung, kidney and skin. Cell activity was determined by methyl thiazolyl tetrazolium (MTT assay. Common molecular markers for fibroblasts such as vimentin, DDR2 and FSP1, stem cell markers nanog, c-kit and sca-1 were detected by RT-PCR, immunofluorescence and western blotting. The osteogenic, adipogenic and cardiogenic differentiations of fibroblasts were performed by inductive culture in special mediums, and analyzed by Alizarin red, Oil red O and immunofluorescence staining of cTnT respectively. Results: The proliferation rate of fibroblasts in lung was faster than in other five organs. Common molecular markers for fibroblasts were expressed differently in different organs. DDR2 was strongly expressed in fibroblasts in the heart, partly expressed in the heart, skin, liver and spleen. Interestingly, no expression of DDR2 was detected in liver and kidney. However, vimentin and FSP1 were consistently expressed in fibroblasts from skin, liver, kidney, spleen and lung. nanog expression in fibroblasts from lung was less than that from heart, skin, liver and spleen (P . c-kit expression in fibroblasts from heart, skin and kidney was higher than that from spleen (P , while the c-kit positive fibroblasts from liver was obviously higher than that from spleen (P . But sca-1 expression in fibroblasts from lung was the lowest among six

  20. The capsule of Porphyromonas gingivalis reduces the immune response of human gingival fibroblasts

    Directory of Open Access Journals (Sweden)

    van Winkelhoff Arie J

    2010-01-01

    Full Text Available Abstract Background Periodontitis is a bacterial infection of the periodontal tissues. The Gram-negative anaerobic bacterium Porphyromonas gingivalis is considered a major causative agent. One of the virulence factors of P. gingivalis is capsular polysaccharide (CPS. Non-encapsulated strains have been shown to be less virulent in mouse models than encapsulated strains. Results To examine the role of the CPS in host-pathogen interactions we constructed an insertional isogenic P. gingivalis knockout in the epimerase-coding gene epsC that is located at the end of the CPS biosynthesis locus. This mutant was subsequently shown to be non-encapsulated. K1 capsule biosynthesis could be restored by in trans expression of an intact epsC gene. We used the epsC mutant, the W83 wild type strain and the complemented mutant to challenge human gingival fibroblasts to examine the immune response by quantification of IL-1β, IL-6 and IL-8 transcription levels. For each of the cytokines significantly higher expression levels were found when fibroblasts were challenged with the epsC mutant compared to those challenged with the W83 wild type, ranging from two times higher for IL-1β to five times higher for IL-8. Conclusions These experiments provide the first evidence that P. gingivalis CPS acts as an interface between the pathogen and the host that may reduce the host's pro-inflammatory immune response. The higher virulence of encapsulated strains may be caused by this phenomenon which enables the bacteria to evade the immune system.

  1. The in vitro photosensitivity of systemic lupus erythematosus skin fibroblasts.

    Science.gov (United States)

    Zamansky, G B; Minka, D F; Deal, C L; Hendricks, K

    1985-03-01

    To investigate the role of DNA damage in the pathogenesis of systemic lupus erythematosus (SLE), we studied the ability of skin fibroblasts derived from SLE patients to recover from ultraviolet (UV) light radiation of varying wavelengths. Four of five SLE cell strains were more sensitive to UV-C (254 nm), sun lamp, and UV-A (320 to 400 nm) light than were normal cells. SLE cellular recovery was most sensitive to broad spectrum, long wavelength light. This hypersensitivity did not appear to result from the UV light activation of a clastogenic factor. Experiments which examined the DNA repair capacity of irradiated cells indicated that SLE fibroblasts may be able to excise certain DNA lesions as well as normal cells. The mechanisms responsible for the hypersensitivity of SLE cells remain under investigation.

  2. Malignant Transformation of an Aneurysmal Bone Cyst to Fibroblastic Osteosarcoma.

    Science.gov (United States)

    Kansagra, Akash P; Wan, Jennifer J; Devulapalli, Kavi K; Horvai, Andrew E; O'Donnell, Richard J; Link, Thomas M

    Aneurysmal bone cysts are uncommon primary bone tumors typically regarded as histologically and clinically benign. Malignant transformation of these lesions occurs almost exclusively in the context of prior radiation exposure. However, 4 cases of an osteosarcoma developing without prior radiation exposure have been reported. In this article, we report a fifth case of degeneration of an aneurysmal bone cyst to a fibroblastic osteosarcoma. In addition to reviewing the earlier cases, we describe the radiologic, pathologic, and immunohistochemical basis of this diagnosis.

  3. Response of dupuytren fibroblasts to different oxygen environments.

    Science.gov (United States)

    Türker, Tolga; Murphy, Erin; Kaufman, Christina L; Kutz, Joseph E; Meister, Edward A; Hoying, James B

    2013-12-01

    It is thought that local ischemia and oxygen radicals are responsible for fibroblast-to-myofibroblast cell transformation and proliferation. We hypothesized that hypoxia could differentially activate the contractility of fibroblasts from normal human palmar fascia and from fibroblasts-myofibroblasts of Dupuytren cords. Normal palmar fascia from 5 patients with carpal tunnel syndrome and Dupuytren cords from 5 patients were harvested. Cells were cultured from all tissue samples, and collagen lattices were prepared containing these cells. Oxygen treatment subgroups were created and incubated under hypoxic (1% O(2), 5% CO(2), and 94% N(2)), normoxic (21% O(2), 5% CO(2), and 74% N(2)), and hyperoxic (100% oxygen using 2.4 atm pressure twice a day for 7 d) conditions. After 7 days, each subgroup was photographed, and lattices were released from dishes. Postrelease photographs were taken immediately, 5 minutes after release, and after 1 hour. Areas of the lattices at each time point were calculated using MetaMorph software. Actin staining and live/dead cell analysis was performed. Linear repeated measures analysis of variance was used for data analysis given that contraction levels were measured over 3 distinct time points. We found a statistically significant difference between normal samples and Dupuytren samples in mean contraction levels over time. There was no statistically significant difference between tissue groups over the 3 time periods based on the oxygen treatment received. Our results showed a greater degree of contractility in Dupuytren disease cells than normal fibroblasts. However, the contraction in either group was not affected by oxygen level. Future in vivo research is needed to better understand the nature of pathophysiology of Dupuytren disease. Copyright © 2013 American Society for Surgery of the Hand. Published by Elsevier Inc. All rights reserved.

  4. Myxoinflammatory fibroblastic sarcoma: An uncommon tumour at an unusual site

    Directory of Open Access Journals (Sweden)

    Varuna Mallya

    2014-01-01

    Full Text Available Myxoinflammatory fibroblastic sarcoma is a low grade sarcoma that is composed of a mixed inflammatory infiltrate along with spindled, epithelioid and bizarre appearing cells in a background of hyaline and myxoid zones. Seen affecting the distal extremities commonly, with an equal sex predilection, these tumors are rare and require an extensive immunohistochemical work up for proper diagnosis. They have a tendency to recur.

  5. Effect of fibroblast-seeded artificial dermis on wound healing.

    Science.gov (United States)

    Jang, Joon Chul; Choi, Rak-Jun; Han, Seung-Kyu; Jeong, Seong-Ho; Kim, Woo-Kyung

    2015-04-01

    In covering wounds, efforts should include use of the safest and least invasive methods with a goal of achieving optimal functional and cosmetic outcome. The recent development of advanced technology in wound healing has triggered the use of cells and/or biological dermis to improve wound healing conditions. The purpose of the study was to evaluate the effects of fibroblast-seeded artificial dermis on wound healing efficacy.Ten nude mice were used in this study. Four full-thickness 6-mm punch wounds were created on the dorsal surface of each mouse (total, 40 wounds). The wounds were randomly assigned to one of the following 4 treatments: topical application of Dulbecco phosphate-buffered saline (control), human fibroblasts (FB), artificial dermis (AD), and human fibroblast-seeded artificial dermis (AD with FB). On the 14th day after treatment, wound healing rate and wound contraction, which are the 2 main factors determining wound healing efficacy, were evaluated using a stereoimage optical topometer system, histomorphological analysis, and immunohistochemistry.The results of the stereoimage optical topometer system demonstrated that the FB group did not have significant influence on wound healing rate and wound contraction. The AD group showed reduced wound contraction, but wound healing was delayed. The AD with FB group showed decreased wound contraction without significantly delayed wound healing. Histomorphological analysis exhibited that more normal skin structure was regenerated in the AD with FB group. Immunohistochemistry demonstrated that the AD group and the AD with FB group produced less α-smooth muscle actin than the control group, but this was not shown in the FB group.Fibroblast-seeded artificial dermis may minimize wound contraction without significantly delaying wound healing in the treatment of skin and soft tissue defects.

  6. Generation of human induced pluripotent stem cells from dermal fibroblasts

    OpenAIRE

    2008-01-01

    The generation of patient-specific pluripotent stem cells has the potential to accelerate the implementation of stem cells for clinical treatment of degenerative diseases. Technologies including somatic cell nuclear transfer and cell fusion might generate such cells but are hindered by issues that might prevent them from being used clinically. Here, we describe methods to use dermal fibroblasts easily obtained from an individual human to generate human induced pluripotent stem (iPS) cells by ...

  7. Fibroblast growth factor signaling during early vertebrate development.

    Science.gov (United States)

    Böttcher, Ralph T; Niehrs, Christof

    2005-02-01

    Fibroblast growth factors (FGFs) have been implicated in diverse cellular processes including apoptosis, cell survival, chemotaxis, cell adhesion, migration, differentiation, and proliferation. This review presents our current understanding on the roles of FGF signaling, the pathways employed, and its regulation. We focus on FGF signaling during early embryonic processes in vertebrates, such as induction and patterning of the three germ layers as well as its function in the control of morphogenetic movements.

  8. Inhibition of normal human lung fibroblast growth by beryllium.

    Science.gov (United States)

    Lehnert, N M; Gary, R K; Marrone, B L; Lehnert, B E

    2001-03-07

    Inhalation of particulate beryllium (Be) and its compounds causes chronic Be disease (CBD) in a relatively small subset ( approximately 1-6%) of exposed individuals. Hallmarks of this pulmonary disease include increases in several cell types, including lung fibroblasts, that contribute to the fibrotic component of the disorder. In this regard, enhancements in cell proliferation appear to play a fundamental role in CBD development and progression. Paradoxically, however, some existing evidence suggests that Be actually has antiproliferative effects. In order to gain further information about the effects of Be on cell growth, we: (1) assessed cell proliferation and cell cycle effects of low concentrations of Be in normal human diploid fibroblasts, and (2) investigated the molecular pathway(s) by which the cell cycle disturbing effects of Be may be mediated. Treatment of human lung and skin fibroblasts with Be added in the soluble form of BeSO(4) (0.1-100 microM) caused inhibitions of their growth in culture in a concentration-dependent manner. Such growth inhibition was found to persist, even after cells were further cultured in Be(2+)-free medium. Flow cytometric analyses of cellular DNA labeled with the DNA-binding fluorochrome DAPI revealed that Be causes a G(0)-G(1)/pre-S phase arrest. Western blot analyses indicated that the Be-induced G(0)-G(1)/pre-S phase arrest involves elevations in TP53 (p53) and the cyclin-dependent kinase inhibitor CDKN1A (p21(Waf-1,Cip1)). That Be at low concentrations inhibits the growth of normal human fibroblasts suggests the possibility of the existence of abnormal cell cycle inhibitory responses to Be in individuals who are sensitive to the metal and ultimately develop CBD.

  9. Expression of intracellular interferon-alpha confers antiviral properties in transfected bovine fetal fibroblasts and does not affect the full development of SCNT embryos.

    Directory of Open Access Journals (Sweden)

    Dawei Yu

    Full Text Available Foot-and-mouth disease, one of the most significant diseases of dairy herds, has substantial effects on farm economics, and currently, disease control measures are limited. In this study, we constructed a vector with a human interferon-α (hIFN-α (without secretory signal sequence gene cassette containing the immediate early promoter of human cytomegalovirus. Stably transfected bovine fetal fibroblasts were obtained by G418 selection, and hIFN-α transgenic embryos were produced by somatic cell nuclear transfer (SCNT. Forty-six transgenic embryos were transplanted into surrogate cows, and five cows (10.9% became pregnant. Two male cloned calves were born. Expression of hIFN-α was detected in transfected bovine fetal fibroblasts, transgenic SCNT embryos, and different tissues from a transgenic SCNT calf at two days old. In transfected bovine fetal fibroblasts, expression of intracellular IFN-α induced resistance to vesicular stomatitis virus infection, increased apoptosis, and induced the expression of double-stranded RNA-activated protein kinase gene (PKR and the 2'-5'-oligoadenylate synthetase gene (2'-5' OAS, which are IFN-inducible genes with antiviral activity. Analysis by qRT-PCR showed that the mRNA expression levels of PKR, 2'-5' OAS, and P53 were significantly increased in wild-type bovine fetal fibroblasts stimulated with extracellular recombinant human IFN-α-2b, showing that intracellular IFN-α induces biological functions similar to extracellular IFN-α. In conclusion, expression of intracellular hIFN-α conferred antiviral properties in transfected bovine fetal fibroblasts and did not significantly affect the full development of SCNT embryos. Thus, IFN-α transgenic technology may provide a revolutionary way to achieve elite breeding of livestock.

  10. Anti-biofilm activity of chitosan gels formulated with silver nanoparticles and their cytotoxic effect on human fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Pérez-Díaz, M.; Alvarado-Gomez, E. [Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi (Mexico); Magaña-Aquino, M. [Servicio de Epidemiologia del Hospital Central “Dr. Ignacio Morones Prieto”, San Luis Potosi (Mexico); Sánchez-Sánchez, R.; Velasquillo, C. [Laboratorio de Biotecnologia, Instituto Nacional de Rehabilitacion, Mexico, D.F. (Mexico); Gonzalez, C. [Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi (Mexico); Ganem-Rondero, A. [Division de Estudios de Posgrado (Tecnologia Farmaceutica), Facultad de Estudios Superiores Cuautitlan, Universidad Nacional Autonoma de Mexico, Cuautitlan Izcalli, Estado de Mexico (Mexico); Martínez-Castañon, G.; Zavala-Alonso, N. [Doctorado en Ciencias Odontológicas Facultad de Estomatologia, UASLP (Mexico); Martinez-Gutierrez, F. [Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi (Mexico)

    2016-03-01

    The development of multi-species biofilms in chronic wounds is a serious health problem that primarily generates strong resistance mechanisms to antimicrobial therapy. The use of silver nanoparticles (AgNPs) as a broad-spectrum antimicrobial agent has been studied previously. However, their cytotoxic effects limit its use within the medical area. The purpose of this study was to evaluate the anti-biofilm capacity of chitosan gel formulations loaded with AgNPs, using silver sulfadiazine (SSD) as a standard treatment, on strains of clinical isolates, as well as their cytotoxic effect on human primary fibroblasts. Multi-species biofilm of Staphylococcus aureus oxacillin resistant (MRSA) and Pseudomonas aeruginosa obtained from a patient with chronic wound infection were carried out using a standard Drip Flow Reactor (DFR) under conditions that mimic the flow of nutrients in the human skin. Anti-biofilm activity of chitosan gels and SSD showed a log-reduction of 6.0 for MRSA when chitosan gel with AgNPs at a concentration of 100 ppm was used, however it was necessary to increase the concentration of the chitosan gel with AgNPs to 1000 ppm to get a log-reduction of 3.3, while the SSD showed a total reduction of both bacteria in comparison with the negative control. The biocompatibility evaluation on primary fibroblasts showed better results when the chitosan gels with AgNPs were tested even in the high concentration, in contrast with SSD, which killed all the primary fibroblasts. In conclusion, chitosan gel formulations loaded with AgNPs effectively prevent the formation of biofilm and kill bacteria in established biofilm, which suggest that chitosan gels with AgNPs could be used for prevention and treatment of infections in chronic wounds. The statistic significance of the biocompatibility of chitosan gel formulations loaded with AgNPs represents an advance; however further research and development are necessary to translate this technology into therapeutic and

  11. Cytosolic malate dehydrogenase regulates senescence in human fibroblasts.

    Science.gov (United States)

    Lee, Seung-Min; Dho, So Hee; Ju, Sung-Kyu; Maeng, Jin-Soo; Kim, Jeong-Yoon; Kwon, Ki-Sun

    2012-10-01

    Carbohydrate metabolism changes during cellular senescence. Cytosolic malate dehydrogenase (MDH1) catalyzes the reversible reduction of oxaloacetate to malate at the expense of reduced nicotinamide adenine dinucleotide (NADH). Here, we show that MDH1 plays a critical role in the cellular senescence of human fibroblasts. We observed that the activity of MDH1 was reduced in old human dermal fibroblasts (HDFs) [population doublings (PD) 56], suggesting a link between decreased MDH1 protein levels and aging. Knockdown of MDH1 in young HDFs (PD 20) and the IMR90 human fibroblast cell line resulted in the appearance of significant cellular senescence features, including senescence-associated β-galactosidase staining, flattened and enlarged morphology, increased population doubling time, and elevated p16(INK4A) and p21(CIP1) protein levels. Cytosolic NAD/NADH ratios were decreased in old HDFs to the same extent as in MDH1 knockdown HDFs, suggesting that cytosolic NAD depletion is related to cellular senescence. We found that AMP-activated protein kinase, a sensor of cellular energy, was activated in MDH1 knockdown cells. We also found that sirtuin 1 (SIRT1) deacetylase, a controller of cellular senescence, was decreased in MDH1 knockdown cells. These results indicate that the decrease in MDH1 and subsequent reduction in NAD/NADH ratio, which causes SIRT1 inhibition, is a likely carbohydrate metabolism-controlled cellular senescence mechanism.

  12. Genomic instability of gold nanoparticle treated human lung fibroblast cells.

    Science.gov (United States)

    Li, Jasmine J; Lo, Soo-Ling; Ng, Cheng-Teng; Gurung, Resham Lal; Hartono, Deny; Hande, Manoor Prakash; Ong, Choon-Nam; Bay, Boon-Huat; Yung, Lin-Yue Lanry

    2011-08-01

    Gold nanoparticles (AuNPs) are one of the most versatile and widely researched materials for novel biomedical applications. However, the current knowledge in their toxicological profile is still incomplete and many on-going investigations aim to understand the potential adverse effects in human body. Here, we employed two dimensional gel electrophoresis to perform a comparative proteomic analysis of AuNP treated MRC-5 lung fibroblast cells. In our findings, we identified 16 proteins that were differentially expressed in MRC-5 lung fibroblasts following exposure to AuNPs. Their expression levels were also verified by western blotting and real time RT-PCR analysis. Of interest was the difference in the oxidative stress related proteins (NADH ubiquinone oxidoreductase (NDUFS1), protein disulfide isomerase associate 3 (PDIA3), heterogeneous nuclear ribonucleus protein C1/C2 (hnRNP C1/C2) and thioredoxin-like protein 1 (TXNL1)) as well as proteins associated with cell cycle regulation, cytoskeleton and DNA repair (heterogeneous nuclear ribonucleus protein C1/C2 (hnRNP C1/C2) and Secernin-1 (SCN1)). This finding is consistent with the genotoxicity observed in the AuNP treated lung fibroblasts. These results suggest that AuNP treatment can induce oxidative stress-mediated genomic instability.

  13. Stretching Fibroblasts Remodels Fibronectin and Alters Cancer Cell Migration

    Science.gov (United States)

    Ao, Mingfang; Brewer, Bryson M.; Yang, Lijie; Franco Coronel, Omar E.; Hayward, Simon W.; Webb, Donna J.; Li, Deyu

    2015-02-01

    Most investigations of cancer-stroma interactions have focused on biochemical signaling effects, with much less attention being paid to biophysical factors. In this study, we investigated the role of mechanical stimuli on human prostatic fibroblasts using a microfluidic platform that was adapted for our experiments and further developed for both repeatable performance among multiple assays and for compatibility with high-resolution confocal microscopy. Results show that mechanical stretching of normal tissue-associated fibroblasts (NAFs) alters the structure of secreted fibronectin. Specifically, unstretched NAFs deposit and assemble fibronectin in a random, mesh-like arrangement, while stretched NAFs produce matrix with a more organized, linearly aligned structure. Moreover, the stretched NAFs exhibited an enhanced capability for directing co-cultured cancer cell migration in a persistent manner. Furthermore, we show that stretching NAFs triggers complex biochemical signaling events through the observation of increased expression of platelet derived growth factor receptor α (PDGFRα). A comparison of these behaviors with those of cancer-associated fibroblasts (CAFs) indicates that the observed phenotypes of stretched NAFs are similar to those associated with CAFs, suggesting that mechanical stress is a critical factor in NAF activation and CAF genesis.

  14. Histamine inhibits differentiation of skin fibroblasts into myofibroblasts.

    Science.gov (United States)

    Lin, Lin; Yamagata, Kaoru; Nakayamada, Shingo; Sawamukai, Norifumi; Yamaoka, Kunihiro; Sakata, Kei; Nakano, Kazuhisa; Tanaka, Yoshiya

    2015-07-31

    Histamine and TGF-β, major mediators secreted by mast cells, are involved in skin inflammation and play critical roles in the pathogenesis of systemic sclerosis. However, the roles of signaling mechanisms in the development of skin fibrosis remain largely unclear. Here we show that histamine suppressed the expression of α smooth muscle actin (αSMA), a marker of myofibroblasts, induced by TGF-β1 in skin fibroblasts. Histamine H1-receptor (H1R), but not H2-receptor (H2R) or H4-receptor (H4R), was expressed on skin fibroblasts at both mRNA and protein levels. Interestingly, an H1R antagonist, but not H2R or H4R antagonists, antagonized the histamine-mediated suppression of αSMA expression by TGF-β1. Correspondingly, phosphorylated Smad2 was detected after treatment with TGF-β1, whereas the addition of histamine inhibited this phosphorylation. Taken together, histamine-H1R decreased TGF-β1-mediated Smad2 phosphorylation and inhibited differentiation of skin fibroblasts into myofibroblasts.

  15. 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.

  16. Transcriptome Alterations In X-Irradiated Human Gingiva Fibroblasts.

    Science.gov (United States)

    Weissmann, Robert; Kacprowski, Tim; Peper, Michel; Esche, Jennifer; Jensen, Lars R; van Diepen, Laura; Port, Matthias; Kuss, Andreas W; Scherthan, Harry

    2016-08-01

    Ionizing radiation is known to induce genomic lesions, such as DNA double strand breaks, whose repair can lead to mutations that can modulate cellular and organismal fate. Soon after radiation exposure, cells induce transcriptional changes and alterations of cell cycle programs to respond to the received DNA damage. Radiation-induced mutations occur through misrepair in a stochastic manner and increase the risk of developing cancers years after the incident, especially after high dose radiation exposures. Here, the authors analyzed the transcriptomic response of primary human gingival fibroblasts exposed to increasing doses of acute high dose-rate x rays. In the dataset obtained after 0.5 and 5 Gy x-ray exposures and two different repair intervals (0.5 h and 16 h), the authors discovered several radiation-induced fusion transcripts in conjunction with dose-dependent gene expression changes involving a total of 3,383 genes. Principal component analysis of repeated experiments revealed that the duration of the post-exposure repair intervals had a stronger impact than irradiation dose. Subsequent overrepresentation analyses showed a number of KEGG gene sets and WikiPathways, including pathways known to relate to radioresistance in fibroblasts (Wnt, integrin signaling). Moreover, a significant radiation-induced modulation of microRNA targets was detected. The data sets on IR-induced transcriptomic alterations in primary gingival fibroblasts will facilitate genomic comparisons in various genotoxic exposure scenarios.

  17. 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.

