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At least 1,225 records · Page 68Linked to original sources
PubMed2001

Overexpression of collagenase (MMP-1) and stromelysin (MMP-3) by pterygium head fibroblasts.

BACKGROUND: The balance between matrix metalloproteinases (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) determines the extent of connective tissue degradation and remodeling. OBJECTIVE: To determine whether pterygium, characterized by fibrovascular invasion into the cornea, may in part be mediated by an increased activity of MMPs. MATERIALS AND METHODS: Expression of transcripts and proteins of MMPs, TIMPs, and urokinase plasminogen activator (uPA) by cultured human pterygium head, body, and subconjunctival fibroblasts, and normal corneal and conjunctival fibroblasts were determined by Northern hybridization, enzyme-linked immunosorbent assay, Western blotting, zymography, and quantitative collagenase assay, respectively. RESULTS: Compared with normal conjunctival fibroblasts from 6 subjects, the expression of MMP-1 and MMP-3 transcripts was dramatically increased in pterygium head fibroblasts of 8 patients, but not in pterygium body fibroblasts of 6 patients. The protein levels and collagenolytic and caseinolytic activities of MMP-1 and/or MMP-3 were also markedly increased in pterygium head fibroblasts. The MMP-1 and MMP-3 proteins and activity decreased in order from pterygium head to body to subconjunctival fibroblasts. There was no difference in the transcript and protein expression of MMP-2, TIMP-1, TIMP-2, and uPA among these groups. CONCLUSION: Pterygium head fibroblasts express increased mRNA, protein, and activities of MMP-1 and MMP-3. CLINICAL RELEVANCE: Overexpression of MMP-1 and MMP-3, a phenotype previously linked with UV exposure in dermal fibroblasts to explain the pathologic finding of elastotic degeneration, suggests that pterygium head fibroblasts might be intrinsically altered by UV, which might be responsible for corneal invasion.

Adult↗
PubMedDate not supplied

Cortical fibroblast culture from human biopsies.

BACKGROUND: Tubulointerstitial fibrosis is an integral part of progressive renal disease. Human cortical fibroblasts are believed to be key effector cells in fibrogenesis. Thus, a reliable culture of these cells is necessary for studies of their pathophysiology. METHODS: Cortical fibroblast culture from routine kidney biopsies were analyzed and the cells were characterized. Indirect immunofluorescence staining was done after the first passage for cytokeratin, vimentin, alpha-smooth muscle actin, CD 44, CD 54, CD 68, collagen types I, III, and HLA-DR. We then assessed the utility of the putative fibroblast markers CD 90, prolyl-4-hydroxylase (P4H) and F1b in simultaneous stainings of tubular epithelial cells. RESULTS: During the study period, 49 biopsy cores were cultured and cortical fibroblasts could be successfully established in 21 cases (42.9%). There was no relation between the success rate of culture and the degree of interstitial fibrosis, but an association was seen with the time of completion of the first passage. There was a negative correlation between the extent of scarring and the percentage of cytokeratin positive cells (r = -0.66, p < 0.001). All primary fibroblasts were negative for factor VIII, HLA-DR, CD 68, and cytokeratin. They expressed alpha-smooth muscle actin and collagen types I and III to variable degrees. There was a robust correlation between the percentage of alpha-smooth muscle actin positive cells and interstitial scarring but no such association with collagen type I or type III positive cells. The three putative fibroblast markers did not prove useful in differentiating between tubular epithelial cells and fibroblasts. However, since only fibroblasts stained positive for CD 90 and negative for cytokeratin, these two markers may suffice to distinguish fibroblasts from other renal cellular elements. CONCLUSIONS: Cortical renal fibroblasts can be easily cultured from kidney biopsy cores, though the success rate of pure cultures is below 50%. Staining for CD 90 and cytokeratin may suffice for initial characterization of these cells.

Biopsy↗
PubMed2003

Cyclooxygenase-2 expression and prostanoid biogenesis reflect clinical phenotype in human colorectal fibroblast strains.