  18. Radiation-Induced Differentiation in Human Lung Fibroblast

    Energy Technology Data Exchange (ETDEWEB)

    Park, Sa-Rah; Ahn, Ji-Yeon; Han, Young-Soo; Shim, Jie-Young; Yun, Yeon-Sook; Song, Jie-Young [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2007-10-15

    One of the most common tumors in many countries is lung cancer and patients with lung cancer may take radiotherapy. Although radiotherapy may have its own advantages, it can also induce serious problems such as acute radiation pneumonitis and pulmonary fibrosis. Pulmonary fibrosis is characterized by excessive production of {alpha}-SMA and accumulation of extracellular matrix (ECM) such as collagen and fibronectin. There has been a great amount of research about fibrosis but the exact mechanism causing the reaction is not elucidated especially in radiation-induced fibrosis. Until now it has been known that several factors such as transforming growth factor (TGF-{beta}), tumor necrosis factor (TNF), interleukin (IL)-1, IL-6, platelet-derived growth factor (PDGF) and fibroblast growth factor (FGF) are related to fibrosis. Among them TGF-{beta} with Smad signaling is known to be the main stream and other signaling molecules such as MAPK, ERK and JNK (3) also participates in the process. In addition to those above factors, it is thought that more diverse and complicate mechanisms may involve in the radiationinduced fibrosis. Therefore, to investigate the underlying mechanisms in radiation induced fibrosis, first of all, we confirmed whether radiation induces trans differentiation in human normal lung fibroblasts. Here, we suggest that not only TGF-{beta} but also radiation can induce trans differentiation in human lung fibroblast WI-38 and IMR-90.

  19. The Effect of Gynostemma pentaphyllum Extract on Mouse Dermal Fibroblasts.

    Science.gov (United States)

    Lobo, Sara Nadia; Qi, Yu Qing; Liu, Quan Zhong

    2014-01-01

    Background. The objective of this paper is to demonstrate the effect of Gynostemma pentaphyllum extract on mouse dermal fibroblasts. Recent studies have shown that this plant may possess great antioxidant properties, which can be very beneficial in combating oxidative stress. Methods. Gynostemma pentaphyllum extract was prepared and mouse dermal fibroblasts were obtained and cultured as per our laboratory protocols. Twelve samples of cells were cultured under the same conditions and both negative and positive controls were established. Induction of oxidative stress was carried out using ultraviolet C (UVC) light. Viable cell count was carried out, using microscopy. The analysis of the overall results was processed using SPSS version 16.0. Results. Statistical analysis showed strong positive correlation between the concentration of Gynostemma pentaphyllum and the mean duration of cell viability (rs = 1), with a high level of statistical significance (P Gynostemma pentaphyllum extract prolongs viability of mouse dermal fibroblasts damaged by UVC light-induced oxidative stress. The results show the potential benefits of this extract on dermal cell aging.

  20. Carcinoma-Associated Fibroblasts Are a Promising Therapeutic Target

    Energy Technology Data Exchange (ETDEWEB)

    Togo, Shinsaku, E-mail: shinsaku@juntendo.ac.jp [Department of Respiratory Medicine, Juntendo University School of Medicine, Tokyo 113-8412 (Japan); Polanska, Urszula M. [CR-UK Stromal-Tumour Interaction Group, Paterson Institute for Cancer Research, The University of Manchester, Wilmslow Road, Manchester M20 4BX (United Kingdom); Horimoto, Yoshiya [Department of Pathology and Oncology, Juntendo University School of Medicine, Tokyo 113-8412 (Japan); Atopy Research Centre, Juntendo University School of Medicine, Tokyo 113-8412 (Japan); Department of Breast Oncology, Juntendo University School of Medicine, Tokyo 113-8412 (Japan); Orimo, Akira, E-mail: shinsaku@juntendo.ac.jp [CR-UK Stromal-Tumour Interaction Group, Paterson Institute for Cancer Research, The University of Manchester, Wilmslow Road, Manchester M20 4BX (United Kingdom); Department of Pathology and Oncology, Juntendo University School of Medicine, Tokyo 113-8412 (Japan); Atopy Research Centre, Juntendo University School of Medicine, Tokyo 113-8412 (Japan)

    2013-01-31

    Human carcinomas frequently exhibit significant stromal reactions such as the so-called “desmoplastic stroma” or “reactive stroma”, which is characterised by the existence of large numbers of stromal cells and extracellular matrix proteins. Carcinoma-associated fibroblasts (CAFs), which are rich in activated fibroblast populations exemplified by myofibroblasts, are among the predominant cell types present within the tumour-associated stroma. Increased numbers of stromal myofibroblasts are often associated with high-grade malignancies with poor prognoses in humans. CAF myofibroblasts possess abilities to promote primary tumour development, growth and progression by stimulating the processes of neoangiogenesis as well as tumour cell proliferation, survival, migration and invasion. Moreover, it has been demonstrated that CAFs serve as a niche supporting the metastatic colonisation of disseminated carcinoma cells in distant organs. Their contribution to primary and secondary malignancies makes these fibroblasts a potential therapeutic target and they also appear to be relevant to the development of drug resistance and tumour recurrence. This review summarises our current knowledge of tumour-promoting CAFs and discusses the therapeutic feasibility of targeting these cells as well as disrupting heterotypic interactions with other cell types in tumours that may improve the efficacy of current anti-tumour therapies.

  1. Responses of fibroblasts and glial cells to nanostructured platinum surfaces

    Science.gov (United States)

    Pennisi, C. P.; Sevcencu, C.; Dolatshahi-Pirouz, A.; Foss, M.; Lundsgaard Hansen, J.; Nylandsted Larsen, A.; Zachar, V.; Besenbacher, F.; Yoshida, K.

    2009-09-01

    The chronic performance of implantable neural prostheses is affected by the growth of encapsulation tissue onto the stimulation electrodes. Encapsulation is associated with activation of connective tissue cells at the electrode's metallic contacts, usually made of platinum. Since surface nanotopography can modulate the cellular responses to materials, the aim of the present work was to evaluate the 'in vitro' responses of connective tissue cells to platinum strictly by modulating its surface nanoroughness. Using molecular beam epitaxy combined with sputtering, we produced platinum nanostructured substrates consisting of irregularly distributed nanopyramids and investigated their effect on the proliferation, cytoskeletal organization and cellular morphology of primary fibroblasts and transformed glial cells. Cells were cultured on these substrates and their responses to surface roughness were studied. After one day in culture, the fibroblasts were more elongated and their cytoskeleton less mature when cultured on rough substrates. This effect increased as the roughness of the surface increased and was associated with reduced cell proliferation throughout the observation period (4 days). Morphological changes also occurred in glial cells, but they were triggered by a different roughness scale and did not affect cellular proliferation. In conclusion, surface nanotopography modulates the responses of fibroblasts and glial cells to platinum, which may be an important factor in optimizing the tissue response to implanted neural electrodes.

  2. Mechanism of induction of fibroblast to corneal endothelial cell.

    Science.gov (United States)

    Jiang, Yan; Fu, Wei-Cai; Zhang, Lin

    2014-08-01

    To explore mechanism of nduction of fibroblast to corneal endothelial cell. Rabbit conjunctiva fibroblasts were used as feeder cells, rabbit oral mucosa epithelial cells were used as seed cells, and human denuded amniotic membrane was used as carrier to establish tissue engineering corneal endothelium. The transformation effect was observed. As concentration of mitomycin C increased, cell survival rate gradually decreased, cell proliferation was obviously inhibited when concentration≥25 μg/mL; 5 days after being treated by 5 μg/mL mitomycin C, cell body was enlarged and extended without cell fusion, however after being treated by 0.5 μg/mL mitomycin C, cell body was significantly proliferated and gradually fused; after 3 weeks of culture, stratified epithelium appeared on rabbit oral mucosa epithelial cells, differentiation layers were 4-5 and were well differentiated, the morphology was similar to corneal endothelial cells; Under electron microscope, surface layer of cells were polygonal, tightly connected to another with microvilli on the border, there was hemidesmosome between basal cells and human denuded amniotic membrane. Fibroblast cells have the potential of multi-directional differentiation, effective induction can promote emergence of intercellular desmosomes between seed cells and emergence of epithelial surface microvilli, and differentiate to the corneal endothelial cell. However, clinical application still needs more research and safety evaluation. Copyright © 2014 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  3. Effect of phenytoin and age on gingival fibroblast enzymes.

    Directory of Open Access Journals (Sweden)

    Surena Vahabi

    2014-06-01

    Full Text Available The alteration of cytokine balance is stated to exert greater influence on gingival overgrowth compared to the direct effect of the drug on the regulation of extracellular matrix metabolism. The current study evaluated the effect of phenytoin on the regulation of collagen, lysyl oxidase and elastin in gingival fibroblasts.Normal human gingival fibroblasts (HGFs were obtained from 4 healthy children and 4 adults. Samples were cultured with phenytoin. MTT test was used to evaluate the proliferation and ELISA was performed to determine the level of IL1β and PGE2 production by HGFs. Total RNA of gingival fibroblasts was extracted and RT-PCR was performed on samples. Mann-Whitney U test was used to analyze the data with an alpha error level less than 0.05.There was a significant difference in the expression of elastin between the controls and treated samples in both adult and pediatric groups and also in the lysyl oxidase expression of adult controls and treated adults. No significant difference was found between collagen expression in adults.The significant difference in elastin and lysyl oxidase expression between adult and pediatric samples indicates the significant effect of age on their production.

  4. Expression of Connexin43 in Rat Epithelial Cells and Fibroblasts

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    To explore the role of connexin43 (Cx43) in gap junctional intercellular communication (GJIC) and propagated sensation along meridians, the expression of Cx43 in the rat epithelial cells and fibroblasts was studied both in vitro and in vivo. With the in vitro study, the rat epithelial cells and fibroblasts were cultured together, and the localization of Cx43 was detected by immunohistochemistry and indirect immunofluorescent cytochemistry and under confocal microscopy . And the expression of Cx43 on the surface of the cells was examined by flow cytometry. With the in vivo examination, 20 SD rats were randomized into control group (n = 10) and electrical acupuncture group (EAgroup, n=10). EA ( 0.5-1.5 V, 4-16 Hz , 30 min) was applied to"Zusanli"acupoint for 30 min at rat's hind paw, the localization of Cx43 was immunohistochemically detected.The immunohistochemical staining and indirect immunfluorescent cytochemistry showed that Cx43was localized on the surface of the cells and in the cytoplasm. The relative expression level of Cx43on the cellular membrane surfaces of the rat epithelial cells and fibroblasts, as determined by FACS, were 13.91 % and 29.53 % respectively. Our studied suggested that Cx43 might be involved in GJIC and propagated sensation along meridians.

  5. Equine sarcoids: Bovine Papillomavirus type 1 transformed fibroblasts are sensitive to cisplatin and UVB induced apoptosis and show aberrant expression of p53

    Directory of Open Access Journals (Sweden)

    Finlay Margaret

    2012-12-01

    Full Text Available Abstract Bovine papillomavirus type 1 infects not only cattle but also equids and is a causative factor in the pathogenesis of commonly occurring equine sarcoid tumours. Whilst treatment of sarcoids is notoriously difficult, cisplatin has been shown to be one of the most effective treatment strategies for sarcoids. In this study we show that in equine fibroblasts, BPV-1 sensitises cells to cisplatin-induced and UVB-induced apoptosis, a known cofactor for papillomavirus associated disease, however BPV-1 transformed fibroblasts show increased clonogenic survival, which may potentially limit the therapeutic effects of repeated cisplatin treatment. Furthermore we show that BPV-1 increases p53 expression in sarcoid cell lines and p53 expression can be either nuclear or cytoplasmic. The mechanism and clinical significance of increase/abnormal p53 expression remains to be established.

  6. Arecoline and oral keratinocytes may affect the collagen metabolism of fibroblasts.

    Science.gov (United States)

    Xia, Li; Tian-You, Ling; Yi-Jun, Gao; Dong-Sheng, Tang; Wen-Hui, Li

    2009-05-01

    The characteristic of oral submucous fibrosis (OSF) is related with the disturbance of synthesis and degradation of extracellular matrix. Arecoline, the areca nut (betel nut) component of betel quid, plays a major role in pathogenesis of OSF. But the exact mechanism how arecoline influences the collagen metabolism is unclear. Oral keratinocytes and fibroblasts were cocultured and keratinocytes were pre-treated by arecoline. Fibroblasts alone, fibroblasts stimulated by arecoline, fibroblasts cocultured with keratinocytes and fibroblasts cocultured with keratinocyte pre-treated by arecoline were included as the four groups in the present study. The concentration of collagen, the content and activity of matrix metalloproteinase (MMP) and the concentration of tissue inhibitor of metalloproteinase (TIMP) were assessed. The collagen production of fibroblasts decreased when cocultured with keratinocytes; when cocultured with keratinocytes pre-treated by arecoline, fibroblasts produced more soluble collagen than non-pretreated coculture group. MMP-9 was produced only in coculture groups. There was no significant difference in the two coculture groups. The activation ratio of pro-MMP-2 in arecoline pre-treated keratinocytes-fibroblasts coculture group was significantly higher than that of non-coculture groups, but no significant difference existed in the two coculture groups. TIMP-1 produced by arecoline pre-treated keratinocytes-fibroblasts coculture group was significantly higher than those by the other three groups. TIMP-1 and the interaction of oral keratinocytes and fibroblasts play important role in pathogenesis of OSF.

  7. A FTIR imaging characterization of fibroblasts stimulated by various breast cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Saroj Kumar

    Full Text Available It is well known that the microenvironment plays a major role in breast cancer progression. Yet, the mechanism explaining the transition from normal fibroblasts to cancer-stimulated fibroblasts remains to be elucidated. Here we report a FTIR imaging study of the effects of three different breast cancer cell lines on normal fibroblasts in culture. Fibroblast activation process was monitored by FTIR imaging and spectra compared by multivariate statistical analyses. Principal component analysis evidenced that the fibroblasts stimulated by these cancer cell lines grouped together and remained distinctly separated from normal fibroblasts indicating a modified different chemical composition in the cancer-stimulated fibroblasts. Similar changes in fibroblasts were induced by the various breast cancer cell lines belonging to different sub-types. Most significant changes were observed in the region of 2950 and 1230 cm(-1, possibly related to changes in lipids and in the 1230 cm(-1 area assigned to phosphate vibrations (nucleotides. Interestingly, the cancer-cell induced changes in the fibroblasts also occurred when there was no possible direct contact between the two cell lines in the co-culture. When contact was possible, the spectral changes were similar, suggesting that soluble factors but not direct cell-cell interactions were responsible for fibroblast activation. Overall, the results indicate that IR imaging could be used in the future for analyzing the microenvironment of breast tumors.

  8. Agent based modelling helps in understanding the rules by which fibroblasts support keratinocyte colony formation.

    Directory of Open Access Journals (Sweden)

    Tao Sun

    Full Text Available BACKGROUND: Autologous keratincoytes are routinely expanded using irradiated mouse fibroblasts and bovine serum for clinical use. With growing concerns about the safety of these xenobiotic materials, it is desirable to culture keratinocytes in media without animal derived products. An improved understanding of epithelial/mesenchymal interactions could assist in this. METHODOLOGY/PRINCIPAL FINDINGS: A keratincyte/fibroblast o-culture model was developed by extending an agent-based keratinocyte colony formation model to include the response of keratinocytes to both fibroblasts and serum. The model was validated by comparison of the in virtuo and in vitro multicellular behaviour of keratinocytes and fibroblasts in single and co-culture in Greens medium. To test the robustness of the model, several properties of the fibroblasts were changed to investigate their influence on the multicellular morphogenesis of keratinocyes and fibroblasts. The model was then used to generate hypotheses to explore the interactions of both proliferative and growth arrested fibroblasts with keratinocytes. The key predictions arising from the model which were confirmed by in vitro experiments were that 1 the ratio of fibroblasts to keratinocytes would critically influence keratinocyte colony expansion, 2 this ratio needed to be optimum at the beginning of the co-culture, 3 proliferative fibroblasts would be more effective than irradiated cells in expanding keratinocytes and 4 in the presence of an adequate number of fibroblasts, keratinocyte expansion would be independent of serum. CONCLUSIONS: A closely associated computational and biological approach is a powerful tool for understanding complex biological systems such as the interactions between keratinocytes and fibroblasts. The key outcome of this study is the finding that the early addition of a critical ratio of proliferative fibroblasts can give rapid keratinocyte expansion without the use of irradiated mouse

  9. 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.

  10. Design and characteristics of cytotoxic fibroblast growth factor 1 conjugate for fibroblast growth factor receptor-targeted cancer therapy

    Directory of Open Access Journals (Sweden)

    Szlachcic A

    2016-08-01

    Full Text Available Anna Szlachcic, Malgorzata Zakrzewska, Michal Lobocki, Piotr Jakimowicz, Jacek Otlewski Department of Protein Engineering, Faculty of Biotechnology, University of Wroclaw, Wroclaw, Poland Abstract: Fibroblast growth factor receptors (FGFRs are attractive candidate cancer therapy targets as they are overexpressed in multiple types of tumors, such as breast, prostate, bladder, and lung cancer. In this study, a natural ligand of FGFR, an engineered variant of fibroblast growth factor 1 (FGF1V, was conjugated to a potent cytotoxic drug, monomethyl auristatin E (MMAE, and used as a targeting agent for cancer cells overexpressing FGFRs, similar to antibodies in antibody–drug conjugates. The FGF1V–valine–citrulline–MMAE conjugate showed a favorable stability profile, bound FGFRs on the cell surface specifically, and efficiently released the drug (MMAE upon cleavage by the lysosomal protease cathepsin B. Importantly, the conjugate showed a prominent cytotoxic effect toward cell lines expressing FGFR. FGF1V–vcMMAE was highly cytotoxic at concentrations even an order of magnitude lower than those found for free MMAE. This effect was FGFR-specific as cells lacking FGFR did not show any increased mortality. Keywords: fibroblast growth factor 1, FGF receptor, targeted cancer therapy, cytotoxic conjugates, FGFR-dependent cancer, MMAE, auristatin

  11. Lithium Attenuates TGF-β1-Induced Fibroblasts to Myofibroblasts Transition in Bronchial Fibroblasts Derived from Asthmatic Patients

    Directory of Open Access Journals (Sweden)

    Marta Michalik

    2012-01-01

    Full Text Available Bronchial asthma is a chronic disorder accompanied by phenotypic transitions of bronchial epithelial cells, smooth muscle cells, and fibroblasts. Human bronchial fibroblasts (HBFs derived from patients with diagnosed asthma display predestination towards TGF-β-induced phenotypic switches. Since the interference between TGF-β and GSK-3β signaling contributes to pathophysiology of chronic lung diseases, we investigated the effect of lithium, a nonspecific GSK-3β inhibitor, on TGF-β1-induced fibroblast to myofibroblast transition (FMT in HBF and found that the inhibition of GSK-3β attenuates TGF-β1-induced FMT in HBF populations derived from asthmatic but not healthy donors. Cytoplasmically sequestrated β-catenin, abundant in TGF-β1/LiCl-stimulated asthmatic HBFs, most likely interacts with and inhibits the nuclear accumulation and signal transduction of Smad proteins. These data indicate that the specific cellular context determines FMT-related responses of HBFs to factors interfering with the TGF-β signaling pathway. They may also provide a mechanistic explanation for epidemiological data revealing coincidental remission of asthmatic syndromes and their recurrence upon the discontinuation of lithium therapy in certain psychiatric diseases.

  12. Hand Infections

    Science.gov (United States)

    ... a skin infection that can cause skin redness, warmth, and pain. People with cellulitis may have a ... cause the skin around the nail to be red, swollen, and tender. If the infection is treated ...

  13. Staph Infections

    Science.gov (United States)

    ... Culture Household Safety: Preventing Cuts Dealing With Cuts Osteomyelitis Tetanus First Aid: Skin Infections Toxic Shock Syndrome ... Abscess Paronychia Dealing With Cuts and Wounds Cellulitis Osteomyelitis Impetigo Staph Infections MRSA Cuts, Scratches, and Scrapes ...

  14. Streptococcal Infections

    Science.gov (United States)

    ... red rash on the body. Impetigo - a skin infection Toxic shock syndrome Cellulitis and necrotizing fasciitis (flesh-eating disease) Group B strep can cause blood infections, pneumonia and meningitis in newborns. A screening test ...

  15. Tapeworm Infection

    Science.gov (United States)

    Diseases and Conditions Tapeworm infection By Mayo Clinic Staff Tapeworm infection is caused by ingesting food or water contaminated with tapeworm eggs or larvae. If you ingest certain tapeworm eggs, they can migrate outside ...

  16. Infection Control

    Science.gov (United States)

    ... lives are lost because of the spread of infections in hospitals. Health care workers can take steps ... of infectious diseases. These steps are part of infection control. Proper hand washing is the most effective ...

  17. Tinea Infections

    Science.gov (United States)

    ... The skin may become itchy and red, with blisters and cracking of the skin. The infection may ... my sheets and towels every day? ResourcesDiagnosis and Management of Common Tinea Infections by SL Noble, Pharm. ...

  18. Campylobacter Infections

    Science.gov (United States)

    Campylobacter infection is a common foodborne illness. You get it from eating raw or undercooked poultry. You ... whether you need to take antibiotics. To prevent campylobacter infection, cook poultry thoroughly. Use a separate cutting ...

  19. Spinal infections.

    Science.gov (United States)

    Tay, Bobby K-B; Deckey, Jeffrey; Hu, Serena S

    2002-01-01

    Spinal infections can occur in a variety of clinical situations. Their presentation ranges from the infant with diskitis who is unwilling to crawl or walk to the adult who develops an infection after a spinal procedure. The most common types of spinal infections are hematogenous bacterial or fungal infections, pediatric diskitis, epidural abscess, and postoperative infections. Prompt and accurate diagnosis of spinal infections, the cornerstone of treatment, requires a high index of suspicion in at-risk patients and the appropriate evaluation to identify the organism and determine the extent of infection. Neurologic function and spinal stability also should be carefully evaluated. The goals of therapy should include eradicating the infection, relieving pain, preserving or restoring neurologic function, improving nutrition, and maintaining spinal stability.

  20. Anaerobic Infections

    Science.gov (United States)

    ... on the face and neck, sometimes after a dental infection or procedure such as a tooth extraction or ... adults of all ages. The most common are dental infections, inflammation of the abdominal lining (peritonitis), and abscesses ...

  1. Hantavirus Infections

    Science.gov (United States)

    ... but deadly viral infection. It is spread by mice and rats. They shed the virus in their ... breathe infected air or come into contact with rodents or their urine or droppings. You cannot catch ...

  2. Rotavirus Infections

    Science.gov (United States)

    Rotavirus is a virus that causes gastroenteritis. Symptoms include severe diarrhea, vomiting, fever, and dehydration. Almost all ... the U.S. are likely to be infected with rotavirus before their 5th birthday. Infections happen most often ...

  3. TGF-β1 promotes Staphylococcus aureus adhesion to and invasion into bovine mammary fibroblasts via the ERK pathway.

    Science.gov (United States)

    Zhao, Shuang; Gao, Yuanyuan; Xia, Xiaojing; Che, Yanyi; Wang, Yuping; Liu, Hongtao; Sun, Yingying; Ren, Wenbo; Han, Wenyu; Yang, Junling; Lei, Liancheng

    2017-01-25

    Fibroblasts are the structural base of mammary breast tissues. TGF-β1 can regulate the fibrotic process; however, it remains unclear whether TGF-β1 influences the susceptibility of fibroblasts to bacteria. Staphylococcus aureus (S. aureus) is a major bacterium in both chronic and subclinical mastitis in lactating cows that acts by invading host cells. To better understand the function of TGF-β1 in bovine mammary fibroblasts' (BMFBs) susceptibility to bacteria as well as the mechanisms involved, a primary BMFB model was established by treating cells with TGF-β1 followed by infection with S. aureus. The results revealed that the adhesion and invasion of S. aureus into BMFBs was significantly increased after cells were treated with 5 ng/ml TGF-β1 for 12 h. Moreover, TGF-β1 can increase Collagen I and α-SMA expression via activation of ERK signaling. However, the increased adhesion and invasion of S. aureus can be blocked by specific antibodies against either Collagen I or α-SMA, indicating that the increased adhesion and invasion are dependent on TGF-β1-induced upregulation of both Collagen I and α-SMA. Using PD98059, an ERK inhibitor, could also decrease the adhesion and invasion of S. aureus. These results indicate that TGF-β1 could promote S. aureus adhesion to and invasion into BMFBs by increasing Collagen I and α-SMA expression and may provide a novel target for controlling bovine mastitis.

  4. Adenoviral-mediated correction of methylmalonyl-CoA mutase deficiency in murine fibroblasts and human hepatocytes

    Directory of Open Access Journals (Sweden)

    Korson Mark

    2007-04-01

    Full Text Available Abstract Background Methylmalonic acidemia (MMA, a common organic aciduria, is caused by deficiency of the mitochondrial localized, 5'deoxyadenosylcobalamin dependent enzyme, methylmalonyl-CoA mutase (MUT. Liver transplantation in the absence of gross hepatic dysfunction provides supportive therapy and metabolic stability in severely affected patients, which invites the concept of using cell and gene delivery as future treatments for this condition. Methods To assess the effectiveness of gene delivery to restore the defective metabolism in this disorder, adenoviral correction experiments were performed using murine Mut embryonic fibroblasts and primary human methylmalonyl-CoA mutase deficient hepatocytes derived from a patient who harbored two early truncating mutations, E224X and R228X, in the MUT gene. Enzymatic and expression studies were used to assess the extent of functional correction. Results Primary hepatocytes, isolated from the native liver after removal subsequent to a combined liver-kidney transplantation procedure, or Mut murine fibroblasts were infected with a second generation recombinant adenoviral vector that expressed the murine methylmalonyl-CoA mutase as well as eGFP from distinct promoters. After transduction, [1-14C] propionate macromolecular incorporation studies and Western analysis demonstrated complete correction of the enzymatic defect in both cell types. Viral reconstitution of enzymatic expression in the human methylmalonyl-CoA mutase deficient hepatocytes exceeded that seen in fibroblasts or control hepatocytes. Conclusion These experiments provide proof of principle for viral correction in methylmalonic acidemia and suggest that hepatocyte-directed gene delivery will be an effective therapeutic treatment strategy in both murine models and in human patients. Primary hepatocytes from a liver that was unsuitable for transplantation provided an important resource for these studies.

  5. Proliferative effect of green light emitting diode irradiation on chicken fibroblasts in hyperglycaemic circumstances: a preliminary in vitro study

    Science.gov (United States)

    Vinck, Elke; Cagnie, Barbara; Declercq, Heidi; Cornelissen, Ria; Cambier, Dirk

    2004-09-01

    A reduced mortality due to hyperglycaemia was noted since the development of insulin treatment for type I diabetes and various oral hypoglycaemic agents for type II diabetes. Nevertheless the chronic metabolic disorder, Diabetes Mellitus, remains an important cause of morbidity and mortality due to a series of common secondary metabolic complications. Patients with diabetes have an increased tendency to develop infections of the skin. Healing of skin lesions in diabetics evolves often relatively slow and the lesions tend to be more severe than in non-diabetics. Endeavouring to accelerate the healing process of skin lesions in diabetic patients, this preliminary in vitro study investigates the efficacy of green Light Emitting Diode (LED) irradiation on fibroblast proliferation of cells in hyperglycaemic circumstances. In an attempt to imitate the diabetic environment, embryonic chicken fibroblasts were cultured in hyperglycaemic medium (30.000mg Glucose per litre Hanks Medium). LED irradiation was performed three consecutive days with a wavelength of 540 nm and a power output of 10 mW, at 0,6 cm distance from the fibroblasts. Each treatment lasted 3 minutes, resulting in a surface energy density of 0,2 J/cm2. Statistical analysis revealed that LED irradiation at the applied parameters induced a higher rate of proliferation in hyperglycaemic circumstances after irradiation than in the same circumstances without irradiation. Regarding these results the effectiveness of green LED irradiation on cells in hyperglycaemic circumstances is proven. To ensure the effectiveness and to evaluate the value of LED irradiation in vivo, further research is required.

  6. Effect of Avastin on the migration and invasion of pterygium fibroblasts

    Institute of Scientific and Technical Information of China (English)

    Wei Liu; Xiangyin Sha; Ye Wen; Wenting Zhao; Wei Luo; Zanmei Hua

    2014-01-01

    Purpose:.To evaluate the effect of Avastin on human ptery-gium fibroblast migration and invasiveness. Methods:.VEGF secretion was compared between human pterygium fibroblasts and conjunctival fibroblasts by measur-ing VEGF-A by ELISA. The influence of Avastin on HPF mi-gration and invasiveness was observed by wound scratch and Transwell migration assays..The expression of p-ERK1/2 and p-FAK was analyzed by western blotting. Results: (1)VEGF was secreted in higher amounts by human pterygium fibroblasts than by conjunctival fibroblasts. (2) Avastin treatment decreased HPF migration and invasion. (3) Avastin significantly decreased the expression of p-ERK1/2 and p-FAK in human pterygium fibroblasts. Conclusion:.Avastin can inhibit migration and invasion of HPFs by decreasing the expression of p-ERK1/2 and p-FAK.

  7. Effect of storage media on the proliferation of periodontal ligament fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Lauer, H.C.; Mueller, J.G.; Gross, J.; Horster, M.F.

    1987-07-01

    The effect of storage media, which are routinely used in replantation, upon the proliferative capacity of periodontal ligament fibroblasts, was compared with the effect of a tissue culture medium. The periodontal tissue was obtained from mandibular central incisors of White New Zealand rabbits. The experiments were performed in fibroblasts derived during second subculture. The storage media were physiologic salt solution, Ringer's solution and Rivanol; the tissue culture medium was alpha-minimum essential medium without nucleosides. The incubation period was 1 hour. (/sup 3/H)-thymidine incorporation and cell counts were taken to indicate changes in the proliferative capacity of the fibroblasts. The tissue culture experiments showed that the proliferative ability of the periodontal ligament fibroblasts was dependent upon the composition of the storage medium. Physiologic salt solution, Ringer's solution and Rivanol were unable to maintain the metabolism of the fibroblasts. alpha-MEM medium, however, was capable of stimulating proliferation of the periodontal ligament fibroblasts.

  8. In Vitro Effects of Pirfenidone on Cardiac Fibroblasts: Proliferation, Myofibroblast Differentiation, Migration and Cytokine Secretion

    OpenAIRE

    Qiang Shi; Xiaoyan Liu; Yuanyuan Bai; Chuanjue Cui; Jun Li; Yishi Li; Shengshou Hu; Yingjie Wei

    2011-01-01

    Cardiac fibroblasts (CFs) are the primary cell type responsible for cardiac fibrosis during pathological myocardial remodeling. Several studies have illustrated that pirfenidone (5-methyl-1-phenyl-2-[1H]-pyridone) attenuates cardiac fibrosis in different animal models. However, the effects of pirfenidone on cardiac fibroblast behavior have not been examined. In this study, we investigated whether pirfenidone directly modulates cardiac fibroblast behavior that is important in myocardial remode...

  9. Tracking and Increasing Viability of Topically Injected Fibroblasts Suspended in Hyaluronic Acid Filler.

    Science.gov (United States)

    You, Hi-Jin; Namgoong, Sik; Rhee, Sung-Mi; Han, Seung-Kyu

    2016-03-01

    A new injectable tissue-engineered soft tissue consisting of a mixture of hyaluronic acid (HA) filler and cultured human fibroblasts have been developed by the authors. To establish this method as a standard treatment, a further study was required to determine whether the injected fibroblasts could stay at the injected place or move to other sites. In addition, effective strategies were needed to increase viability of the injected fibroblasts. The purpose of this study was to track the injected fibroblasts and to determine the effect of adding prostaglandin E1 (PGE1) or vitamin C on the viability of fibroblasts.Human fibroblasts labeled with fluorescence dye were suspended in HA filler and injected into 4 sites on the back of nude mice. The injected bioimplants consisted of one of the 4 followings: HA filler without cells (HA group), fibroblasts suspended in HA filler (HA + FB group), PGE1-supplemented fibroblasts in HA filler (HA + FB + PGE1 group), and vitamin C-supplemented fibroblasts in HA filler (HA + FB + VC group). At 4 weeks after injection, locations and intensities of the fluorescence signals were evaluated using a live imaging system.The fluorescence signals of the fibroblast-containing groups were visible only at the injected sites without dispersing to other sites. The HA +FB + PGE1 group showed a significantly higher fluorescence signal than the HA + FB and the HA + FB +VC groups (P FB and HA + FB +VC groups (P = 0.69).The results of the current study collectively suggest that injected fibroblasts suspended in HA filler stay at the injected place without moving to other sites. In addition, PGE1 treatment may increase the remaining rhodamine B isothiocynanate dye at the injected site of the human dermal fibroblasts.

  10. 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. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. 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.

  12. Metformin Inhibits Angiotensin II-Induced Differentiation of Cardiac Fibroblasts into Myofibroblasts

    OpenAIRE

    Jian Bai; Na Zhang; Ying Hua; Bingjian Wang; Lin Ling; Albert Ferro; Biao Xu

    2013-01-01

    Differentiation of cardiac fibroblasts into myofibroblasts is a critical event in the progression of cardiac fibrosis that leads to pathological cardiac remodeling. Metformin, an antidiabetic agent, exhibits a number of cardioprotective properties. However, much less is known regarding the effect of metformin on cardiac fibroblast differentiation. Thus, in the present study, we examined the effect of metformin on angiotensin (Ang) II-induced differentiation of cardiac fibroblasts into myofibr...

  13. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2015-12-01

    Full Text Available BACKGROUND Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. METHODS In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. RESULTS Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1- fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity CONCLUSIONS Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  14. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2014-08-01

    Full Text Available Background Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. Methods In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. Results Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1-fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity Conclusions Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  15. Controlled induction of focal adhesion disassembly and migration in primary fibroblasts

    DEFF Research Database (Denmark)

    Dunlevy, J R; Couchman, J R

    1993-01-01

    Fibroblast migration is an integral component of biological processes such as wound healing and embryogenesis. Previous experiments examining fibroblast locomotion from tissue explants have shown that migrating fibroblasts lack, or contain only transient, focal adhesions (focal contacts). Focal...... adhesions are specialized regions of tight cell-matrix interaction, assembled by a complex process of transmembrane signalling. Although the explant model has been used for studying several aspects of fibroblast locomotion, it is limited by the lack of control over migration, and only a small percentage...

  16. Differential expression of wound fibrotic factors between facial and trunk dermal fibroblasts.

    Science.gov (United States)

    Kurita, Masakazu; Okazaki, Mutsumi; Kaminishi-Tanikawa, Akiko; Niikura, Mamoru; Takushima, Akihiko; Harii, Kiyonori

    2012-01-01

    Clinically, wounds on the face tend to heal with less scarring than those on the trunk, but the causes of this difference have not been clarified. Fibroblasts obtained from different parts of the body are known to show different properties. To investigate whether the characteristic properties of facial and trunk wound healing are caused by differences in local fibroblasts, we comparatively analyzed the functional properties of superficial and deep dermal fibroblasts obtained from the facial and trunk skin of seven individuals, with an emphasis on tendency for fibrosis. Proliferation kinetics and mRNA and protein expression of 11 fibrosis-associated factors were investigated. The proliferation kinetics of facial and trunk fibroblasts were identical, but the expression and production levels of profibrotic factors, such as extracellular matrix, transforming growth factor-β1, and connective tissue growth factor mRNA, were lower in facial fibroblasts when compared with trunk fibroblasts, while the expression of antifibrotic factors, such as collagenase, basic fibroblast growth factor, and hepatocyte growth factor, showed no clear trends. The differences in functional properties of facial and trunk dermal fibroblasts were consistent with the clinical tendencies of healing of facial and trunk wounds. Thus, the differences between facial and trunk scarring are at least partly related to the intrinsic nature of the local dermal fibroblasts.

  17. Actomyosin organisation for adhesion, spreading, growth and movement in chick fibroblasts

    DEFF Research Database (Denmark)

    Couchman, J R; Rees, D A

    1979-01-01

    Examination of the actomyosin structures and their relation to adhesion, movement and growth in the first fibroblasts migrating from chick heart explants shows striking differences with fibroblasts adapted to grow in culture. The latter have focal adhesions which seem to immobilize them for ancho......Examination of the actomyosin structures and their relation to adhesion, movement and growth in the first fibroblasts migrating from chick heart explants shows striking differences with fibroblasts adapted to grow in culture. The latter have focal adhesions which seem to immobilize them...

  18. Endoglin haploinsufficiency promotes fibroblast accumulation during wound healing through Akt activation.

    Directory of Open Access Journals (Sweden)

    Miguel Pericacho

    Full Text Available Accurate regulation of dermal fibroblast function plays a crucial role in wound healing. Many fibrotic diseases are characterized by a failure to conclude normal tissue repair and the persistence of fibroblasts inside lesions. In the present study we demonstrate that endoglin haploinsufficiency promotes fibroblast accumulation during wound healing. Moreover, scars from endoglin-heterozygous (Eng(+/- mice show persisting fibroblasts 12 days after wounding, which could lead to a fibrotic scar. Endoglin haploinsufficiency results in increased proliferation and migration of primary cultured murine dermal fibroblasts (MDFs. Moreover, Eng(+/- MDF have diminished responses to apoptotic signals compared with control cells. Altogether, these modifications could explain the augmented presence of fibroblasts in Eng(+/- mice wounds. We demonstrate that endoglin expression regulates Akt phosphorylation and that PI3K inhibition abolishes the differences in proliferation between endoglin haploinsufficient and control cells. Finally, persistent fibroblasts in Eng(+/- mice wound co-localize with a greater degree of Akt phosphorylation. Thus, endoglin haploinsufficiency seems to promote fibroblast accumulation during wound healing through the activation of the PI3K/Akt pathway. These studies open new non-Smad signaling pathway for endoglin regulating fibroblast cell function during wound healing, as new therapeutic opportunities for the treatment of fibrotic wounds.

  19. Endoglin haploinsufficiency promotes fibroblast accumulation during wound healing through Akt activation.

    Science.gov (United States)

    Pericacho, Miguel; Velasco, Soraya; Prieto, Marta; Llano, Elena; López-Novoa, José M; Rodríguez-Barbero, Alicia

    2013-01-01

    Accurate regulation of dermal fibroblast function plays a crucial role in wound healing. Many fibrotic diseases are characterized by a failure to conclude normal tissue repair and the persistence of fibroblasts inside lesions. In the present study we demonstrate that endoglin haploinsufficiency promotes fibroblast accumulation during wound healing. Moreover, scars from endoglin-heterozygous (Eng(+/-)) mice show persisting fibroblasts 12 days after wounding, which could lead to a fibrotic scar. Endoglin haploinsufficiency results in increased proliferation and migration of primary cultured murine dermal fibroblasts (MDFs). Moreover, Eng(+/-) MDF have diminished responses to apoptotic signals compared with control cells. Altogether, these modifications could explain the augmented presence of fibroblasts in Eng(+/-) mice wounds. We demonstrate that endoglin expression regulates Akt phosphorylation and that PI3K inhibition abolishes the differences in proliferation between endoglin haploinsufficient and control cells. Finally, persistent fibroblasts in Eng(+/-) mice wound co-localize with a greater degree of Akt phosphorylation. Thus, endoglin haploinsufficiency seems to promote fibroblast accumulation during wound healing through the activation of the PI3K/Akt pathway. These studies open new non-Smad signaling pathway for endoglin regulating fibroblast cell function during wound healing, as new therapeutic opportunities for the treatment of fibrotic wounds.

  20. Pearl extract enhances the migratory ability of fibroblasts in a wound healing model.

    Science.gov (United States)

    Li, Yi-Chen; Chen, Chi-Ruei; Young, Tai-Horng

    2013-03-01

    For 2000 years, traditional Chinese medicine has been used as a remedy for general health improvement, including the fight against aging. Pearl powder has recently been used as a health food that has antioxidant, antiaging, antiradioactive, and tonic activities for cells; it is also applied to cure aphthous ulcer, gastric ulcer, and duodenal ulcer on clinical therapy. In addition, the mother of pearl, nacre, could enhance the cell adhesion and tissue regeneration of skin fibroblasts. Fibroblast is regarded as indispensable in the processes of wound healing. Therefore, the effect of pearl extract (PL) on fibroblasts is investigated in this study. PL is produced by a room temperature super extraction system (Taiwan patent no. I271 220). DMEM medium containing PL (300 μg/mL) was used to examine the effect of migration-promoting potential on human fibroblast cell line or human primary fibroblast cells in a wound healing model in vitro. Medium containing PL (300 μg/mL) demonstrated that the migratory cell numbers of fibroblasts were three times more than that without PL, and mRNA expression of collagen type III was higher than in collagen type I in fibroblasts. It revealed a migration-promoting potential of human fibroblasts in a wound healing model in vitro. The present study found that the migration-promoting effect in PL, which could be a supplement in cell culture. These data suggest PL could be useful for enhancing the wound healing of fibroblasts.

  1. New Developments in Peritoneal Fibroblast Biology: Implications for Inflammation and Fibrosis in Peritoneal Dialysis

    Directory of Open Access Journals (Sweden)

    Janusz Witowski

    2015-01-01

    Full Text Available Uraemia and long-term peritoneal dialysis (PD can lead to fibrotic thickening of the peritoneal membrane, which may limit its dialytic function. Peritoneal fibrosis is associated with the appearance of myofibroblasts and expansion of extracellular matrix. The extent of contribution of resident peritoneal fibroblasts to these changes is a matter of debate. Recent studies point to a significant heterogeneity and complexity of the peritoneal fibroblast population. Here, we review recent developments in peritoneal fibroblast biology and summarize the current knowledge on the involvement of peritoneal fibroblasts in peritoneal inflammation and fibrosis.

  2. PKCδ inhibition normalizes the wound-healing capacity of diabetic human fibroblasts.

    Science.gov (United States)

    Khamaisi, Mogher; Katagiri, Sayaka; Keenan, Hillary; Park, Kyoungmin; Maeda, Yasutaka; Li, Qian; Qi, Weier; Thomou, Thomas; Eschuk, Danielle; Tellechea, Ana; Veves, Aris; Huang, Chenyu; Orgill, Dennis Paul; Wagers, Amy; King, George L

    2016-03-01

    Abnormal fibroblast function underlies poor wound healing in patients with diabetes; however, the mechanisms that impair wound healing are poorly defined. Here, we evaluated fibroblasts from individuals who had type 1 diabetes (T1D) for 50 years or more (Medalists, n = 26) and from age-matched controls (n = 7). Compared with those from controls, Medalist fibroblasts demonstrated a reduced migration response to insulin, lower VEGF expression, and less phosphorylated AKT (p-AKT), but not p-ERK, activation. Medalist fibroblasts were also functionally less effective at wound closure in nude mice. Activation of the δ isoform of protein kinase C (PKCδ) was increased in postmortem fibroblasts from Medalists, fibroblasts from living T1D subjects, biopsies of active wounds of living T1D subjects, and granulation tissues from mice with streptozotocin-induced diabetes. Diabetes-induced PKCD mRNA expression was related to a 2-fold increase in the mRNA half-life. Pharmacologic inhibition and siRNA-mediated knockdown of PKCδ or expression of a dominant-negative isoform restored insulin signaling of p-AKT and VEGF expression in vitro and improved wound healing in vivo. Additionally, increasing PKCδ expression in control fibroblasts produced the same abnormalities as those seen in Medalist fibroblasts. Our results indicate that persistent PKCδ elevation in fibroblasts from diabetic patients inhibits insulin signaling and function to impair wound healing and suggest PKCδ inhibition as a potential therapy to improve wound healing in diabetic patients.

  3. Smac/DIABLO regulates the apoptosis of hypertrophic scar fibroblasts.

    Science.gov (United States)

    Liu, Bao-Heng; Chen, Liang; Li, Shi-Rong; Wang, Zhen-Xiang; Cheng, Wen-Guang

    2013-09-01

    In abnormal skin wound healing, hypertrophic scars (HS) are characterized by excessive fibroblast hypercellularity and an overproduction of collagen, leading to atypical extracellular matrix (ECM) remodeling. Although the exact mechanisms of HS remain unclear, decreased HS fibroblast (HSFB) apoptosis and increased proliferation are evident in the development of HS. In this study, the contribution of the second mitochondria-derived activator of caspases/direct inhibitor of apoptosis protein (IAP)-binding protein with a low isoelectric point (pI) (Smac/DIABLO), an apoptosis-promoting protein released from the mitochondria, was investigated in human normal skin and HSFB cultures. The expression of Smac/DIABLO is usually decreased in many malignant tumors compared with normal tissues. Immunohistochemical analysis of skin tissues and the western blot analyses of fibroblasts revealed that the expression of Smac/DIABLO was lower in HS tissues compared with normal skin tissues. Of note, adenovirus-mediated Smac/DIABLO overexpression in the cultured HSFBs significantly reduced cell proliferation, as detected by the cell counting kit-8, and increased caspase-3 and -9 activity, as detected by spectrofluorimetry. In addition, it increased apoptosis, as detected by fluorescence-activated cell sorting (FACS). Furthermore, we found that the silencing of Smac with siRNA in the HSFBs induced a noticeable decrease in caspase-3 and -9 activity, leading to a significant reduction in apoptosis. In addition, the mRNA expression of type I and III pro-collagen detected in the HSFBs was significantly increased following the silencing of Smac with siRNA and was inhibited following Smac/DIABLO overexpression, as shown by real-time RT-PCR. In conclusion, Smac/DIABLO decreases the proliferation and increases the apoptosis of HSFBs. To our knowledge, the data from our study suggest for the first time that Smac/DIABLO is a novel therapeutic target for HS.

  4. Aggregatibacter actinomycetemcomitans lipopolysaccharide affects human gingival fibroblast cytoskeletal organization.

    Science.gov (United States)

    Gutiérrez-Venegas, Gloria; Contreras-Marmolejo, Luis Arturo; Román-Alvárez, Patricia; Barajas-Torres, Carolina

    2008-04-01

    The cytoskeleton is a dynamic structure that plays a key role in maintaining cell morphology and function. This study investigates the effect of bacterial wall lipopolysaccharide (LPS), a strong inflammatory agent, on the dynamics and organization of actin, tubulin, vimentin, and vinculin proteins in human gingival fibroblasts (HGF). A time-dependent study showed a noticeable change in actin architecture after 1.5 h of incubation with LPS (1 microg/ml) with the formation of orthogonal fibers and further accumulation of actin filament at the cell periphery by 24 h. When 0.01-10 microg/ml of LPS was added to human gingival fibroblast cultures, cells acquired a round, flat shape and gradually developed cytoplasmic ruffling. Lipopolysaccharides extracted from Aggregatibacter actinomycetemcomitans periodontopathogenic bacteria promoted alterations in F-actin stress fibres of human gingival cells. Normally, human gingival cells have F-actin fibres that are organized in linear distribution throughout the cells, extending along the cell's length. LPS-treated cells exhibited changes in cytoskeletal protein organization, and F-actin was reorganized by the formation of bundles underneath and parallel to the cell membrane. We also found the reorganization of the vimentin network into vimentin bundling after 1.5 h of treatment. HGF cells exhibited diffuse and granular gamma-tubulin stain. There was no change in LPS-treated HGF. However, vinculin plaques distributed in the cell body diminished after LPS treatment. We conclude that the dynamic and structured organization of cytoskeletal filaments and actin assembly in human gingival fibroblasts is altered by LPS treatment and is accompanied by a decrease in F-actin pools.

  5. Metallic nanoparticles reduce the migration of human fibroblasts in vitro

    Science.gov (United States)

    Vieira, Larissa Fernanda de Araújo; Lins, Marvin Paulo; Viana, Iana Mayane Mendes Nicácio; dos Santos, Jeniffer Estevão; Smaniotto, Salete; Reis, Maria Danielma dos Santos

    2017-03-01

    Nanoparticles have extremely wide applications in the medical and biological fields. They are being used in biosensors, local drug delivery, diagnostics, and medical therapy. However, the potential effects of nanoparticles on target cell and tissue function, apart from cytotoxicity, are not completely understood. Thus, the aim of this study was to investigate the in vitro effects of silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) on human fibroblasts with respect to their interaction with the extracellular matrix and in cell migration. Immunofluorescence analysis revealed that treatment with AgNPs or AuNPs decreased collagen and laminin production at all the concentrations tested (0.1, 1, and 10 μg/mL). Furthermore, cytofluorometric analysis showed that treatment with AgNPs reduced the percentage of cells expressing the collagen receptor very late antigen 2, α2β1 integrin (VLA-2) and the laminin receptor very late antigen 6, α6β1 integrin (VLA-6). In contrast, AuNP treatment increased and decreased the percentages of VLA-2-positive and VLA-6-positive cells, respectively, as compared to the findings for the controls. Analysis of cytoskeletal reorganization showed that treatment with both types of nanoparticles increased the formation of stress fibres and number of cell protrusions and impaired cell polarity. Fibroblasts exposed to different concentrations of AuNPs and AgNPs showed reduced migration through transwell chambers in the functional chemotaxis assay. These results demonstrated that metal nanoparticles may influence fibroblast function by negatively modulating the deposition of extracellular matrix molecules (ECM) and altering the expression of ECM receptors, cytoskeletal reorganization, and cell migration.

  6. Circadian-independent cell mitosis in immortalized fibroblasts.

    Science.gov (United States)

    Yeom, Mijung; Pendergast, Julie S; Ohmiya, Yoshihiro; Yamazaki, Shin

    2010-05-25

    Two prominent timekeeping systems, the cell cycle, which controls cell division, and the circadian system, which controls 24-h rhythms of physiology and behavior, are found in nearly all living organisms. A distinct feature of circadian rhythms is that they are temperature-compensated such that the period of the rhythm remains constant (approximately 24 h) at different ambient temperatures. Even though the speed of cell division, or growth rate, is highly temperature-dependent, the cell-mitosis rhythm is temperature-compensated. Twenty-four-hour fluctuations in cell division have also been observed in numerous species, suggesting that the circadian system is regulating the timing of cell division. We tested whether the cell-cycle rhythm was coupled to the circadian system in immortalized rat-1 fibroblasts by monitoring cell-cycle gene promoter-driven luciferase activity. We found that there was no consistent phase relationship between the circadian and cell cycles, and that the cell-cycle rhythm was not temperature-compensated in rat-1 fibroblasts. These data suggest that the circadian system does not regulate the cell-mitosis rhythm in rat-1 fibroblasts. These findings are inconsistent with numerous studies that suggest that cell mitosis is regulated by the circadian system in mammalian tissues in vivo. To account for this discrepancy, we propose two possibilities: (i) There is no direct coupling between the circadian rhythm and cell cycle but the timing of cell mitosis is synchronized with the rhythmic host environment, or (ii) coupling between the circadian rhythm and cell cycle exists in normal cells but it is disconnected in immortalized cells.

  7. Platelets stimulate fibroblast-mediated contraction of collagen gels

    Directory of Open Access Journals (Sweden)

    Lundahl Joachim

    2003-10-01

    Full Text Available Abstract Background Platelets are thought to play a role in a variety of inflammatory conditions in the lung, some of which may lead to fibrosis. In the current study we tested the hypothesis that whole platelets and platelet lysate can mediate remodelling of extracellular matrix in vitro by affecting fibroblast-mediated contraction of a collagen gel. We also sought to determine to what extent platelet-derived growth factor (PDGF and transforming growth factor-β (TGF-β contribute to this effect. Methods Washed platelets, isolated from healthy blood donors, and platelet lysate (freezing and thawing, were cast together with human lung fibroblasts in three-dimensional collagen gels. The gels were then released and cultured for four days. PDGF and TGF-β1 concentrations were measured in culture supernatants by ELISA. Results Both platelets and platelet lysate augmented fibroblast-mediated gel contraction in a time and concentration dependent manner (19.9% ± 0.1 (mean ± SEM of initial area vs. 48.0% ± 0.4 at 48 hours; P 1 and PDGF-AA/AB were released in co-culture. PDGF-AA/AB had a maximum release at 24 hours whereas TGF-β1 release increased with longer culture periods. Neutralising antibodies to these mediators partially inhibited platelet-induced gel contraction. Conclusion We conclude that platelets may promote remodelling of extracellular matrix in vitro and that PDGF and TGF-β partially mediate this effect, also indicating a role for other mediators. The findings may be an important mechanism in regulating repair processes after injury.

  8. Influence of titanium surface charge on fibroblast adhesion.

    Science.gov (United States)

    Hamdan, Mohammad; Blanco, Luis; Khraisat, Ameen; Tresguerres, Isabel F

    2006-01-01

    Although dental implants have a high success rate, failure owing to the absence of adhesion between the gingival connective tissue and the implant surface is still being reported. This study was designed to evaluate the effect of a titanium surface charge on fibroblast adhesion. An electrical chamber was custom-made to generate negative and positive surface charges on commercially pure titanium cylinders with a potential difference of 4.5 V. Twenty-seven titanium cylinders were divided into three experimental groups. In each group, cell attachment to a positively charged titanium cylinder, a negatively charged titanium cylinder, and a titanium cylinder (control) was studied at three time intervals of 15, 30, and 60 minutes. NCTC clone 929 fibroblasts were used in these experiments. The effect of the potential difference in the pH of Dulbecco's Modified Eagle Medium (DMEM) was also evaluated using two new specimens at time intervals of 15, 30, 60, and 80 minutes. The fibroblast cell attachment was more statistically significant to the positively charged titanium cylinder than the negatively charged titanium cylinder (p =.002) and the control (p=.000), whereas the cell adhesion difference between the control and the negatively charged titanium cylinder was not statistically significant (p=.808). The range of pH difference of the DMEM in the negative and positive parts of the electrical chamber was 0.46 and 0.30, respectively. Within the limitations of this in vitro study, the positive surface charge of the titanium cylinder results in significantly favorable cell adhesion.

  9. Association of spirochetal infection with Morgellons disease.

    Science.gov (United States)

    Middelveen, Marianne J; Burugu, Divya; Poruri, Akhila; Burke, Jennie; Mayne, Peter J; Sapi, Eva; Kahn, Douglas G; Stricker, Raphael B

    2013-01-01

    Morgellons disease (MD) is an emerging multisystem illness characterized by skin lesions with unusual filaments embedded in or projecting from epithelial tissue. Filament formation results from abnormal keratin and collagen expression by epithelial-based keratinocytes and fibroblasts. Recent research comparing MD to bovine digital dermatitis, an animal infectious disease with similar skin features, provided clues that spirochetal infection could play an important role in the human disease as it does in the animal illness. Based on histological staining, immunofluorescent staining, electron microscopic imaging and polymerase chain reaction, we report the detection of Borrelia spirochetes in dermatological tissue of  four randomly-selected MD patients. The association of MD with spirochetal infection provides evidence that this infection may be a significant factor in the illness and refutes claims that MD lesions are self-inflicted and that people suffering from this disorder are delusional. Molecular characterization of the Borrelia spirochetes found in MD patients is warranted.

  10. Glutamine deprivation induces interleukin-8 expression in ataxia telangiectasia fibroblasts.

    Science.gov (United States)

    Kim, Min-Hyun; Kim, Aryung; Yu, Ji Hoon; Lim, Joo Weon; Kim, Hyeyoung

    2014-05-01

    To investigate whether glutamine deprivation induces expression of inflammatory cytokine interleukin-8 (IL-8) by determining NF-κB activity and levels of oxidative indices (ROS, reactive oxygen species; hydrogen peroxide; GSH, glutathione) in fibroblasts isolated from patients with ataxia telangiectasia (A-T). We used A-T fibroblasts stably transfected with empty vector (Mock) or with human full-length ataxia telangiectasia mutated (ATM) cDNA (YZ5) and mouse embryonic fibroblasts (MEFs) transiently transfected with ATM small interfering RNA (siRNA) or with non-specific control siRNA. The cells were cultured with or without glutamine or GSH. ROS levels were determined using a fluorescence reader and confocal microscopy. IL-8 or murine IL-8 homolog, keratinocyte chemoattractant (KC), and hydrogen peroxide levels in the medium were determined by enzyme-linked immunosorbent assay and colorimetric assay. GSH level was assessed by enzymatic assay, while IL-8 (KC) mRNA level was measured by reverse transcription-polymerase chain reaction (RT-PCR) and/or quantitative real-time PCR. NF-κB DNA-binding activity was determined by electrophoretic mobility shift assay. Catalase activity and ATM protein levels were determined by O2 generation and Western blotting. While glutamine deprivation induced IL-8 expression and increased NF-κB DNA-binding activity in Mock cells, both processes were decreased by treatment of cells with glutamine or GSH or both glutamine and GSH. Glutamine deprivation had no effect on IL-8 expression or NF-κB DNA-binding activity in YZ5 cells. Glutamine-deprived Mock cells had higher oxidative stress indices (increases in ROS and hydrogen peroxide, reduction in GSH) than glutamine-deprived YZ5 cells. In Mock cells, glutamine deprivation-induced oxidative stress indices were suppressed by treatment with glutamine or GSH or both glutamine and GSH. GSH levels and catalase activity were lower in Mock cells than YZ5 cells. MEFs transfected with ATM siRNA and

  11. Isolation of cardiac myocytes and fibroblasts from neonatal rat pups.

    Science.gov (United States)

    Golden, Honey B; Gollapudi, Deepika; Gerilechaogetu, Fnu; Li, Jieli; Cristales, Ricardo J; Peng, Xu; Dostal, David E

    2012-01-01

    Neonatal rat ventricular myocytes (NRVM) and fibroblasts (FBs) serve as in vitro models for studying fundamental mechanisms underlying cardiac pathologies, as well as identifying potential therapeutic targets. Both cell types are relatively easy to culture as monolayers and can be manipulated using molecular and pharmacological tools. Because NRVM cease to proliferate after birth, and FBs undergo phenotypic changes and senescence after a few passages in tissue culture, primary cultures of both cell types are required for experiments. Below we describe methods that provide good cell yield and viability of primary cultures of NRVM and FBs from 0 to 3-day-old neonatal rat pups.

  12. Cancer Associated Fibroblasts and Tumor Growth: Focus on Multiple Myeloma

    Energy Technology Data Exchange (ETDEWEB)

    De Veirman, Kim, E-mail: kdeveirm@vub.ac.be [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Rao, Luigia [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Department of Biomedical Sciences and Human Oncology, Section of Internal Medicine, University of Bari Medical School, Bari I-70124 (Italy); De Bruyne, Elke; Menu, Eline; Van Valckenborgh, Els [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Van Riet, Ivan [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium); Stem Cell Laboratory, Division of Clinical Hematology, Universitair Ziekenhuis Brussel (UZ Brussel), Brussels 1090 (Belgium); Frassanito, Maria Antonia [Department of Biomedical Sciences and Human Oncology, Section of General Pathology, University of Bari Medical School, Bari I-70124 (Italy); Di Marzo, Lucia; Vacca, Angelo [Department of Biomedical Sciences and Human Oncology, Section of Internal Medicine, University of Bari Medical School, Bari I-70124 (Italy); Vanderkerken, Karin, E-mail: kdeveirm@vub.ac.be [Department of Hematology and Immunology, Myeloma Center Brussels, Vrije Universiteit Brussel (VUB), Brussels 1090 (Belgium)

    2014-06-27

    Cancer associated fibroblasts (CAFs) comprise a heterogeneous population that resides within the tumor microenvironment. They actively participate in tumor growth and metastasis by production of cytokines and chemokines, and the release of pro-inflammatory and pro-angiogenic factors, creating a more supportive microenvironment. The aim of the current review is to summarize the origin and characteristics of CAFs, and to describe the role of CAFs in tumor progression and metastasis. Furthermore, we focus on the presence of CAFs in hypoxic conditions in relation to multiple myeloma disease.

  13. Cancer Associated Fibroblasts and Tumor Growth: Focus on Multiple Myeloma

    Directory of Open Access Journals (Sweden)

    Kim De Veirman

    2014-06-01

    Full Text Available Cancer associated fibroblasts (CAFs comprise a heterogeneous population that resides within the tumor microenvironment. They actively participate in tumor growth and metastasis by production of cytokines and chemokines, and the release of pro-inflammatory and pro-angiogenic factors, creating a more supportive microenvironment. The aim of the current review is to summarize the origin and characteristics of CAFs, and to describe the role of CAFs in tumor progression and metastasis. Furthermore, we focus on the presence of CAFs in hypoxic conditions in relation to multiple myeloma disease.

  14. Secretome Analysis of Human Primary Fibroblasts Undergoing Senescence

    DEFF Research Database (Denmark)

    Rogowska-Wrzesinska, Adelina; Micutkova, Lucia; Diener, Thomas

    complementary approaches: 1)  Two-dimensional gel electro-phoresis (2DGE) was used to separate proteins and quantitate their expression; selected proteins were identified using mass spectrometry; 2)  Proteins were digested into peptides and analysed using high performance liquid chromatography (HPLC) and mass...... forms of the same protein. Using the gel free approach we were able to identify over 400 proteins and 29 of them showed significant differences in secretion between young and old fibroblasts. Among them MMP-1 was found up regulated and several ECM proteins were found down regulated: lamin A, collagen...

  15. Oxidized DNA induces an adaptive response in human fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Kostyuk, Svetlana V., E-mail: svet.kostyuk@gmail.com [Research Centre for Medical Genetics, Russian Academy of Medical Sciences, Moscow (Russian Federation); Tabakov, Viacheslav J.; Chestkov, Valerij V.; Konkova, Marina S.; Glebova, Kristina V.; Baydakova, Galina V.; Ershova, Elizaveta S.; Izhevskaya, Vera L. [Research Centre for Medical Genetics, Russian Academy of Medical Sciences, Moscow (Russian Federation); Baranova, Ancha, E-mail: abaranov@gmu.edu [Research Centre for Medical Genetics, Russian Academy of Medical Sciences, Moscow (Russian Federation); Center for the Study of Chronic Metabolic Diseases, School of System Biology, George Mason University, Fairfax, VA 22030 (United States); Veiko, Natalia N. [Research Centre for Medical Genetics, Russian Academy of Medical Sciences, Moscow (Russian Federation)

    2013-07-15

    Highlights: • We describe the effects of gDNAOX on human fibroblasts cultivated in serum withdrawal conditions. • gDNAOX evokes an adaptive response in human fibroblasts. • gDNAOX increases the survival rates in serum starving cell populations. • gDNAOX enhances the survival rates in cell populations irradiated at 1.2 Gy dose. • gDNAOX up-regulates NRF2 and inhibits NF-kappaB-signaling. - Abstract: Cell-free DNA (cfDNA) released from dying cells contains a substantial proportion of oxidized nucleotides, thus, forming cfDNA{sup OX}. The levels of cfDNA{sup OX} are increased in the serum of patients with chronic diseases. Oxidation of DNA turns it into a stress signal. The samples of genomic DNA (gDNA) oxidized by H{sub 2}O{sub 2}in vitro (gDNA{sup OX}) induce effects similar to that of DNA released from damaged cells. Here we describe the effects of gDNA{sup OX} on human fibroblasts cultivated in the stressful conditions of serum withdrawal. In these cells, gDNA{sup OX} evokes an adaptive response that leads to an increase in the rates of survival in serum starving cell populations as well as in populations irradiated at the dose of 1.2 Gy. These effects are not seen in control populations of fibroblasts treated with non-modified gDNA. In particular, the exposure to gDNA{sup OX} leads to a decrease in the expression of the proliferation marker Ki-67 and an increase in levels of PSNA, a decrease in the proportion of subG1- and G2/M cells, a decrease in proportion of cells with double strand breaks (DSBs). Both gDNA{sup OX} and gDNA suppress the expression of DNA sensors TLR9 and AIM2 and up-regulate nuclear factor-erythroid 2 p45-related factor 2 (NRF2), while only gDNA{sup OX} inhibits NF-κB signaling. gDNA{sup OX} is a model for oxidized cfDNA{sup OX} that is released from the dying tumor cells and being carried to the distant organs. The systemic effects of oxidized DNA have to be taken into account when treating tumors. In particular, the damaged DNA

  16. Fibroblastic rheumatism: case report and review of the literature.

    Science.gov (United States)

    Lee, Joyce M; Sundel, Robert P; Liang, Marilyn G

    2002-01-01

    A previously healthy 7-year-old boy presented with polyarthritis and skin lesions. Multiple, skin- to pink-colored, firm papules were noted on the periungual areas, hands, feet, and nose. There was swelling of the proximal interphalangeal joints, wrists, elbows, ankles, and knees. A skin biopsy specimen revealed dermal fibrosis and interspersed histiocytes and lymphocytes. These findings were consistent with fibroblastic rheumatism, a condition characterized by cutaneous nodules and a symmetric polyarthritis. He was treated with methotrexate and corticosteroids with improvement in the symptoms of his arthritis and skin lesions. This early treatment was beneficial in our patient.

  17. Secretome Analysis of Human Primary Fibroblasts Undergoing Senescence

    DEFF Research Database (Denmark)

    Rogowska-Wrzesinska, Adelina; Micutkova, Lucia; Diener, Thomas

    metalloproteinases (MMP1, 2 and 3), secreted protein acidic and rich in cysteine (SPARC) and two isoforms of metalloproteinase inhibitors (TIMP1 and 2). Major proteins decreased in senescent secretome were follistatin-like 1 and nucleobindin-1. Over 800 spots were separated by 2DGE of which many represent modified...... alpha-1(XII) chain and fibulin 3. Results obtained until now are in agreement with the suggested shift from matix - synthesizing to matrix - degrading phenotype in senescent fibroblasts except the increased secretion of metalloproteinase inhibitors. Work is going on to identify the remaining...

  18. Orbital fibroblast chemokine modulation: effects of dexamethasone and cyclosporin A

    OpenAIRE

    BURNSTINE, M.; Elner, S.; Elner, V.

    1998-01-01

    AIM—Orbital inflammation is common, but the mechanisms underlying leucocytic infiltration of orbital tissue are poorly understood. Human orbital fibroblasts (OF) express chemokines, interleukin 8 (IL-8) and monocyte chemotactic protein 1 (MCP-1), when exposed to proinflammatory cytokines. The effects of dexamethasone (DEX) and cyclosporin A (CSA) on OF IL-8 and MCP-1 were examined.
METHODS—Cultured human OF were incubated with recombinant interleukin 1β (rIL-1β; 0.2, 2.0, 20 ng/ml) alone or i...

  19. Influence of glycosaminoglycan identity on vocal fold fibroblast behavior.

    Science.gov (United States)

    Jimenez-Vergara, Andrea Carolina; Munoz-Pinto, Dany J; Becerra-Bayona, Silvia; Wang, Bo; Iacob, Alexandra; Hahn, Mariah S

    2011-11-01

    Poly(ethylene glycol) (PEG) hydrogels have recently begun to be studied for the treatment of scarred vocal fold lamina propria due, in part, to their tunable mechanical properties, resistance to fibroblast-mediated contraction, and ability to be polymerized in situ. However, pure PEG gels lack intrinsic biochemical signals to guide cell behavior and generally fail to mimic the frequency-dependent viscoelastic response critical to normal superficial lamina propria function. Recent results suggest that incorporation of viscoelastic bioactive substances, such as glycosaminoglycans (GAGs), into PEG networks may allow these gels to more closely approach the mechanical responses of normal vocal fold lamina propria while also stimulating desired vocal fold fibroblast behaviors. Although a number of vocal fold studies have examined the influence of hyaluronan (HA) on implant mechanics and vocal fold fibroblast responses, the effects of other GAG types have been relatively unexplored. This is significant, since recent studies have suggested that chondroitin sulfate C (CSC) and heparan sulfate (HS) are substantially altered in scarred lamina propria. The present study was therefore designed to evaluate the effects of CSC and HS incorporation on the mechanical response of PEG gels and vocal fold fibroblast behavior relative to HA. As with PEG-HA, the viscoelasticity of PEG-CSC and PEG-HS gels more closely approached that of the normal vocal fold lamina propria than pure PEG hydrogels. In addition, collagen I deposition and fibronectin production were significantly higher in CSC than in HA gels, and levels of the myofibroblast marker smooth muscle α-actin (SM α-actin) were greater in CSC and HS gels than in HA gels. Since collagen I, fibronectin, and SM α-actin are generally elevated in scarred lamina propria these results suggest that CSC and HS may be undesirable for vocal fold implants relative to HA. Investigation of various signaling intermediates indicated that

  20. Effect of microemulsions on cell viability of human dermal fibroblasts

    Science.gov (United States)

    Li, Juyi; Mironava, Tatsiana; Simon, Marcia; Rafailovich, Miriam; Garti, Nissim

    Microemulsions are optically clear, thermostable and isotropic mixture consisting of water, oil and surfactants. Their advantages of ease preparation, spontaneous formation, long-term stability and enhanced solubility of bioactive materials make them great potentials as vehicles in food and pharmaceutical applications. In this study, comparative in vitro cytotoxicity tests were performed to select a best formulation of microemulsion with the least toxicity for human dermal fibroblasts. Three different kinds of oils and six different kinds of surfactants were used to form microemulsions by different ratios. The effect of oil type and surfactant type as well as their proportions on cell proliferation and viability were tested.

  1. EPAC expression and function in cardiac fibroblasts and myofibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Olmedo, Ivonne; Muñoz, Claudia; Guzmán, Nancy; Catalán, Mabel; Vivar, Raúl; Ayala, Pedro; Humeres, Claudio; Aránguiz, Pablo [Departamento de Química Farmacológica y Toxicológica, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile); García, Lorena [Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile); Velarde, Victoria [Departamento de Ciencias Fisiológicas, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile (Chile); Díaz-Araya, Guillermo, E-mail: gadiaz@ciq.uchile.cl [Departamento de Química Farmacológica y Toxicológica, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile)

    2013-10-15

    In the heart, cardiac fibroblasts (CF) and cardiac myofibroblasts (CMF) are the main cells responsible for wound healing after cardiac insult. Exchange protein activated by cAMP (EPAC) is a downstream effector of cAMP, and it has been not completely studied on CF. Moreover, in CMF, which are the main cells responsible for cardiac healing, EPAC expression and function are unknown. We evaluated in both CF and CMF the effect of transforming growth factor β1 (TGF-β1) on EPAC-1 expression. We also studied the EPAC involvement on collagen synthesis, adhesion, migration and collagen gel contraction. Method: Rat neonatal CF and CMF were treated with TGF-β1 at different times and concentrations. EPAC-1 protein levels and Rap1 activation were measured by western blot and pull down assay respectively. EPAC cellular functions were determined by adhesion, migration and collagen gel contraction assay; and collagen expression was determined by western blot. Results: TGF-β1 through Smad and JNK significantly reduced EPAC-1 expression in CF, while in CMF this cytokine increased EPAC-1 expression through ERK1/2, JNK, p38, AKT and Smad3. EPAC activation was able to induce higher Rap1-GTP levels in CMF than in CF. EPAC and PKA, both cAMP effectors, promoted CF and CMF adhesion on fibronectin, as well as CF migration; however, this effect was not observed in CMF. EPAC but not PKA activation mediated collagen gel contraction in CF, while in CMF both PKA and EPAC mediated collagen gel contraction. Finally, the EPAC and PKA activation reduced collagen synthesis in CF and CMF. Conclusion: TGF-β1 differentially regulates the expression of EPAC in CF and CMF; and EPAC regulates differentially CF and CMF functions associated with cardiac remodeling. - Highlights: • TGF-β1 regulates EPAC-1 expression in cardiac fibroblast and myofibroblast. • Rap-1GTP levels are higher in cardiac myofibroblast than fibroblast. • EPAC-1 controls adhesion, migration and collagen synthesis in cardiac

  2. Myb proteins inhibit fibroblast transformation by v-Rel

    Directory of Open Access Journals (Sweden)

    Lipsick Joseph S

    2006-11-01

    Full Text Available Abstract Genes that cause cancer have been divided into two general classes – oncogenes that act in a dominant fashion to transform normal cells into a malignant state, and tumor suppressor genes that act in a dominant fashion to prevent such transformation. In this report, we demonstrate that both the v-myb retroviral oncogene, which causes leukemic transformation of hematopoietic cells, and the c-myb proto-oncogene can also function as inhibitors of fibroblast transformation by the v-rel oncogene. These results imply that the myb genes can function either as oncogenes or as tumor suppressors in different cellular contexts.

  3. Co-cultivation of pancreatic cancer cells with orthotopic tumor-derived fibroblasts: fibroblasts stimulate tumor cell invasion via HGF secretion whereas cancer cells exert a minor regulative effect on fibroblasts HGF production.

    Science.gov (United States)

    Qian, Li-Wu; Mizumoto, Kazuhiro; Maehara, Naoki; Ohuchida, Kenoki; Inadome, Naoki; Saimura, Michiyo; Nagai, Eishi; Matsumoto, Kunio; Nakamura, Toshikazu; Tanaka, Masao

    2003-02-10

    The intensive stromal reaction is one of characteristics of pancreatic exocrine carcinoma. The mutual interaction between pancreatic cancer cells and orthotopic tumor-derived fibroblasts have not been clarified yet. In this study, we sought to elucidate the mechanism underlying the tumor-stromal interaction with an in vitro coculture experimental system. Considerable strong c-Met expression was detected in seven out ten lines of human pancreatic carcinoma cells, as determined by Western blotting. For hepatocyte growth factor (HGF)-production, however, none or only trace amounts of HGF could be detected in those ten cell lines. Of the two lots of tumor-derived fibroblasts obtained from two pancreatic cancer patients, the fibroblasts capable to produce HGF could initiate an apparent invasion-stimulating response in strong c-Met-expressed Suit-2 and Panc-1 cells but not in faint expressed Mia PaCa-2 and BxPC-3 cells. A specialized HGF antagonist, NK4 would effectively inhibit the fibroblast-mediated invasive growth, thus proving the key role of the paracrine-fashioned HGF/c-Met pathway in the tumor-stromal interaction. On the other hand, the regulative action of cancer cells on HGF expression of fibroblasts was also investigated using direct or indirect coculture systems. For the fibroblasts that originally did not produce HGF, cancer cells failed to show any HGF-inductive effect. For the HGF-producing fibroblasts, despite of somewhat upregulation or downregulation in fibroblast HGF expression, the feedback regulation by studied pancreatic cancer cells in both coculture modes were relatively limited. This in vitro study sketched out the interaction between cancerous and stromal compartments with an emphasis on HGF/c-Met signal pathway, thus possibly helping to unveil the more complicated mutual modulation in vivo between pancreatic cancer and host mesenchymal tissues.

  4. Current perspectives on the role of orbital fibroblasts in the pathogenesis of Graves' ophthalmopathy.

    Science.gov (United States)

    Dik, Willem A; Virakul, Sita; van Steensel, Leendert

    2016-01-01

    Graves' ophthalmopathy (GO) is an extra-thyroidal complication of Graves' disease (GD; Graves' hyperthyroidism) characterized by orbital tissue inflammation, expansion, remodeling and fibrosis. Although the initiating trigger of GO is still indistinct, excessive orbital fibroblast activity is at the heart of its pathogenesis. Orbital fibroblasts are activated by cellular interactions with immune cells and the soluble factors they secrete. Orbital fibroblasts, especially from GO patients, express the thyrotropin receptor (TSH-receptor; TSHR), and activation of the orbital fibroblast population by stimulatory autoantibodies directed against the TSHR may provide an important link between GD and GO. Furthermore, stimulatory autoantibodies directed against the insulin-like growth factor-1 receptor have been proposed to contribute to orbital fibroblast activation in GO. Activated orbital fibroblasts produce inflammatory mediators thereby contributing to the orbital inflammatory process in GO. Moreover, orbital fibroblasts exhibit robust proliferative activity and extracellular matrix (especially hyaluronan) synthesizing capacity and can differentiate into adipocytes and myofibroblasts with disease progression, thereby contributing to tissue expansion/remodeling and fibrosis in GO. Orbital fibroblasts, especially those from GO patients, exhibit a hyper-responsive phenotype when compared to fibroblasts from other anatomical regions, which may further contribute to GO pathogenesis. Fibrocytes have been identified as additional source of orbital fibroblasts in GO, where they may contribute to orbital tissue inflammation, adipogenesis and remodeling/fibrosis. This review addresses our current view on the role that orbital fibroblasts fulfill in GO pathogenesis and both established as well as less established not fully crystallized concepts that need future studies will be discussed.

  5. Potent, selective inhibitors of fibroblast growth factor receptor define fibroblast growth factor dependence in preclinical cancer models.

    Science.gov (United States)

    Squires, Matthew; Ward, George; Saxty, Gordan; Berdini, Valerio; Cleasby, Anne; King, Peter; Angibaud, Patrick; Perera, Tim; Fazal, Lynsey; Ross, Douglas; Jones, Charlotte Griffiths; Madin, Andrew; Benning, Rajdeep K; Vickerstaffe, Emma; O'Brien, Alistair; Frederickson, Martyn; Reader, Michael; Hamlett, Christopher; Batey, Michael A; Rich, Sharna; Carr, Maria; Miller, Darcey; Feltell, Ruth; Thiru, Abarna; Bethell, Susanne; Devine, Lindsay A; Graham, Brent L; Pike, Andrew; Cosme, Jose; Lewis, Edward J; Freyne, Eddy; Lyons, John; Irving, Julie; Murray, Christopher; Newell, David R; Thompson, Neil T

    2011-09-01

    We describe here the identification and characterization of 2 novel inhibitors of the fibroblast growth factor receptor (FGFR) family of receptor tyrosine kinases. The compounds exhibit selective inhibition of FGFR over the closely related VEGFR2 receptor in cell lines and in vivo. The pharmacologic profile of these inhibitors was defined using a panel of human tumor cell lines characterized for specific mutations, amplifications, or translocations known to activate one of the four FGFR receptor isoforms. This pharmacology defines a profile for inhibitors that are likely to be of use in clinical settings in disease types where FGFR is shown to play an important role.

  6. Expression of herpes simplex virus 1 microRNAs in cell culture models of quiescent and latent infection.

    Science.gov (United States)

    Jurak, Igor; Hackenberg, Michael; Kim, Ju Youn; Pesola, Jean M; Everett, Roger D; Preston, Chris M; Wilson, Angus C; Coen, Donald M

    2014-02-01

    To facilitate studies of herpes simplex virus 1 latency, cell culture models of quiescent or latent infection have been developed. Using deep sequencing, we analyzed the expression of viral microRNAs (miRNAs) in two models employing human fibroblasts and one using rat neurons. In all cases, the expression patterns differed from that in productively infected cells, with the rat neuron pattern most closely resembling that found in latently infected human or mouse ganglia in vivo.

  7. A Marfan syndrome gene expression phenotype in cultured skin fibroblasts

    Directory of Open Access Journals (Sweden)

    Emond Mary

    2007-09-01

    Full Text Available Abstract Background Marfan syndrome (MFS is a heritable connective tissue disorder caused by mutations in the fibrillin-1 gene. This syndrome constitutes a significant identifiable subtype of aortic aneurysmal disease, accounting for over 5% of ascending and thoracic aortic aneurysms. Results We used spotted membrane DNA macroarrays to identify genes whose altered expression levels may contribute to the phenotype of the disease. Our analysis of 4132 genes identified a subset with significant expression differences between skin fibroblast cultures from unaffected controls versus cultures from affected individuals with known fibrillin-1 mutations. Subsequently, 10 genes were chosen for validation by quantitative RT-PCR. Conclusion Differential expression of many of the validated genes was associated with MFS samples when an additional group of unaffected and MFS affected subjects were analyzed (p-value -6 under the null hypothesis that expression levels in cultured fibroblasts are unaffected by MFS status. An unexpected observation was the range of individual gene expression. In unaffected control subjects, expression ranges exceeding 10 fold were seen in many of the genes selected for qRT-PCR validation. The variation in expression in the MFS affected subjects was even greater.

  8. A Marfan syndrome gene expression phenotype in cultured skin fibroblasts

    Science.gov (United States)

    Yao, Zizhen; Jaeger, Jochen C; Ruzzo, Walter L; Morale, Cecile Z; Emond, Mary; Francke, Uta; Milewicz, Dianna M; Schwartz, Stephen M; Mulvihill, Eileen R

    2007-01-01

    Background Marfan syndrome (MFS) is a heritable connective tissue disorder caused by mutations in the fibrillin-1 gene. This syndrome constitutes a significant identifiable subtype of aortic aneurysmal disease, accounting for over 5% of ascending and thoracic aortic aneurysms. Results We used spotted membrane DNA macroarrays to identify genes whose altered expression levels may contribute to the phenotype of the disease. Our analysis of 4132 genes identified a subset with significant expression differences between skin fibroblast cultures from unaffected controls versus cultures from affected individuals with known fibrillin-1 mutations. Subsequently, 10 genes were chosen for validation by quantitative RT-PCR. Conclusion Differential expression of many of the validated genes was associated with MFS samples when an additional group of unaffected and MFS affected subjects were analyzed (p-value < 3 × 10-6 under the null hypothesis that expression levels in cultured fibroblasts are unaffected by MFS status). An unexpected observation was the range of individual gene expression. In unaffected control subjects, expression ranges exceeding 10 fold were seen in many of the genes selected for qRT-PCR validation. The variation in expression in the MFS affected subjects was even greater. PMID:17850668

  9. From mechanotransduction to extracellular matrix gene expression in fibroblasts.

    Science.gov (United States)

    Chiquet, Matthias; Gelman, Laurent; Lutz, Roman; Maier, Silke

    2009-05-01

    Tissue mechanics provide an important context for tissue growth, maintenance and function. On the level of organs, external mechanical forces largely influence the control of tissue homeostasis by endo- and paracrine factors. On the cellular level, it is well known that most normal cell types depend on physical interactions with their extracellular matrix in order to respond efficiently to growth factors. Fibroblasts and other adherent cells sense changes in physical parameters in their extracellular matrix environment, transduce mechanical into chemical information, and integrate these signals with growth factor derived stimuli to achieve specific changes in gene expression. For connective tissue cells, production of the extracellular matrix is a prominent response to changes in mechanical load. We will review the evidence that integrin-containing cell-matrix adhesion contacts are essential for force transmission from the extracellular matrix to the cytoskeleton, and describe novel experiments indicating that mechanotransduction in fibroblasts depends on focal adhesion adaptor proteins that might function as molecular springs. We will stress the importance of the contractile actin cytoskeleton in balancing external with internal forces, and describe new results linking force-controlled actin dynamics directly to the expression of specific genes, among them the extracellular matrix protein tenascin-C. As assembly lines for diverse signaling pathways, matrix adhesion contacts are now recognized as the major sites of crosstalk between mechanical and chemical stimuli, with important consequences for cell growth and differentiation.

  10. Fibroblasts secrete Slit2 to inhibit fibrocyte differentiation and fibrosis.

    Science.gov (United States)

    Pilling, Darrell; Zheng, Zhichao; Vakil, Varsha; Gomer, Richard H

    2014-12-23

    Monocytes leave the blood and enter tissues. In healing wounds and fibrotic lesions, some of the monocytes differentiate into fibroblast-like cells called fibrocytes. In healthy tissues, even though monocytes enter the tissue, for unknown reasons, very few monocytes differentiate into fibrocytes. In this report, we show that fibroblasts from healthy human tissues secrete the neuronal guidance protein Slit2 and that Slit2 inhibits human fibrocyte differentiation. In mice, injections of Slit2 inhibit bleomycin-induced lung fibrosis. In lung tissue from pulmonary fibrosis patients with relatively normal lung function, Slit2 has a widespread distribution whereas, in patients with advanced disease, there is less Slit2 in the fibrotic lesions. These data may explain why fibrocytes are rarely observed in healthy tissues, may suggest that the relative levels of Slit2 present in healthy tissue and at sites of fibrosis may have a significant effect on the decision of monocytes to differentiate into fibrocytes, and may indicate that modulating Slit2 signaling may be useful as a therapeutic for fibrosis.

  11. Differentiation of Human Embryonic Stem Cells on Periodontal Ligament Fibroblasts.

    Science.gov (United States)

    Elçin, Y Murat; İnanç, Bülend; Elçin, A Eser

    2016-01-01

    Human embryonic stem cells' (hESCs) unlimited proliferative potential and differentiation capability to all somatic cell types makes them one of the potential cell sources in cell-based tissue engineering strategies as well as various experimental applications in fields such as developmental biology, pharmacokinetics, toxicology, and genetics. Periodontal tissue engineering is an approach to reconstitute the ectomesenchymally derived alveolar bone, periodontal ligament apparatus, and cementum tissues lost as a result of periodontal diseases. Cell-based therapies may offer potential advantage in overcoming the inherent limitations associated with contemporary regenerative procedures, such as dependency on defect type and size and the pool and capacity of progenitor cells resident in the wound area. Further elucidation of developmental mechanisms associated with tooth formation may also contribute to valuable knowledge based upon which the future therapies can be designed. Protocols for the differentiation of pluripotent hESCs into periodontal ligament fibroblastic cells (PDLF) as common progenitors for ligament, cementum, and alveolar bone tissue represent an initial step in developing hESC-based experimental and tissue engineering strategies. The present protocol describes methods associated with the guided differentiation of hESCs by the use of coculture with adult PDLFs and the resulting change of morphotype and phenotype of the pluripotent embryonic stem cells toward fibroblastic and osteoblastic lineages.

  12. Differential Gene Expression of Fibroblasts: Keloid versus Normal

    Directory of Open Access Journals (Sweden)

    Michael F. Angel

    2002-11-01

    Full Text Available Abstract: This study investigated gene regulation and unique gene products in both keloid (KDF and normal (NDF dermal fibroblasts in established cell lines. For gene regulation, NDF versus KDF were compared using Clontech's Atlas™ Human cDNA Expression Array while unique gene products were studied using RNA Fingerprinting Kit. RNA from each sample was converted to cDNA using oligo-dT primers. Down-regulated genes using Atlas Array in KDF were 1 60 S ribosomal protein, 2 Thioredoxin dependent peroxidase, 3 Nuclease sensitive element DNA binding protein, 4 c-myc purine-binding transcription factor, 5 c-AMP dependent protein kinase, and, 6 Heat Shock Protein 90 kDa. Genes that are up regulated in KDF were 1 Tubulin and 2 Heat Shock Protein 27 kDa. With the differential display, we found 17 bands unique to both KDF and NDF. The specific gene and the manner in which they were differentially regulated have direct implications to understanding keloid fibroblast proliferation.

  13. Proteases induce secretion of collagenase and plasminogen activator by fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Werb, Z.; Aggeler, J.

    1978-04-01

    We have observed that treatment of rabbit synovial fibroblasts with proteolytic enzymes can induce secretion of collagenase (EC 3.4.24.7) and plasminogen activator (EC 3.4.21.-). Cells treated for 2 to 24 hr with plasmin, trypsin, chymotrypsin, pancreatic elastase, papain, bromelain, thermolysin, or ..cap alpha..-protease but not with thrombin or neuraminidase secreted detectable amounts of collagenase within 16 to 48 hr. Treatment of fibroblasts with trypsin also induced secretion of plasminogen activator. Proteases initiated secretion of collagenase (up to 20 units per 10/sup 6/ cells per 24 hr) only when treatment produced decreased cell adhesion. Collagenase production did not depend on continued presence of proteolytic activity or on subsequent cell adhesion, spreading, or proliferation. Routine subculturing with crude trypsin also induced collagenase secretion by cells. Secretion of collagenase was prevented and normal spreading was obtained if the trypsinized cells were placed into medium containing fetal calf serum. Soybean trypsin inhibitor, ..cap alpha../sub 1/-antitrypsin, bovine serum albumin, collagen, and fibronectin did not inhibit collagenase production. Although proteases that induced collagenase secretion also removed surface glycoprotein, the kinetics of induction of cell protease secretion were different from those for removal of fibronectin. Physiological inducers of secretion of collagenase and plasminogen activator by cells have not been identified. These results suggest that extracellular proteases in conjunction with plasma proteins may govern protease secretion by cells.

  14. Proliferative Effects of Histamine on Primary Human Pterygium Fibroblasts

    Directory of Open Access Journals (Sweden)

    Zhenwei Qin

    2016-01-01

    Full Text Available Purpose. It has been confirmed that inflammatory cytokines are involved in the progression of pterygium. Histamine can enhance proliferation and migration of many cells. Therefore, we intend to investigate the proliferative and migratory effects of histamine on primary culture of human pterygium fibroblasts (HPFs. Methods. Pterygium and conjunctiva samples were obtained from surgery, and toluidine blue staining was used to identify mast cells. 3-[4, 5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT was performed to evaluate the proliferative rate of HPFs and human conjunctival fibroblasts (HCFs; ki67 expression was also measured by immunofluorescence analysis. Histamine receptor-1 (H1R antagonist (Diphenhydramine Hydrochloride and histamine receptor-2 (H2R antagonist (Nizatidine were added to figure out which receptor was involved. Wound healing model was used to evaluate the migratory ability of HPFs. Results. The numbers of total mast cells and degranulated mast cells were both higher in pterygium than in conjunctiva. Histamine had a proliferative effect on both HPFs and HCFs, the effective concentration (10 μmol/L on HPFs was lower than on HCFs (100 μmol/L, and the effect could be blocked by H1R antagonist. Histamine showed no migratory effect on HPFs. Conclusion. Histamine may play an important role in the proliferation of HPFs and act through H1R.

  15. Fibroblast growth factor receptors, developmental corruption and malignant disease.

    Science.gov (United States)

    Kelleher, Fergal C; O'Sullivan, Hazel; Smyth, Elizabeth; McDermott, Ray; Viterbo, Antonella

    2013-10-01

    Fibroblast growth factors (FGF) are a family of ligands that bind to four different types of cell surface receptor entitled, FGFR1, FGFR2, FGFR3 and FGFR4. These receptors differ in their ligand binding affinity and tissue distribution. The prototypical receptor structure is that of an extracellular region comprising three immunoglobulin (Ig)-like domains, a hydrophobic transmembrane segment and a split intracellular tyrosine kinase domain. Alternative gene splicing affecting the extracellular third Ig loop also creates different receptor isoforms entitled FGFRIIIb and FGFRIIIc. Somatic fibroblast growth factor receptor (FGFR) mutations are implicated in different types of cancer and germline FGFR mutations occur in developmental syndromes particularly those in which craniosynostosis is a feature. The mutations found in both conditions are often identical. Many somatic FGFR mutations in cancer are gain-of-function mutations of established preclinical oncogenic potential. Gene amplification can also occur with 19-22% of squamous cell lung cancers for example having amplification of FGFR1. Ontologic comparators can be informative such as aberrant spermatogenesis being implicated in both spermatocytic seminomas and Apert syndrome. The former arises from somatic FGFR3 mutations and Apert syndrome arises from germline FGFR2 mutations. Finally, therapeutics directed at inhibiting the FGF/FGFR interaction are a promising subject for clinical trials.

  16. Cytotoxic effects of nickel nanowires in human fibroblasts

    KAUST Repository

    Felix Servin, Laura P.

    2016-03-09

    The increasing interest in the use of magnetic nanostructures for biomedical applications necessitates rigorous studies to be carried out in order to determine their potential toxicity. This work attempts to elucidate the cytotoxic effects of nickel nanowires (NWs) in human fibroblasts WI-38 by a colorimetric assay (MTT) under two different parameters: NW concentration and exposure time. This was complemented with TEM and confocal images to assess the NWs internalization and to identify any changes in the cell morphology. Ni NWs were fabricated by electrodeposition using porous alumina templates. Energy dispersive X-Ray analysis, scanning electron microscopy and transmission electron microscopy imaging were used for NW characterization. The results showed decreased cell metabolic activity for incubation times longer than 24 hours and no negative effects for exposure times shorter than that. The cytotoxicity effects for human fibroblasts were then compared with those reported for HCT 116 cells, and the findings point out that it is relevant to consider the cellular size. In addition, the present study compares the toxic effects of equivalent amounts of nickel in the form of its salt to those of NWs and shows that the NWs are more toxic than the salts. Internalized NWs were found in vesicles inside of the cells where their presence induced inflammation of the endoplasmic reticulum.

  17. Role of fibroblast growth factors in organ regeneration and repair.

    Science.gov (United States)

    El Agha, Elie; Kosanovic, Djuro; Schermuly, Ralph T; Bellusci, Saverio

    2016-05-01

    In its broad sense, regeneration refers to the renewal of lost cells, tissues or organs as part of the normal life cycle (skin, hair, endometrium etc.) or as part of an adaptive mechanism that organisms have developed throughout evolution. For example, worms, starfish and amphibians have developed remarkable regenerative capabilities allowing them to voluntarily shed body parts, in a process called autotomy, only to replace the lost parts afterwards. The bizarre myth of the fireproof homicidal salamander that can survive fire and poison apple trees has persisted until the 20th century. Salamanders possess one of the most robust regenerative machineries in vertebrates and attempting to draw lessons from limb regeneration in these animals and extrapolate the knowledge to mammals is a never-ending endeavor. Fibroblast growth factors are potent morphogens and mitogens that are highly conserved among the animal kingdom. These growth factors play key roles in organogenesis during embryonic development as well as homeostatic balance during postnatal life. In this review, we provide a summary about the current knowledge regarding the involvement of fibroblast growth factor signaling in organ regeneration and repair. We also shed light on the use of these growth factors in previous and current clinical trials in a wide array of human diseases.

  18. Transglutaminase-2 regulation by arecoline in gingival fibroblasts.

    Science.gov (United States)

    Thangjam, G S; Agarwal, P; Khan, I; Verma, U P; Balapure, A K; Rao, S G; Kondaiah, P

    2009-02-01

    Transglutaminase-2 (TGM-2) stabilizes extracellular matrix (ECM) proteins by cross-linking and has been implicated in several fibrotic disorders. Arecoline present in betel quid has been proposed as one of the causative factors for oral submucous fibrosis (OSMF). Hence, we hypothesize that arecoline may regulate TGM-2 and may have a role in the pathogenesis of OSMF. The expression of TGM-2 was studied in OSMF tissues by real-time RT-PCR analysis, and significant overexpression was observed in most OSMF tissues (P=0.0112) compared with normal tissues. Arecoline induced TGM-2 mRNA and protein expression as well as TGM-2 activity in human gingival fibroblast cells. The addition of methocramine hemihydrate (M-2 muscarinic acetylcholine receptor selective antagonist) or 8'-bromo-cAMP abolished arecoline-mediated TGM-2 induction, suggesting a role for M-2 muscarinic acid receptor and a repressor role for cAMP. Our study provides evidence for TGM-2 overexpression in OSMF and its regulation by arecoline in oral fibroblasts.

  19. Physiological loading of tendons induces scleraxis expression in epitenon fibroblasts.

    Science.gov (United States)

    Mendias, Christopher L; Gumucio, Jonathan P; Bakhurin, Konstantin I; Lynch, Evan B; Brooks, Susan V

    2012-04-01

    Scleraxis is a basic helix-loop-helix transcription factor that plays a central role in promoting fibroblast proliferation and matrix synthesis during the embryonic development of tendons. Mice with a targeted inactivation of scleraxis (Scx(-/-)) fail to properly form limb tendons, but the role that scleraxis has in regulating the growth and adaptation of tendons of adult organisms is unknown. To determine if scleraxis expression changes in response to a physiological growth stimulus to tendons, we subjected adult mice that express green fluorescent protein (GFP) under the control of the scleraxis promoter (ScxGFP) to a 6-week-treadmill training program designed to induce adaptive growth in Achilles tendons. Age matched sedentary ScxGFP mice were used as controls. Scleraxis expression was sparsely observed in the epitenon region of sedentary mice, but in response to treadmill training, scleraxis was robustly expressed in fibroblasts that appeared to be emerging from the epitenon and migrating into the superficial regions of tendon fascicles. Treadmill training also led to an increase in scleraxis, tenomodulin, and type I collagen gene expression as measured by qPCR. These results suggest that in addition to regulating the embryonic formation of limb tendons, scleraxis also appears to play an important role in the adaptation of adult tendons to physiological loading.

  20. Capturing the Regenerative Potential of Periodontal Ligament Fibroblasts

    Directory of Open Access Journals (Sweden)

    Christina Springstead Scanlon

    2011-01-01

    Full Text Available The cell population within the periodontal ligament (PDL tissue is remarkably heterogeneous1. Fibroblasts, a mixed population of cells, are the main cellular component of the PDL and the cell type most often studied for periodontal regeneration. Osteoblasts and osteoclasts are found on the bone side, while fibroblasts, macrophages, undifferentiated adult/mesenchymal stem cells, neural elements, and endothelial cells are found throughout the PDL. Epithelial rests of Malassez cells and cementoblasts are focused near the root surface. PDL tissue also includes loose connective tissue between dense fiber bundles that contain branches of the periodontal blood vessels and nerves2. The complexity of the PDL tissue, with its various cell types and cell progenitor components, explains the challenges involved in therapies to restore tissue following periodontal disease. Cementoblasts, osteoblasts, and endothelial cells must migrate, differentiate, and coordinately interact with a variety of soluble mediators to regenerate the periodontium3. Stem cells located in the PDL tissue are key contributors to this process4. Stem cells in the PDL are important not only for formation and maintenance of the tissue but also for repair, remodeling, and regeneration of adjacent alveolar bone and cementum5. Our laboratory has shown that progenitor cells isolated from PDL tissue by selection with cell surface markers STRO-1+ and CD146+ are capable of differentiating into chondrogenic, osteogenic, and adipogenic phenotypes under appropriate culture conditions6.

  1. An Expandable, Inducible Hemangioblast State Regulated by Fibroblast Growth Factor

    Directory of Open Access Journals (Sweden)

    David T. Vereide

    2014-12-01

    Full Text Available During development, the hematopoietic and vascular lineages are thought to descend from common mesodermal progenitors called hemangioblasts. Here we identify six transcription factors, Gata2, Lmo2, Mycn, Pitx2, Sox17, and Tal1, that “trap” murine cells in a proliferative state and endow them with a hemangioblast potential. These “expandable” hemangioblasts (eHBs are capable, once released from the control of the ectopic factors, to give rise to functional endothelial cells, multilineage hematopoietic cells, and smooth muscle cells. The eHBs can be derived from embryonic stem cells, from fetal liver cells, or poorly from fibroblasts. The eHBs reveal a central role for fibroblast growth factor, which not only promotes their expansion, but also facilitates their ability to give rise to endothelial cells and leukocytes, but not erythrocytes. This study serves as a demonstration that ephemeral progenitor states can be harnessed in vitro, enabling the creation of tractable progenitor cell lines.

  2. Effect of Arctium lappa (burdock) extract on canine dermal fibroblasts.

    Science.gov (United States)

    Pomari, Elena; Stefanon, Bruno; Colitti, Monica

    2013-12-15

    Although the biological activities of Arctium lappa (burdock) have been already investigated in human and other species, data evaluating the molecular mechanisms have not been reported in the dog. In this study we analyzed for the first time the effect of a root extract of burdock on molecular responses in canine dermal fibroblasts with H2O2 stimulation (H group), with burdock treatment (B group) and with H2O2 stimulation and burdock treatment (BH group), using RNAseq technology. Differentially expressed genes (P<0.05) of H, B and BH groups in comparison to the untreated sample (negative control, C group) were identified with MeV software and were functional annotated and monitored for signaling pathways and candidate biomarkers using the Ingenuity Pathways Analysis (IPA). The expression profile of canine dermal fibroblasts treated with burdock extract with or without H2O2 stimulation, showed an up-regulation of mitochondrial superoxide dismutase (SOD2), disheveled 3 (DVL3) and chondroitin sulfate N-acetylgalactosaminyltransferase 2 (CSGALNACT2). The data suggested that burdock has implications in cell adhesion and gene expression with the modulation of Wnt/β catenin signaling and Chondroitin Sulphate Biosynthesis that are particularly important for the wound healing process.

  3. Carcinoma-associated fibroblasts provide operational flexibility in metastasis.

    Science.gov (United States)

    De Wever, Olivier; Van Bockstal, Mieke; Mareel, Marc; Hendrix, An; Bracke, Marc

    2014-04-01

    Malignant cancer cells do not act as lone wolves to achieve metastasis, as they exist within a complex ecosystem consisting of an extracellular matrix scaffold populated by carcinoma-associated fibroblasts (CAFs), endothelial cells and immune cells. We recognize local (primary tumor) and distant ecosystems (metastasis). CAFs, also termed myofibroblasts, may have other functions in the primary tumor versus the metastasis. Cellular origin and tumor heterogeneity lead to the expression of specific markers. The molecular characteristics of a CAF remain in evolution since CAFs show operational flexibility. CAFs respond dynamically to a cancer cell's fluctuating demands by shifting profitable signals necessary in metastasis. Local, tissue-resident fibroblasts and mesenchymal stem cells (MSCs) coming from reservoir sites such as bone marrow and adipose tissue are the main progenitor cells of CAFs. CAFs may induce awakening from metastatic dormancy, a major cause of cancer-specific death. Cancer management protocols influence CAF precursor recruitment and CAF activation. Since CAF signatures represent early changes in metastasis, including formation of pre-metastatic niches, we discuss whether liquid biopsies, including exosomes, may detect and monitor CAF reactions allowing optimized prognosis of cancer patients.

  4. The ORF 113 of Heliocoverpa armigera Single Nucleopolyhedrovirus Encodes a Functional Fibroblast Growth Factor

    Institute of Scientific and Technical Information of China (English)

    Xiang LI; Chang-yong LIANG; Jian-hua SONG; Xin-wen CHEN

    2008-01-01

    Fibroblast growth factor (FGF) is a key regulator of developmental processes. A FGF homolog (vFGF) is found in all lepidopteran baculoviruses. Autographa californica nucleopolyhedrovirus (AcMNPV) and Bombyx mori NPV (BmNPV) vFGFs are chemotactic factors. Here we analyzed the vfgf of Helicoverpa armigera NPV (HearNPV), a group Ⅱ NPV. The HearNPV vfgftranscripts were detected from 18 to 96 h post-infection (hpi) of Hz-AMI cells with HearNPV and encoded a 36 kDa protein, which was secreted into the culture medium. HearNPV vFGF had strong affinity to heparin, a property important for FGF signaling via an FGF receptor. Unlike its AcMNPV homolog, HearNPV vFGF specially chemoattracted Hz-AM 1, but not other insect cells such as Sf9 and Se-UCR and not the mammalian cells 293 and HepG2. HearNPV vFGF is also associated with the envelope of BV but is absent in occlusion-derived virus, which coordinated to the chemotatic activity analysis.

  5. Protozoan Infections

    Science.gov (United States)

    1989-01-01

    cooked meat of infected animals, especially sheep and pigs. Toxoplasma infects one-third to one-half of normal humans by age 40, but causes...Toxoplasma in the first trimester, 30-50 per cent deliver an obviously affected infant. Congenital infection is associated with hydrocephalus...chorioretinitis, hepatosplenomegaly, and rash. In survivors, mental retardation, deaf- ness, and cardiac malformations may be noted. Chorioretinitis may also

  6. [Hand infections].

    Science.gov (United States)

    Schiele, Philippe; Le Nen, Dominique

    2013-11-01

    Superficial and deep hand infections are frequent in general medical practice. Clinical examination is a crucial step for an adapted provided care. Most of the time, surgery is the only way to heal infections. However, in some cases (like bites), empiric antibiotherapy is first indicated to limit infection. Staphyloccocus aureus as well as Group Beta Streptococcus are the most frequently pathogenes associated with hand infections. Methicillin resistant S. Aureus must always be considered in the diagnoses. Whatever treatment is provided, clinical assessement must be repeated within two days. An early adaquated treatment prevent functional complications and in some cases death of the patients.

  7. Rhinovirus Infections

    Science.gov (United States)

    ... Genitals and Urinary Tract Glands & Growth Head Neck & Nervous System Heart Infections Learning Disabilities Obesity Orthopedic Prevention Sexually Transmitted Skin Tobacco Treatments Injuries & ...

  8. Eye Infections

    Science.gov (United States)

    ... Genitals and Urinary Tract Glands & Growth Head Neck & Nervous System Heart Infections Learning Disabilities Obesity Orthopedic Prevention Sexually Transmitted Skin Tobacco Treatments Injuries & ...

  9. Campylobacter Infections

    Science.gov (United States)

    ... Genitals and Urinary Tract Glands & Growth Head Neck & Nervous System Heart Infections Learning Disabilities Obesity Orthopedic Prevention Sexually Transmitted Skin Tobacco Treatments Injuries & ...

  10. Interferon gamma blocks the growth of Toxoplasma gondii in human fibroblasts by inducing the host cells to degrade tryptophan.

    Science.gov (United States)

    Pfefferkorn, E R

    1984-01-01

    Treatment of human fibroblasts with human recombinant gamma interferon blocked the growth of Toxoplasma gondii, an obligate intracellular protozoan parasite. Growth of the parasite was measured by a plaque assay 7 days after infection or by the incorporation of [3H]uracil 1 or 2 days after infection. The antitoxoplasma activity induced in the host cells by gamma interferon was strongly dependent upon the tryptophan concentration of the medium. Progressively higher minimal inhibitory concentrations of gamma interferon were observed as the tryptophan concentration in the culture medium was increased. Treatment with gamma interferon did not make the cells impermeable to tryptophan. The kinetics of [3H]tryptophan uptake into the acid-soluble pools of control and gamma interferon-treated cultures were identical during the first 48 sec. Thereafter uptake of [3H]tryptophan into the acid-soluble pool of control fibroblasts reached the expected plateau after 96 sec. In contrast, uptake of [3H]tryptophan continued for at least 12 min in the gamma interferon-treated cultures. At that time, the acid-soluble pool of the gamma interferon-treated cultures contained 8 times the radioactivity of the control cultures. This continued accumulation was the result of rapid intracellular degradation of [3H]tryptophan into kynurenine and N-formylkynurenine that leaked slowly from the cells. These two metabolites were also recovered from the medium of cultures treated for 1 or 2 days with gamma interferon. Human recombinant alpha and beta interferons, which have no antitoxoplasma activity, did not induce any detectable degradation of tryptophan. Several hypotheses are presented to explain how the intracellular degradation of tryptophan induced by gamma interferon could restrict the growth of an obligate intracellular parasite. Images PMID:6422465

  11. In vitro effects of pirfenidone on cardiac fibroblasts: proliferation, myofibroblast differentiation, migration and cytokine secretion.

    Directory of Open Access Journals (Sweden)

    Qiang Shi

    Full Text Available Cardiac fibroblasts (CFs are the primary cell type responsible for cardiac fibrosis during pathological myocardial remodeling. Several studies have illustrated that pirfenidone (5-methyl-1-phenyl-2-[1H]-pyridone attenuates cardiac fibrosis in different animal models. However, the effects of pirfenidone on cardiac fibroblast behavior have not been examined. In this study, we investigated whether pirfenidone directly modulates cardiac fibroblast behavior that is important in myocardial remodeling such as proliferation, myofibroblast differentiation, migration and cytokine secretion. Fibroblasts were isolated from neonatal rat hearts and bioassays were performed to determine the effects of pirfenidone on fibroblast function. We demonstrated that treatment of CFs with pirfenidone resulted in decreased proliferation, and attenuated fibroblast α-smooth muscle actin expression and collagen contractility. Boyden chamber assay illustrated that pirfenidone inhibited fibroblast migration ability, probably by decreasing the ratio of matrix metalloproteinase-9 to tissue inhibitor of metalloproteinase-1. Furthermore, pirfenidone attenuated the synthesis and secretion of transforming growth factor-β1 but elevated that of interleukin-10. These direct and pleiotropic effects of pirfenidone on cardiac fibroblasts point to its potential use in the treatment of adverse myocardial remodeling.

  12. High inorganic phosphate causes DNMT1 phosphorylation and subsequent fibrotic fibroblast activation

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Xiaoying [Department of Nephrology and Rheumatology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); Department of Cardiology and Pneumology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); Xu, Xingbo [Department of Cardiology and Pneumology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); Zeisberg, Elisabeth M. [Department of Cardiology and Pneumology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); German Center for Cardiovascular Research (DZHK), Göttingen (Germany); Zeisberg, Michael, E-mail: mzeisberg@med.uni-goettingen.de [Department of Nephrology and Rheumatology, Göttingen University Medical Center, Georg August University, Göttingen (Germany); German Center for Cardiovascular Research (DZHK), Göttingen (Germany)

    2016-04-08

    Phosphate is an essential constituent of critical cellular functions including energy metabolism, nucleic acid synthesis and phosphorylation-dependent cell signaling. Increased plasma phosphate levels are an independent risk factor for lowered life-expectancy as well as for heart and kidney failure. Nevertheless, direct cellular effects of elevated phosphate concentrations within the microenvironment are poorly understood and have been largely neglected in favor of phosphor-regulatory hormones. Because interstitial fibrosis is the common determinant of chronic progressive kidney disease, and because fibroblasts are major mediators of fibrogenesis, we here explored the effect of high extracellular phosphate levels on renal fibroblasts. We demonstrate that high inorganic phosphate directly induces fibrotic fibroblast activation associated with increased proliferative activity, increased expression of α-smooth muscle actin and increased synthesis of type I collagen. We further demonstrate that such fibroblast activation is dependent on phosphate influx, aberrant phosphorylation of DNA methyltransferase DNMT1 and aberrant CpG island promoter methylation. In summary, our studies demonstrate that elevated phosphate concentrations induce pro-fibrotic fibroblast activation independent of phospho-regulatory hormones. - Highlights: • We exposed human kidney fibroblasts to media containing 1 mM or 3 mM phosphate. • Increased phosphate influx causes phosphorylation of DNA methyltransferase Dnmt1. • Phosphorylated Dnmt1 causes promoter methylation and transcriptional silencing of RASAL1. • Depletion of RASAL1 causes increased intrinsic Ras-GTP activity and fibroblast activation. • Inorganic phosphate causes fibroblast activation independent of phospho-regulatory hormones.

  13. Endoglin negatively regulates transforming growth factor beta1-induced profibrotic responses in intestinal fibroblasts.

    LENUS (Irish Health Repository)

    Burke, J P

    2012-02-01

    BACKGROUND: Fibroblasts isolated from strictures in Crohn\\'s disease (CD) exhibit reduced responsiveness to stimulation with transforming growth factor (TGF) beta1. TGF-beta1, acting through the smad pathway, is critical to fibroblast-mediated intestinal fibrosis. The membrane glycoprotein, endoglin, is a negative regulator of TGF-beta1. METHODS: Intestinal fibroblasts were cultured from seromuscular biopsies of patients undergoing intestinal resection for CD strictures or from control patients. Endoglin expression was assessed using confocal microscopy, flow cytometry and western blot. The effect of small interfering (si) RNA-mediated knockdown and plasmid-mediated overexpression of endoglin on fibroblast responsiveness to TGF-beta1 was assessed by examining smad phosphorylation, smad binding element (SBE) promoter activity, connective tissue growth factor (CTGF) expression and ability to contract collagen. RESULTS: Crohn\\'s stricture fibroblasts expressed increased constitutive cell-surface and whole-cell endoglin relative to control cells. Endoglin co-localized with filamentous actin. Fibroblasts treated with siRNA directed against endoglin exhibited enhanced TGF-beta1-mediated smad-3 phosphorylation, and collagen contraction. Cells transfected with an endoglin plasmid did not respond to TGF-beta1 by exhibiting SBE promoter activity or producing CTGF. CONCLUSION: Fibroblasts from strictures in CD express increased constitutive endoglin. Endoglin is a negative regulator of TGF-beta1 signalling in the intestinal fibroblast, modulating smad-3 phosphorylation, SBE promoter activity, CTGF production and collagen contraction.

  14. Stimulation of skin repair is dependent on fibroblast source and presence of extracellular matrix.

    NARCIS (Netherlands)

    Wang, H.; Pieper, J.S.; Schotel, R.; Blitterswijk, C.A. van; Lamme, E.N.

    2004-01-01

    In this study in vitro and in vivo functions were compared between cultured dermal equivalents produced with human fibroblasts isolated either from papillary dermis or adipose tissue of the same donors. Papillary dermal fibroblasts had a normal spindle cell shape; in contrast, adipose tissue fibrobl

  15. Stimulation of Skin Repair Is Dependent on Fibroblast Source and Presence of Extracellular Matrix

    NARCIS (Netherlands)

    Wang, Hong-Jun; Pieper, Jeroen; Schotel, Roka; Blitterswijk, van Clemens A.; Lamme, Evert N.

    2004-01-01

    In this study in vitro and in vivo functions were compared between cultured dermal equivalents produced with human fibroblasts isolated either from papillary dermis or adipose tissue of the same donors. Papillary dermal fibroblasts had a normal spindle cell shape; in contrast, adipose tissue fibrobl

  16. Curcumin induces differential expression of cytoprotective enzymes but similar apoptotic responses in fibroblasts and myofibroblasts

    NARCIS (Netherlands)

    Lundvig, D.M.S.; Pennings, S.W.C.; Brouwer, K.M.; Mtaya-Mlangwa, M.; Mugonzibwa, E.A.; Kuijpers-Jagtman, A.M.; Hoff, J.W. Von den; Wagener, F.A.D.T.G.

    2015-01-01

    Excessive extracellular matrix (ECM) deposition and tissue contraction after injury can lead to esthetic and functional problems. Fibroblasts and myofibroblasts activated by transforming growth factor (TGF)-beta1 play a key role in these processes. The persistence of (myo)fibroblasts and their exces

  17. Gingival and periodontal ligament fibroblasts differ in their inflammatory response to viable Porphyromonas gingivalis

    NARCIS (Netherlands)

    Scheres, N.; Laine, M.L.; de Vries, T.J.; Everts, V.; van Winkelhoff, A.J.

    2010-01-01

    Background and Objective: Porphyromonas gingivalis is an oral pathogen strongly associated with destruction of the tooth-supporting tissues in human periodontitis. Gingival fibroblasts (GF) and periodontal ligament fibroblasts (PDLF) are functionally different cell types in the periodontium that can

  18. Hepatocyte differentiation of human fibroblasts from cirrhotic liver in vitro and in vivo

    Institute of Scientific and Technical Information of China (English)

    Yu-Ling Sun; Sheng-Yong Yin; Lin Zhou; Hai-Yang Xie; Feng Zhang; Li-Ming Wu; Shu-Sen Zheng

    2011-01-01

    BACKGROUND: Mesenchymal stem cells (MSCs) and fibro-blasts have intimate relationships, and the phenotypic homology between fibroblasts and MSCs has been recently described. The aim of this study was to investigate the hepatic differentiating potentialofhumanfibroblastsincirrhoticliver. METHODS: The phenotypes of fibroblasts in cirrhotic liver were labeled by biological methods. After that, the differentiation potential of these fibroblasts in vitro was characterized in terms of liver-specific gene and protein expression. Finally, an animal model of hepatocyte regeneration in severe combined immunodeficient (SCID) mice was created by retrorsine injection and partial hepatectomy, and the expression of human hepatocyte proteins in SCID mouse livers was checked by immunohistochemicalanalysisafterfibroblastadministration. RESULTS: Surface immunophenotyping revealed that a minority of fibroblasts expressed markers of MSCs and hepatic epithelial cytokeratins as well as alpha-smooth muscle actin, but homogeneously expressed vimentin, desmin, prolyl 4-hydroxylase and fibronectin. These fibroblasts presented the characteristics of hepatocytes in vitro and differentiated directly into functional hepatocytes in the liver of hepatecto-mized SCID mice. CONCLUSIONS: This study demonstrated that fibroblasts in cirrhotic liver have the potential to differentiate into hepatocyte-like cells in vitro and in vivo. Our findings infer that hepatic differentiation of fibroblasts may serve as a new target for reversion of liver fibrosis and a cell source for tissue engineering.

  19. Versican V1 Overexpression Induces a Myofibroblast-Like Phenotype in Cultured Fibroblasts.

    Directory of Open Access Journals (Sweden)

    Jon M Carthy

    Full Text Available Versican, a chondroitin sulphate proteoglycan, is one of the key components of the provisional extracellular matrix expressed after injury. The current study evaluated the hypothesis that a versican-rich matrix alters the phenotype of cultured fibroblasts.The full-length cDNA for the V1 isoform of human versican was cloned and the recombinant proteoglycan was expressed in murine fibroblasts. Versican expression induced a marked change in fibroblast phenotype. Functionally, the versican-expressing fibroblasts proliferated faster and displayed enhanced cell adhesion, but migrated slower than control cells. These changes in cell function were associated with greater N-cadherin and integrin β1 expression, along with increased FAK phosphorylation. The versican-expressing fibroblasts also displayed expression of smooth muscle α-actin, a marker of myofibroblast differentiation. Consistent with this observation, the versican fibroblasts displayed increased synthetic activity, as measured by collagen III mRNA expression, as well as a greater capacity to contract a collagen lattice. These changes appear to be mediated, at least in part, by an increase in active TGF-β signaling in the versican expressing fibroblasts, and this was measured by phosphorylation and nuclear accumulation of SMAD2.Collectively, these data indicate versican expression induces a myofibroblast-like phenotype in cultured fibroblasts.

  20. Fibroblast growth factor receptor 3 protein is overexpressed in oral and oropharyngeal squamous cell carcinoma

    NARCIS (Netherlands)

    Koole, Koos; van Kempen, Pauline M W; Swartz, Justin E; Peeters, Ton; van Diest, Paul J; Koole, Ron; van Es, Robert J. J.; Willems, Stefan M

    Fibroblast growth factor receptor 3 (FGFR3) is a member of the fibroblast growth factor receptor tyrosine kinase family. It has been identified as a promising therapeutic target in multiple types of cancer. We have investigated FGFR3 protein expression and FGFR3 gene copy-numbers in a single

  1. Stimulation of skin repair is dependent on fibroblast source and presence of extracellular matrix.

    NARCIS (Netherlands)

    Wang, H.; Pieper, J.S.; Schotel, R.; Blitterswijk, C.A. van; Lamme, E.N.

    2004-01-01

    In this study in vitro and in vivo functions were compared between cultured dermal equivalents produced with human fibroblasts isolated either from papillary dermis or adipose tissue of the same donors. Papillary dermal fibroblasts had a normal spindle cell shape; in contrast, adipose tissue

  2. Stimulation of skin repair is dependent on fibroblast source and presence of extracellular matrix

    NARCIS (Netherlands)

    Wang, Hongjun; Pieper, Jeroen; Schotel, Roka; van Blitterswijk, Clemens; Lamme, Evert N.

    2004-01-01

    In this study in vitro and in vivo functions were compared between cultured dermal equivalents produced with human fibroblasts isolated either from papillary dermis or adipose tissue of the same donors. Papillary dermal fibroblasts had a normal spindle cell shape; in contrast, adipose tissue

  3. Curcumin induces differential expression of cytoprotective enzymes but similar apoptotic responses in fibroblasts and myofibroblasts

    NARCIS (Netherlands)

    Lundvig, D.M.S.; Pennings, S.W.C.; Brouwer, K.M.; Mtaya-Mlangwa, M.; Mugonzibwa, E.A.; Kuijpers-Jagtman, A.M.; Hoff, J.W. Von den; Wagener, F.A.D.T.G.

    2015-01-01

    Excessive extracellular matrix (ECM) deposition and tissue contraction after injury can lead to esthetic and functional problems. Fibroblasts and myofibroblasts activated by transforming growth factor (TGF)-beta1 play a key role in these processes. The persistence of (myo)fibroblasts and their

  4. Murine gamma interferon fails to inhibit Toxoplasma gondii growth in murine fibroblasts.

    Science.gov (United States)

    Schwartzman, J D; Gonias, S L; Pfefferkorn, E R

    1990-01-01

    Although treatment of human macrophages or fibroblasts with human gamma interferon results in the inhibition of intracellular Toxoplasma gondii, murine gamma interferon stimulated only murine macrophages, not murine fibroblasts, to inhibit T. gondii. This species difference may be important in understanding the control of acute and chronic toxoplasmosis. PMID:2106497

  5. TRIF promotes angiotensin II-induced cross-talk between fibroblasts and macrophages in atrial fibrosis

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xiao-Qing; Zhang, Dao-Liang [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China); Zhang, Ming-Jian; Guo, Meng; Zhan, Yang-Yang; Liu, Fang [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Jiang, Wei-Feng; Zhou, Li [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China); Zhao, Liang, E-mail: zhaol_zg@163.com [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China); Wang, Quan-Xing, E-mail: wqxejd@126.com [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Liu, Xu, E-mail: liuxu_xk@163.com [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University, Shanghai (China)

    2015-08-14

    Aims: Atrial fibroblasts and macrophages have long been thought to participate in atrial fibrillation (AF). However, which specific mediator may regulate the interaction between them remains unclear. Methods and results: We provided the evidence for the involvement of Toll/IL-1 receptor domain-containing adaptor inducing IFN-β (TRIF), an important inflammation-related molecule, in the pathophysiology of AF. Patients with AF showed higher levels of angiotensin II (AngII) and TRIF expression and larger number of macrophages infiltration in left atria appendage than individuals with sinus rhythm (SR). In the cell study, AngII induced chemokines expressions in mouse atrial fibroblasts and AngII-stimulated atrial fibroblasts induced the chemotaxis of macrophages, which were reduced by losartan and TRIF siRNA. Meanwhile, AngII-stimulated atrial fibroblasts proliferation was enhanced by macrophages. Conclusions: Our data demonstrated that TRIF may be a crucial factor promoting the interaction between atrial fibroblasts and macrophages, leading to atrial fibrosis. - Highlights: • Compared with SR, AF showed higher TRIF expression in left atrial appendage. • TRIF siRNA reversed macrophage chemotaxis induced by AngII-treated fibroblast. • TRIF siRNA reversed chemokines expressions induced by AngII in fibroblast. • AngII-stimulated atrial fibroblast proliferation was enhanced by macrophage.

  6. Bovine trophectoderm cell lines induced from bovine fibroblasts with reprogramming factors

    Science.gov (United States)

    Bovine trophectoderm (TE) cells were induced [induced bovine trophectoderm-like (iBT)] from bovine fetal liver-derived fibroblasts, and other bovine fetal fibroblasts, after viral-vector transduction with either four or six reprogramming factors (RF), including POU5F1, KLF4, SOX2, C-MYC, SV40 large ...

  7. Mitochondrial Bioenergetics Is Altered in Fibroblasts from Patients with Sporadic Alzheimer's Disease

    Directory of Open Access Journals (Sweden)

    María J. Pérez

    2017-10-01

    Full Text Available The identification of an early biomarker to diagnose Alzheimer's disease (AD remains a challenge. Neuropathological studies in animal and AD patients have shown that mitochondrial dysfunction is a hallmark of the development of the disease. Current studies suggest the use of peripheral tissues, like skin fibroblasts as a possibility to detect the early pathological alterations present in the AD brain. In this context, we studied mitochondrial function properties (bioenergetics and morphology in cultured fibroblasts obtained from AD, aged-match and young healthy patients. We observed that AD fibroblasts presented a significant reduction in mitochondrial length with important changes in the expression of proteins that control mitochondrial fusion. Moreover, AD fibroblasts showed a distinct alteration in proteolytic processing of OPA1, a master regulator of mitochondrial fusion, compared to control fibroblasts. Complementary to these changes AD fibroblasts showed a dysfunctional mitochondrial bioenergetics profile that differentiates these cells from aged-matched and young patient fibroblasts. Our findings suggest that the human skin fibroblasts obtained from AD patients could replicate mitochondrial impairment observed in the AD brain. These promising observations suggest that the analysis of mitochondrial bioenergetics could represent a promising strategy to develop new diagnostic methods in peripheral tissues of AD patients.

  8. Governance of Cutaneous Photocarcinogenesis by Chronic UVA-Exposed Dermal Fibroblasts

    Science.gov (United States)

    2014-09-01

    fibroblasts and determine if bystander oxidative stress is induced by UVA irradiation and abrogated by antioxidants . In initial studies, we selected...primary human skin cells in order to avoid untoward effects of immortalization or transformation. Initial experiments determined direct effects...of UVA on normal human melanocytes (NHMs), fibroblasts (NHFs) and keratinocytes (NHKs). Figure 1A shows dose- dependent toxicity for three skin

  9. Aging alters functionally human dermal papillary fibroblasts but not reticular fibroblasts: a new view of skin morphogenesis and aging.

    Directory of Open Access Journals (Sweden)

    Solène Mine

    Full Text Available 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 a higher proportion of small cells with low granularity and a higher growth potential than Fr populations. However, these differences became less marked with increasing age of donors. Aging was also associated with changes in the secretion activity of both Fp and Fr. Using a reconstructed skin model, we evidenced that Fp and Fr cells do not possess equivalent capacities to sustain keratinopoiesis. Comparing Fp and Fr from young donors, we noticed that dermal equivalents containing Fp were more potent to promote epidermal morphogenesis than those containing Fr. These data emphasize the complexity of dermal fibroblast biology and document the specific functional properties of Fp and Fr. Our results suggest a new model of skin aging in which marked alterations of Fp may affect the histological characteristics of skin.

  10. Fibroblast growth factor receptor 4 (FGFR4) and fibroblast growth factor 19 (FGF19) autocrine enhance breast cancer cells survival.

    Science.gov (United States)

    Tiong, Kai Hung; Tan, Boon Shing; Choo, Heng Lungh; Chung, Felicia Fei-Lei; Hii, Ling-Wei; Tan, Si Hoey; Khor, Nelson Tze Woei; Wong, Shew Fung; See, Sze-Jia; Tan, Yuen-Fen; Rosli, Rozita; Cheong, Soon-Keng; Leong, Chee-Onn

    2016-09-06

    Basal-like breast cancer is an aggressive tumor subtype with poor prognosis. The discovery of underlying mechanisms mediating tumor cell survival, and the development of novel agents to target these pathways, is a priority for patients with basal-like breast cancer. From a functional screen to identify key drivers of basal-like breast cancer cell growth, we identified fibroblast growth factor receptor 4 (FGFR4) as a potential mediator of cell survival. We found that FGFR4 mediates cancer cell survival predominantly via activation of PI3K/AKT. Importantly, a subset of basal-like breast cancer cells also secrete fibroblast growth factor 19 (FGF19), a canonical ligand specific for FGFR4. siRNA-mediated silencing of FGF19 or neutralization of extracellular FGF19 by anti-FGF19 antibody (1A6) decreases AKT phosphorylation, suppresses cancer cell growth and enhances doxorubicin sensitivity only in the FGFR4+/FGF19+ breast cancer cells. Consistently, FGFR4/FGF19 co-expression was also observed in 82 out of 287 (28.6%) primary breast tumors, and their expression is strongly associated with AKT phosphorylation, Ki-67 staining, higher tumor stage and basal-like phenotype. In summary, our results demonstrated the presence of an FGFR4/FGF19 autocrine signaling that mediates the survival of a subset of basal-like breast cancer cells and suggest that inactivation of this autocrine loop may potentially serve as a novel therapeutic intervention for future treatment of breast cancers.

  11. EXPRESSION AND SIGNIFICANCE OF BASIC FIBROBLAST GWOWTH FACTOR AND FIBROBLAST GROWTH FACTOR RECEPTOR-1 IN OVARIAN EPITHELIAL NEOPLASM

    Institute of Scientific and Technical Information of China (English)

    高尚风; 杨蓉; 高博; 刘惠喜

    2003-01-01

    fibroblast growth factor (bFGF), fibroblast growth factor receptor-1 (FGFR-1) and carcinogenesis and progression of ovarian epithelial neoplasm. Methods Ten cases of normal ovarian tissues and 75 cases of ovarian epithelial neoplasm tissues were detected by immunohistochemical methods: S-P for bFGF, FGFR-1,double immunohistochemistry Lab-SA for Ki-67 antigen and bFGF. Results The expression level of bFGF, FGFR-1in ovarian epithelium and ovarian epithelial neoplasm showed a step-wise increase in the following order:normal〈benign〈borderline〈malignant; The expression level and intensity of bFGF and FGFR-1 were increased with the decrease of differentiation degree and increase of clinical stage in ovarian carcinoma; There was no statistical difference between the expression of bFGF, FGFR-1 in serous cystadenocarcinoma and that of mucinous cystadenocarcinoma; The expression of bFGF was correlated with that of FGFR-1 in neoplastic tissues; There were positive expression rates of bFGF and Ki-67 antigen in ovarian epithelial neoplasm. Conclusion As an important proliferative factor, bFGF plays an important role in carcinogenisis and progression of ovarian epithelial neoplasm.

  12. The initiation of embryonic-like collagen fibrillogenesis by adult human tendon fibroblasts when cultured under tension

    DEFF Research Database (Denmark)

    Bayer, Monika L; Yeung, Chin-Yan C; Kadler, Karl E

    2010-01-01

    Tendon fibroblasts synthesize collagen and form fibrils during embryonic development, but to what extent mature fibroblasts are able to recapitulate embryonic development and develop normal tendon structure is unknown. The present study examined the capability of mature human tendon fibroblasts t...... collagen fibrillogenesis similar to that of developing tendon, which implies that the hormonal/mechanical milieu, rather than intrinsic cellular function, inhibits regenerative potential in mature tendon.......Tendon fibroblasts synthesize collagen and form fibrils during embryonic development, but to what extent mature fibroblasts are able to recapitulate embryonic development and develop normal tendon structure is unknown. The present study examined the capability of mature human tendon fibroblasts...... to initiate collagen fibrillogenesis when cultured in fixed-length fibrin gels. Fibroblasts were dissected from semitendinosus and gracilis tendons from healthy humans and cultured in 3D linear fibrin gels. The fibroblasts synthesized an extracellular matrix of parallel collagen fibrils that were aligned...

  13. Ethanol exposure induces the cancer-associated fibroblast phenotype and lethal tumor metabolism: Implications for breast cancer prevention

    National Research Council Canada - National Science Library

    Sanchez-Alvarez, Rosa; Martinez-Outschoorn, Ubaldo E; Lin, Zhao; Lamb, Rebecca; Hulit, James; Howell, Anthony; Sotgia, Federica; Rubin, Emanuel; Lisanti, Michael P

    2013-01-01

    ...) and hTERT-immortalized fibroblasts. Here, we show that ethanol treatment (100 mM) promotes ROS production and oxidative stress in cancer-associated fibroblasts, which is sufficient to induce myofibroblastic differentiation...

  14. Inscribing Optical Excitability to Non-Excitable Cardiac Cells: Viral Delivery of Optogenetic Tools in Primary Cardiac Fibroblasts

    Science.gov (United States)

    Yu, Jinzhu; Entcheva, Emilia

    2016-01-01

    We describe in detail a method to introduce optogenetic actuation tools, a mutant version of channelrhodopsin- 2, ChR2(H134R), and archaerhodopsin (ArchT), into primary cardiac fibroblasts (cFB) in vitro by adenoviral infection to yield quick, robust, and consistent expression. Instructions on adjusting infection parameters such as the multiplicity of infection and virus incubation duration are provided to generalize the method for different lab settings or cell types. Specific conditions are discussed to create hybrid co-cultures of the optogenetically modified cFB and non-transformed cardiomyocytes to obtain light- sensitive excitable cardiac syncytium, including stencil-patterned cell growth. We also describe an all-optical framework for the functional testing of responsiveness of these opsins in cFB. The presented methodology provides cell-specific tools for the mechanistic investigation of the functional bioelectric contribution of different non-excitable cells in the heart and their electrical coupling to cardiomyocytes under different conditions. PMID:26965132

  15. Synergistic antitumor effects of in vivo production of human endostatin and tissue inhibitor of metalloproteinase-1 in mice after subcutaneous implantation of primary fibroblasts transfected by adenovirus-mediated gene delivery

    Institute of Scientific and Technical Information of China (English)

    SHEN Wei-gan; ZHU Jun; ZHANG Yu; SU Qing

    2010-01-01

    Background Tissue inhibitor of metalloproteinase (TIMP)-1 is a multifunctional protein. The aim of the study was to examine the feasibility of using a combination of adenovirus-mediated gene delivery of TIMP-1 plus endostatin and cell transplantation techniques to treat tumor growth and metastasis in mouse melanoma.Methods A enzyme-linked immunosorbent assay (ELISA) was used to detect the level of TIMP-1 and endostatin in vitro and in vivo. A tumor bearing mouse model and an experimental lung metastasis model in animal experiments were used to explore the therapeutic effect of in vivo production of human TIMP-1 and endostatin after the implantation of primary fibroblasts infected with the indicated adenovirus into tumor-bearing mice and a cytochemical method was used to observe histopathological changes of the tumor. An experimental lung metastasis model was established by injecting B16BL6 cells into the tail vein of mice and adenovirus-infected primary fibroblasts were subcutaneously implanted into the mice 24 hours later. Twenty-one days after tumor cell injection, mice were sacrificed to examine the effect on nodules visible as black forms on the surface of the lungs in B16BL6 cells.Results TIMP-1 and endostatin were secreted into the supernatants of cultures of Ad-TIMP-1 and Ad-End-infected mouse primary fibroblasts. We also observed that implantation of fibroblasts infected with Ad-TIMP-1 alone, Ad-End alone, or Ad-TIMP-1 plus Ad-End resulted in detectable blood levels which may clearly inhibit the tumor growth and metastasis in a murine melanoma model.Conclusion These results suggest the high capacity of transfection for the delivery of TIMP-1 or endostatin gene constructs into primary fibroblasts, and demonstrate that the implantation of TIMP-1 and endostatin producing fibroblasts at a site in vivo where direct secretion of TIMP-1 and endostatin into the blood is possible represented a promising approach for the development of cancer therapy.

  16. Stromal-epithelial interactions in aging and cancer: Senescent fibroblasts alter epithelial cell differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Parrinello, Simona; Coppe, Jean-Philippe; Krtolica, Ana; Campisi, Judith

    2004-07-14

    Cellular senescence suppresses cancer by arresting cells at risk for malignant tumorigenesis. However, senescent cells also secrete molecules that can stimulate premalignant cells to proliferate and form tumors, suggesting the senescence response is antagonistically pleiotropic. We show that premalignant mammary epithelial cells exposed to senescent human fibroblasts in mice irreversibly lose differentiated properties, become invasive and undergo full malignant transformation. Moreover, using cultured mouse or human fibroblasts and non-malignant breast epithelial cells, we show that senescent fibroblasts disrupt epithelial alveolar morphogenesis, functional differentiation, and branching morphogenesis. Further, we identify MMP-3 as the major factor responsible for the effects of senescent fibroblasts on branching morphogenesis. Our findings support the idea that senescent cells contribute to age-related pathology, including cancer, and describe a new property of senescent fibroblasts--the ability to alter epithelial differentiation--that might also explain the loss of tissue function and organization that is a hallmark of aging.

  17. A function for filamentous alpha-smooth muscle actin: Retardation of motility in human breast fibroblasts

    DEFF Research Database (Denmark)

    Rønnov-Jessen, Lone; Petersen, Ole William

    1996-01-01

    Actins are known to comprise six mammalian isoforms of which beta- and gamma-nonmuscle actins are present in all cells, whereas alpha-smooth muscle (alpha-sm) actin is normally restricted to cells of the smooth muscle lineages. alpha-Sm actin has been found also to be expressed transiently...... reactions. Here, we show that the presence of alpha-sm actin is a signal for retardation of migratory behavior in fibroblasts. Comparison in a migration assay of fibroblast cell strains with and without alpha-sm actin revealed migratory restraint in alpha-sm actin-positive fibroblasts. Electroporation...... in certain nonmuscle cells, in particular fibroblasts, which are referred to as myofibroblasts. The functional significance of alpha-sm actin in fibroblasts is unknown. However, myofibroblasts appear to play a prominent role in stromal reaction in breast cancer, at the site of wound repair, and in fibrotic...

  18. Mitochondrial Oxidative Stress due to Complex I Dysfunction Promotes Fibroblast Activation and Melanoma Cell Invasiveness

    Directory of Open Access Journals (Sweden)

    Maria Letizia Taddei

    2012-01-01

    Full Text Available Increased ROS (cellular reactive oxygen species are characteristic of both fibrosis and tumour development. ROS induce the trans-differentiation to myofibroblasts, the activated form of fibroblasts able to promote cancer progression. Here, we report the role of ROS produced in response to dysfunctions of mitochondrial complex I, in fibroblast activation and in tumour progression. We studied human fibroblasts with mitochondrial dysfunctions of complex I, leading to hyperproduction of ROS. We demonstrated that ROS level produced by the mutated fibroblasts correlates with their activation. The increase of ROS in these cells provides a greater ability to remodel the extracellular matrix leading to an increased motility and invasiveness. Furthermore, we evidentiated that in hypoxic conditions these fibroblasts cause HIF-1α stabilization and promote a proinvasive phenotype of human melanoma cells through secretion of cytokines. These data suggest a possible role of deregulated mitochondrial ROS production in fibrosis evolution as well as in cancer progression and invasion.

  19. Staph Infections

    Science.gov (United States)

    ... ill or the area spreads and gets very red or and hot — it's a good idea to see a doctor. Wound infections generally show up 2 or more days after the injury or surgery. The signs of a wound infection (redness, pain, swelling, and warmth) are similar to those found in cellulitis. A ...

  20. Staphylococcal Infections

    Science.gov (United States)

    ... you should be familiar with include the following: Cellulitis is a bacterial skin infection that first affects the outer layers of the ... skin. Although other types of bacteria can cause cellulitis, Saureus ... may diagnose the infection by examining the area. The doctor may take ...

  1. Skin Infections

    Science.gov (United States)

    ... Old Feeding Your 8- to 12-Month-Old Feeding Your 1- to 2-Year-Old First Aid: Skin ... (bacterial infection of the deeper layers of the skin and tissues beneath) are typical childhood skin infections. The usual bacterial culprits in skin ...

  2. Mutant Parkin impairs mitochondrial function and morphology in human fibroblasts.

    Directory of Open Access Journals (Sweden)

    Anne Grünewald

    Full Text Available BACKGROUND: Mutations in Parkin are the most common cause of autosomal recessive Parkinson disease (PD. The mitochondrially localized E3 ubiquitin-protein ligase Parkin has been reported to be involved in respiratory chain function and mitochondrial dynamics. More recent publications also described a link between Parkin and mitophagy. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the impact of Parkin mutations on mitochondrial function and morphology in a human cellular model. Fibroblasts were obtained from three members of an Italian PD family with two mutations in Parkin (homozygous c.1072delT, homozygous delEx7, compound-heterozygous c.1072delT/delEx7, as well as from two relatives without mutations. Furthermore, three unrelated compound-heterozygous patients (delEx3-4/duplEx7-12, delEx4/c.924C>T and delEx1/c.924C>T and three unrelated age-matched controls were included. Fibroblasts were cultured under basal or paraquat-induced oxidative stress conditions. ATP synthesis rates and cellular levels were detected luminometrically. Activities of complexes I-IV and citrate synthase were measured spectrophotometrically in mitochondrial preparations or cell lysates. The mitochondrial membrane potential was measured with 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide. Oxidative stress levels were investigated with the OxyBlot technique. The mitochondrial network was investigated immunocytochemically and the degree of branching was determined with image processing methods. We observed a decrease in the production and overall concentration of ATP coinciding with increased mitochondrial mass in Parkin-mutant fibroblasts. After an oxidative insult, the membrane potential decreased in patient cells but not in controls. We further determined higher levels of oxidized proteins in the mutants both under basal and stress conditions. The degree of mitochondrial network branching was comparable in mutants and

  3. Increased fibroblast telomerase expression precedes myofibroblast α-smooth muscle actin expression in idiopathic pulmonary fibrosis

    Directory of Open Access Journals (Sweden)

    Daniel Reis Waisberg

    2012-09-01

    Full Text Available OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast α-smooth muscle actin expression and the tissue expression of inter leu kin-4, transforming growth factor-β, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-β expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast α-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast α-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast α-smooth muscle actin expression predominates in areas of late remodeling

  4. Epigenetic and phenotypic profile of fibroblasts derived from induced pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Kyle J Hewitt

    Full Text Available Human induced pluripotent stem (hiPS cells offer a novel source of patient-specific cells for regenerative medicine. However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES cells remain unclear. We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK and their hES-derived counterparts (EDK showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts. Array-based methylation analysis was performed for EDK, iPDK and their parental hES and iPS cell lines, and hierarchical clustering revealed that EDK and iPDK had closely-related methylation profiles. DNA methylation analysis of promoter regions associated with extracellular matrix (ECM-production (COL1A1 by iPS- and hESC-derived fibroblasts and fibroblast lineage commitment (PDGFRβ, revealed promoter demethylation linked to their expression, and patterns of transcription and methylation of genes related to the functional properties of mature stromal cells were seen in both hiPS- and hES-derived fibroblasts. iPDK cells also showed functional properties analogous to those of hES-derived and mature fibroblasts, as seen by their capacity to direct the morphogenesis of engineered human skin equivalents. Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

  5. Upregulation of the N-formyl Peptide receptors in scleroderma fibroblasts fosters the switch to myofibroblasts.

    Science.gov (United States)

    Rossi, Francesca Wanda; Napolitano, Filomena; Pesapane, Ada; Mascolo, Massimo; Staibano, Stefania; Matucci-Cerinic, Marco; Guiducci, Serena; Ragno, Pia; di Spigna, Gaetano; Postiglione, Loredana; Marone, Gianni; Montuori, Nunzia; de Paulis, Amato

    2015-06-01

    Systemic sclerosis (SSc) is characterized by chronic inflammation and fibrosis. N-Formyl peptide (fMLF) receptors (FPRs) are chemotactic receptors involved in inflammation. Three FPRs have been identified: FPR1, FPR2, and FPR3. We have examined, by RT-PCR, Western blot and immunohistochemistry, FPRs expression in skin fibroblasts from 10 normal subjects and 10 SSc patients, showing increased expression in SSc fibroblasts. Several functions of FPRs occur through the interaction with a region of the urokinase-type plasminogen activator receptor (uPAR88-92), able to interact with FPRs and to mediate urokinase (uPA) or fMLF-dependent cell migration. Soluble uPAR84-95 peptide can act as a direct ligand of FPRs. Furthermore, uPA or its aminoterminal fragment (ATF) can promote the exposure of the uPAR88-92 region. The WKYMVm peptide is a FPRs pan-agonist. We investigated the functional effects of these agonists on normal and SSc fibroblasts. ATF, uPAR84-95, and WKYMVm regulated adhesion, migration, and proliferation of normal fibroblasts. Despite FPR overexpression, the response of SSc fibroblasts to the same agonists was greatly reduced, except for the proliferative response to ATF. SSc fibroblasts showed increased α-smooth muscle actin expression and improved capability to induce wound closure. Indeed, they overexpressed a cleaved uPAR form, exposing the uPAR88-92 region, and vitronectin, both involved in fibrosis and in the fibroblast-to-myofibroblast transition. FPR stimulation promoted α-smooth muscle actin expression in normal fibroblasts as well as motility, matrix deposition, αvβ5 integrin expression, and radical oxygen species generation in normal and SSc fibroblasts. This study provides evidence that FPRs may play a role in fibrosis and in the fibroblast-to-myofibroblast transition. Copyright © 2015 by The American Association of Immunologists, Inc.

  6. CTRP6 inhibits fibrogenesis in TGF-β1-stimulated human dermal fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Rong-hui, E-mail: fan_ronghuixa@163.com [Department of Burn and Plastic Surgery, Shaanxi Provincial People’s Hospital, Xi’an 710068 (China); Zhu, Xiu-mei; Sun, Yao-wen [Department of Burn and Plastic Surgery, Shaanxi Provincial People’s Hospital, Xi’an 710068 (China); Peng, Hui-zi [Department of Cosmetology Plastic Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shaanxi 710061 (China); Wu, Hang-li; Gao, Wen-jie [Department of Burn and Plastic Surgery, Shaanxi Provincial People’s Hospital, Xi’an 710068 (China)

    2016-07-08

    Skin fibrosis is characterized by excessive proliferation of fibroblasts and overproduction of extracellular matrix (ECM). C1q/tumor necrosis factor-related protein 6 (CTRP6), a member of CTRPs, has been involved in the development of cardiac fibrosis. However, the function and detailed regulatory mechanism of CTRP6 in skin fibrosis remain unclear. The aim of this study was to investigate the effect of CTRP6 on the activation of human dermal fibroblasts. Our results showed that CTRP6 was lowly expressed in scar tissues and transforming growth factor-β1 (TGF-β1)-treated dermal fibroblasts. CTRP6 overexpression significantly inhibited the proliferation of dermal fibroblasts, as well as suppressed the expression of ECM in TGF-β1-treated dermal fibroblasts. Furthermore, CTRP6 overexpression markedly inhibited TGF-β1-induced phosphorylation of Smad3 in dermal fibroblasts. In conclusion, the data reported here demonstrate that CTRP6 is able to inhibit the proliferation and ECM expression in human dermal fibroblasts through suppressing the TGF-β1/Smad3 signaling pathway. These findings suggest that CTRP6 may be a potential therapeutic target for the prevention of skin fibrosis. -- Highlights: •CTRP6 expression was decreased in scar tissues and TGF-β1-treated dermal fibroblasts. •CTRP6 inhibits TGF-β1-induced the proliferation of dermal fibroblasts. •CTRP6 inhibits expression of collagen type I and α-SMA. •CTRP6 inhibits the activation of TGF-β1/Smad3 signaling pathway in dermal fibroblasts.

  7. Temporal alterations in cardiac fibroblast function following induction of pressure overload

    Science.gov (United States)

    Stewart, James A.; Massey, Erin P.; Fix, Charity; Zhu, Jinyu; Goldsmith, Edie C.

    2014-01-01

    Increases in cardiovascular load (pressure overload) are known to elicit ventricular remodeling including cardiomyocyte hypertrophy and interstitial fibrosis. While numerous studies have focused on the mechanisms of myocyte hypertrophy, comparatively little is known regarding the response of the interstitial fibroblasts to increased cardiovascular load. Fibroblasts are the most numerous cell type in the mammalian myocardium and have long been recognized as producing the majority of the myocardial extracellular matrix. It is only now becoming appreciated that other aspects of fibroblast behavior are important to overall cardiac function. The present studies were performed to examine the temporal alterations in fibroblast activity in response to increased cardiovascular load. Rat myocardial fibroblasts were isolated at specific time-points (3, 7, 14, and 28 days) after induction of pressure overload by abdominal aortic constriction. Bioassays were performed to measure specific parameters of fibroblast function including remodeling and contraction of 3-dimensional collagen gels, migration, and proliferation. In addition, the expression of extracellular matrix receptors of the integrin family was examined. Myocardial hypertrophy and fibrosis were evident within 7 days after constriction of the abdominal aorta. Collagen gel contraction, migration, and proliferation were enhanced in fibroblasts from pressure-overloaded animals compared to fibroblasts from sham animals. Differences in fibroblast function and protein expression were evident within 7 days of aortic constriction, concurrent with the onset of hypertrophy and fibrosis of the intact myocardium. These data provide further support for the idea that rapid and dynamic changes in fibroblast phenotype accompany and contribute to the progression of cardiovascular disease. PMID:20217135

  8. Identification of human fibroblast cell lines as a feeder layer for human corneal epithelial regeneration.

    Directory of Open Access Journals (Sweden)

    Rong Lu

    Full Text Available There is a great interest in using epithelium generated in vitro for tissue bioengineering. Mouse 3T3 fibroblasts have been used as a feeder layer to cultivate human epithelia including corneal epithelial cells for more than 3 decades. To avoid the use of xeno-components, we evaluated human fibroblasts as an alternative feeder supporting human corneal epithelial regeneration. Five human fibroblast cell lines were used for evaluation with mouse 3T3 fibroblasts as a control. Human epithelial cells isolated from fresh corneal limbal tissue were seeded on these feeders. Colony forming efficiency (CFE and cell growth capacity were evaluated on days 5-14. The phenotype of the regenerated epithelia was evaluated by morphology and immunostaining with epithelial markers. cDNA microarray was used to analyze the gene expression profile of the supportive human fibroblasts. Among 5 strains of human fibroblasts evaluated, two newborn foreskin fibroblast cell lines, Hs68 and CCD1112Sk, were identified to strongly support human corneal epithelial growth. Tested for 10 passages, these fibroblasts continually showed a comparative efficiency to the 3T3 feeder layer for CFE and growth capacity of human corneal epithelial cells. Limbal epithelial cells seeded at 1 × 10(4 in a 35-mm dish (9.6 cm(2 grew to confluence (about 1.87-2.41 × 10(6 cells in 12-14 days, representing 187-241 fold expansion with over 7-8 doublings on these human feeders. The regenerated epithelia expressed K3, K12, connexin 43, p63, EGFR and integrin β1, resembling the phenotype of human corneal epithelium. DNA microarray revealed 3 up-regulated and 10 down-regulated genes, which may be involved in the functions of human fibroblast feeders. These findings demonstrate that commercial human fibroblast cell lines support human corneal epithelial regeneration, and have potential use in tissue bioengineering for corneal reconstruction.

  9. Cathelicidin suppresses colon cancer development by inhibition of cancer associated fibroblasts

    Science.gov (United States)

    Cheng, Michelle; Ho, Samantha; Yoo, Jun Hwan; Tran, Deanna Hoang-Yen; Bakirtzi, Kyriaki; Su, Bowei; Tran, Diana Hoang-Ngoc; Kubota, Yuzu; Ichikawa, Ryan; Koon, Hon Wai

    2015-01-01

    Background Cathelicidin (LL-37 in humans and mCRAMP in mice) represents a family of endogenous antimicrobial and anti-inflammatory peptides. Cancer-associated fibroblasts can promote the proliferation of colon cancer cells and growth of colon cancer tumors. Methods We examined the role of cathelicidin in the development of colon cancer, using subcutaneous human HT-29 colon-cancer-cell-derived tumor model in nude mice and azoxymethane- and dextran sulfate-mediated colon cancer model in C57BL/6 mice. We also determined the indirect antitumoral mechanism of cathelicidin via the inhibition of epithelial–mesenchymal transition (EMT) of colon cancer cells and fibroblast-supported colon cancer cell proliferation. Results Intravenous administration of cathelicidin expressing adeno-associated virus significantly reduced the size of tumors, tumor-derived collagen expression, and tumor-derived fibroblast expression in HT-29-derived subcutaneous tumors in nude mice. Enema administration of the mouse cathelicidin peptide significantly reduced the size and number of colonic tumors in azoxymethane- and dextran sulfate-treated mice without inducing apoptosis in tumors and the adjacent normal colonic tissues. Cathelicidin inhibited the collagen expression and vimentin-positive fibroblast expression in colonic tumors. Cathelicidin did not directly affect HT-29 cell viability, but did significantly reduce tumor growth factor-β1-induced EMT of colon cancer cells. Media conditioned by the human colonic CCD-18Co fibroblasts promoted human colon cancer HT-29 cell proliferation. Cathelicidin pretreatment inhibited colon cancer cell proliferation mediated by media conditioned by human colonic CCD-18Co fibroblasts. Cathelicidin disrupted tubulin distribution in colonic fibroblasts. Disruption of tubulin in fibroblasts reduced fibroblast-supported colon cancer cell proliferation. Conclusion Cathelicidin effectively inhibits colon cancer development by interfering with EMT and fibroblast

  10. Ascorbic acid inhibits replication and infectivity of avian RNA tumor virus

    Energy Technology Data Exchange (ETDEWEB)

    BISSELL, MINA J; HATIE, CARROLL; FARSON, DEBORAH A.; SCHWARZ, RICHARD I.; SOO, WHAI-JEN

    1980-04-01

    Ascorbic acid, at nontoxic concentrations, causes a substantial reduction in the ability of avian tumor viruses to replicate in both primary avian tendon cells and chicken embryo fibroblasts. The virus-infected cultures appear to be less transformed in the presence of ascorbic acid by the criteria of morphology, reduced glucose uptake, and increased collagen synthesis. The vitamin does not act by altering the susceptibility of the cells to initial infection and transformation, but instead appears to interfere with the spread of infection through a reduction in virus replication and virus infectivity. The effect is reversible and requires the continuous presence of the vitamin in the culture medium.

  11. The initiation of embryonic-like collagen fibrillogenesis by adult human tendon fibroblasts when cultured under tension

    DEFF Research Database (Denmark)

    Bayer, Monika L; Yeung, Chin-Yan C; Kadler, Karl E

    2010-01-01

    Tendon fibroblasts synthesize collagen and form fibrils during embryonic development, but to what extent mature fibroblasts are able to recapitulate embryonic development and develop normal tendon structure is unknown. The present study examined the capability of mature human tendon fibroblasts t...

  12. File list: Oth.PSC.10.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.10.AllAg.iPS_derived_fibroblasts hg19 TFs and others Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.PSC.10.AllAg.iPS_derived_fibroblasts.bed ...

  13. File list: Oth.PSC.50.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.50.AllAg.iPS_derived_fibroblasts hg19 TFs and others Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.PSC.50.AllAg.iPS_derived_fibroblasts.bed ...

  14. File list: DNS.PSC.20.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.PSC.20.AllAg.hESC_derived_fibroblasts hg19 DNase-seq Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.PSC.20.AllAg.hESC_derived_fibroblasts.bed ...

  15. File list: InP.PSC.10.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.PSC.10.AllAg.hESC_derived_fibroblasts hg19 Input control Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.PSC.10.AllAg.hESC_derived_fibroblasts.bed ...

  16. File list: NoD.PSC.20.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available NoD.PSC.20.AllAg.hESC_derived_fibroblasts hg19 No description Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.PSC.20.AllAg.hESC_derived_fibroblasts.bed ...

  17. File list: DNS.PSC.20.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.PSC.20.AllAg.iPS_derived_fibroblasts hg19 DNase-seq Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.PSC.20.AllAg.iPS_derived_fibroblasts.bed ...

  18. File list: DNS.PSC.10.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available DNS.PSC.10.AllAg.hESC_derived_fibroblasts hg19 DNase-seq Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/DNS.PSC.10.AllAg.hESC_derived_fibroblasts.bed ...

  19. File list: NoD.PSC.10.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available NoD.PSC.10.AllAg.hESC_derived_fibroblasts hg19 No description Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.PSC.10.AllAg.hESC_derived_fibroblasts.bed ...

  20. File list: NoD.PSC.50.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available NoD.PSC.50.AllAg.iPS_derived_fibroblasts hg19 No description Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.PSC.50.AllAg.iPS_derived_fibroblasts.bed ...

  1. File list: InP.PSC.10.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.PSC.10.AllAg.iPS_derived_fibroblasts hg19 Input control Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.PSC.10.AllAg.iPS_derived_fibroblasts.bed ...

  2. File list: NoD.PSC.20.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available NoD.PSC.20.AllAg.iPS_derived_fibroblasts hg19 No description Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/NoD.PSC.20.AllAg.iPS_derived_fibroblasts.bed ...

  3. File list: Oth.PSC.50.AllAg.hESC_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.PSC.50.AllAg.hESC_derived_fibroblasts hg19 TFs and others Pluripotent stem cell hESC derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Oth.PSC.50.AllAg.hESC_derived_fibroblasts.bed ...

  4. File list: InP.PSC.50.AllAg.iPS_derived_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.PSC.50.AllAg.iPS_derived_fibroblasts hg19 Input control Pluripotent stem cell iPS derived... fibroblasts http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.PSC.50.AllAg.iPS_derived_fibroblasts.bed ...

  5. File list: ALL.Oth.10.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.10.AllAg.3T3_fibroblasts mm9 All antigens Others 3T3 fibroblasts SRX105582,...SRX099255,SRX099259,SRX099260,SRX099261,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.10.AllAg.3T3_fibroblasts.bed ...

  6. File list: His.Oth.50.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Oth.50.AllAg.3T3_fibroblasts mm9 Histone Others 3T3 fibroblasts SRX099255,SRX10...5582,SRX099259,SRX099260 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Oth.50.AllAg.3T3_fibroblasts.bed ...

  7. File list: InP.Oth.20.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.Oth.20.AllAg.3T3_fibroblasts mm9 Input control Others 3T3 fibroblasts SRX099261...,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Oth.20.AllAg.3T3_fibroblasts.bed ...

  8. File list: InP.Oth.50.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available InP.Oth.50.AllAg.3T3_fibroblasts mm9 Input control Others 3T3 fibroblasts SRX099261...,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Oth.50.AllAg.3T3_fibroblasts.bed ...

  9. Basement membrane reconstruction in human skin equivalents is regulated by fibroblasts and/or exogenously activated keratinocytes

    NARCIS (Netherlands)

    El Ghalbzouri, A; Jonkman, MF; Dijkman, R; Ponec, M

    2005-01-01

    This study was undertaken to examine the role fibroblasts play in the formation of the basement membrane (BM) in human skin equivalents. For this purpose, keratinocytes were seeded on top of fibroblast-free or fibroblast-populated collagen matrix or de-epidermized dermis and cultured in the absence

  10. File list: ALL.Oth.50.AllAg.3T3_fibroblasts [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Oth.50.AllAg.3T3_fibroblasts mm9 All antigens Others 3T3 fibroblasts SRX099255,...SRX105582,SRX099259,SRX099260,SRX099261,SRX099262 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Oth.50.AllAg.3T3_fibroblasts.bed ...

  11. 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.

  12. Malignant transformation of diploid human fibroblasts by transfection of oncogenes

    Energy Technology Data Exchange (ETDEWEB)

    McCormick, J.J.

    1992-01-01

    This document consist of brief reports prepared by postdoctoral students supported by the project, each describing his accomplishments under the grant. Topics include (1) Malignant Transformation of MSU-1. 1 Cells by Gamma Radiation, (2) Correlation between Levels of ras Expression and Presence of Transformed Phenotypes Including Tumorigenicity, Using a Modulatable Promoter, (3) Relation between Specific rad Oncogene Expression, (4) Correlation of Genetic Changes in Fibroblastic Tumors with Malignancies, (5)Transformation of MSU-1.1 Cells by sis Oncogene, (6) Malignant Transformation of MSU-1.0 Cells, (7) Correlation of Urokinase Plasminogen Activation (mu-PA) with Malignant Phenotype, (8)Two Dimensional Gel Electrophoresis Studies of the Proteins of the Major Cell Strains of the MSU-1 Family of Cells, and (9) Correlation between Proteinase Activity Levels and Malignancy.

  13. Fibroblast Growth Factor 23 in Long-Duration Spaceflight

    Science.gov (United States)

    Bokhari, R.; Zwart, S. R.; Fields, E.; Heer, M.; Sibonga, J.; Smith, S. M.

    2015-01-01

    Many nutritional factors influence bone, from the basics of calcium and vitamin D, to factors which influence bone through acid/base balance, including protein, sodium, and more. Fibroblast growth factor 23 (FGF23) is a recently identified factor, secreted from osteocytes, which is involved in classic (albeit complex) feedback loops controlling phosphorus homeostasis through both vitamin D and parathyroid hormone (PTH) (1, 2). As osteocytes are gravity sensing cells, it is important to determine if there are changes in FGF23 during spaceflight. In extreme cases, such as chronic kidney disease, FGF23 levels are highly elevated. FGF23 imbalances, secondary to dietary influences, may contribute to skeletal demineralization and kidney stone risk during spaceflight.

  14. Cancer-associated fibroblasts as targets for immunotherapy.

    Science.gov (United States)

    Kakarla, Sunitha; Song, Xiao-Tong; Gottschalk, Stephen

    2012-11-01

    Immunotherapy for solid tumors has shown promise in preclinical as well as early clinical studies. However, its efficacy remains limited. The hindrance to achieving objective, long-lasting therapeutic responses in solid tumors is, in part, mediated by the dynamic nature of the tumor and its complex microenvironment. Tumor-directed therapies fail to eliminate components of the microenvironment, which can reinstate a tumorigenic milieu and contribute to recurrence. Cancer-associated fibroblasts (CAFs) form the most preponderant cell type in the solid tumor microenvironment. Given their pervasive role in facilitating tumor growth and metastatic dissemination, CAFs have emerged as attractive therapeutic targets in the tumor microenvironment. In this article, we highlight the cross-talk between CAFs and cancer cells, and discuss how targeting CAFs has the potential to improve current immunotherapy approaches for cancer.

  15. EB1 is required for primary cilia assembly in fibroblasts

    DEFF Research Database (Denmark)

    Schrøder, Jacob M; Schneider, Linda; Christensen, Søren T

    2007-01-01

    EB1 is a small microtubule (MT)-binding protein that associates preferentially with MT plus ends and plays a role in regulating MT dynamics. EB1 also targets other MT-associated proteins to the plus end and thereby regulates interactions of MTs with the cell cortex, mitotic kinetochores, and diff......EB1 is a small microtubule (MT)-binding protein that associates preferentially with MT plus ends and plays a role in regulating MT dynamics. EB1 also targets other MT-associated proteins to the plus end and thereby regulates interactions of MTs with the cell cortex, mitotic kinetochores...... in the cilium/flagellum is unknown. We depleted EB1 from NIH3T3 fibroblasts by using siRNA and found that EB1 depletion causes a approximately 50% reduction in the efficiency of primary cilia assembly in serum-starved cells. Expression of dominant-negative EB1 also inhibited cilia formation, and expression...

  16. Cytoskeletal elements of chick embryo fibroblasts revealed by detergent extraction.

    Science.gov (United States)

    Brown, S; Levinson, W; Spudich, J A

    1976-01-01

    Treatment of chick embryo fibroblasts with 0.5% Triton X-100 extracts most of the cell protein, leaving an organized part of the cell structure attached to the tissue culture dish. This "Triton cytoskeleton" consists largely of intermediate-sized filaments and bundles of microfilaments. SDS polyacrylamide gel electrophoresis reveals that this cytoskeleton is made up of three main proteins. One protein component is 42,000 daltons and co-migrates with muscle actin. The other two components are 52,000 and 230,000 daltons and remain quantitatively associated with the cytoskeleton during the detergent extraction. The possible identity of these three protein components and their organization into a supramolecular structure is discussed.

  17. Fibroblast Growth Factors: Biology, Function, and Application for Tissue Regeneration

    Directory of Open Access Journals (Sweden)

    Ye-Rang Yun

    2010-01-01

    Full Text Available Fibroblast growth factors (FGFs that signal through FGF receptors (FGFRs regulate a broad spectrum of biological functions, including cellular proliferation, survival, migration, and differentiation. The FGF signal pathways are the RAS/MAP kinase pathway, PI3 kinase/AKT pathway, and PLCγ pathway, among which the RAS/MAP kinase pathway is known to be predominant. Several studies have recently implicated the in vitro biological functions of FGFs for tissue regeneration. However, to obtain optimal outcomes in vivo, it is important to enhance the half-life of FGFs and their biological stability. Future applications of FGFs are expected when the biological functions of FGFs are potentiated through the appropriate use of delivery systems and scaffolds. This review will introduce the biology and cellular functions of FGFs and deal with the biomaterials based delivery systems and their current applications for the regeneration of tissues, including skin, blood vessel, muscle, adipose, tendon/ligament, cartilage, bone, tooth, and nerve tissues.

  18. Tattoo ink nanoparticles in skin tissue and fibroblasts.

    Science.gov (United States)

    Grant, Colin A; Twigg, Peter C; Baker, Richard; 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 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.

  19. A method for quantifying mechanical properties of tissue following viral infection.

    Directory of Open Access Journals (Sweden)

    Vy Lam

    Full Text Available Viral infection and replication involves the reorganization of the actin network within the host cell. Actin plays a central role in the mechanical properties of cells. We have demonstrated a method to quantify changes in mechanical properties of fabricated model three-dimensional (3D connective tissue following viral infection. Using this method, we have characterized the impact of infection by the human herpesvirus, cytomegalovirus (HCMV. HCMV is a member of the herpesvirus family and infects a variety of cell types including fibroblasts. In the body, fibroblasts are necessary for maintaining connective tissue and function by creating mechanical force. Using this 3D connective tissue model, we observed that infection disrupted the cell's ability to generate force and reduced the cumulative contractile force of the tissue. The addition of HCMV viral particles in the absence of both viral gene expression and DNA replication was sufficient to disrupt tissue function. We observed that alterations of the mechanical properties are, in part, due to a disruption of the underlying complex actin microfilament network established by the embedded fibroblasts. Finally, we were able to prevent HCMV-mediated disruption of tissue function by the addition of human immune globulin against HCMV. This study demonstrates a method to quantify the impact of viral infection on mechanical properties which are not evident using conventional cell culture systems.

  20. Effects of plant lectins on in vitro fibroblast proliferation

    Directory of Open Access Journals (Sweden)

    Ana Maria Sell

    2003-06-01

    Full Text Available Lectins are carbohydrate-binding proteins that have been isolated from various sources and presented a wide spectrum of biological activities. The effects of four lectins, namely, Phaseolus vulgaris phytohemagglutinin, PHA, wheat germ agglutinin, WGA, Artocarpus integrifolia seed lectins, jacalin and artocarpin, on in vitro fibroblasts proliferation were investigated. The lectins did not influence the initial cell adhesion to the plate. PHA and WGA at 10-20 µg/mL concentrations significantly decreased fibroblasts proliferation. At these concentrations, they caused morphological alterations on cells and over 80 µg/mL, promoted cell death. Neither jacalin nor artocarpin significantly affected cell proliferation.O objetivo deste trabalho foi avaliar a influência das lectinas PHA, WGA, jacalina e artocarpina sobre a proliferação de fibroblastos in vitro. Para tanto, fibroblastos gengivais de voluntários saudáveis foram cultivados, por cinco dias, em DMEM suplementado com soro bovino fetal (10% v/v e na presença das lectinas nas concentrações finais de 0.1 a 300 µg/mL. A adesão, o crescimento e a morfologia celular foram acompanhados por microscopia de inversão e contraste de fase. O índice de proliferação foi avaliado pelo método calorimétrico usando MTT. As lectinas não alteraram a adesão inicial dos fibroblastos à placa de poliestireno. PHA e WGA, nas concentrações de 10 a 20 µg/mL, diminuíram significativamente a proliferação celular. Nestas concentrações a morfologia celular é alterada e acima de 80 µg/mL, houve100% de morte celular. As lectinas jacalina e artocarpina não influenciaram a proliferação celular.