Up-regulated cyclooxygenase (COX)-2 expression and prostaglandin E2 (PGE2)synthesis contribute causally to the early stages of colorectal neoplasia and carcinogenesis, yet COX-2 expression is barely detectable in normal and premalignant colorectal epithelium. Rather, COX-2 expression in nonmalignant colonic tissue is probably confined to subepithelial cells, such as fibroblasts. We established a panel of 33 primary subepithelial fibroblast strains from human colonoscopic biopsies of normal colon (group I), normal segments of colons that harbored synchronous advanced neoplasms in remote segments (group II), advanced neoplasms (group III), and segments of active ulcerative colitis (group IV). In group I strains, mean basal and peak PGE2 levels after 24 h of interleukin (IL)-1beta stimulation were 5.4 +/- 1.1 and 32.8 +/- 4.9 ng/mg protein, respectively. Mean IL-1beta-stimulated peak levels in groups II, III, and IV strains were, respectively, 6-, 9-, and 7-fold greater than that in group I (P < 0.001 for each comparison), and inductions of COX-2 mRNA and protein were consistent with these findings. IL-1beta-mediated stimulation of PGE2 was fully blocked in the presence of a nonselective COX inhibitor (indomethacin) or a selective COX-2 inhibitor (NS-398). IL-1beta treatment elicited from group I (normal) and group III (cancer-associated) fibroblasts, respectively, approximately 2- and 3-fold inductions of COX-2 transcriptional activity and approximately 1.4- and 1.7-fold inductions of COX-2 promoter activity. This modestly greater COX-2 transcription rate could not alone account for the dramatically higher levels of COX-2 mRNA and protein and PGE2 in cancer-associated compared with normal fibroblasts. However, incubation of fibroblasts with PGE2 after IL-1beta stimulation prolonged COX-2 mRNA half-life from approximately 1 to 9 h. Our results strengthen the evidence that fibroblasts and other mesenchymal cells are the source of COX-2 expression in normal and premalignant colorectal tissue. Group II fibroblasts from normal segments of colons that harbored synchronous remote advanced neoplasms behaved like group III fibroblasts from advanced neoplasms and not group I fibroblasts from normal colons. We hypothesize that the effects of modestly greater COX-2 transcription in groups II-IV fibroblasts yield corresponding modest increases in PGE2 synthesis whose effects are progressively amplified through robust stabilization of COX-2 mRNA.

Cells, Cultured↗
PubMed2002

Constitutive production of parathyroid hormone-related protein (PTHrP) by fibroblasts derived from normal and pathological human breast tissue.

Parathyroid hormone-related protein (PTHrP) is expressed in the mammary gland and appears to be critical to the morphogenesis of this structure. PTHrP production in the breast is generally attributed to epithelial cells. Because the stromal component of the breast produces factors implicated in proliferation and differentiation of mammary epithelial tissue and tumors, the aim of this study was to investigate the PTHrP expression by mammary fibroblasts from breast cancer tumors and normal breast. PTHrP antibodies labeled intralobular fibroblasts in normal breast and stromal fibroblasts that surround tumor cells. PTHrP was constitutively produced by the cultured mammary fibroblasts, independent of serum stimulation. Normal (15.83 +/- 1.72 fmol/10(6) cells) and pathological breast fibroblasts (19.87 +/- 5.76) secreted similar amounts of PTHrP. PTH/PTHrP receptor mRNA was detected by RT-PCR in all the samples tested. Fibroblasts from normal breast were both PTH and PTHrP-cAMP responsive (453 +/- 133% and 513 +/- 133%, respectively, from basal stimulation), whereas pathological breast fibroblasts were minimally PTHrP-cAMP responsive (183 +/- 36%). The production of other fibroblastic factors implicated in tumor growth and invasiveness was also examined. Interleukin-6 (IL-6), tumor necrosis factor-alpha (INF-alpha), and pro-matrix metalloproteinase (MMP)-1 were not affected by the status of the tissue. In contrast, increased levels of pro-MMP-2 were produced in fibroblasts that originated from pathological (290 +/- 62 ng/10(6) cells) samples compared with those from normal donors (125 +/- 41 ng/10(6) cells). PTHrP production was correlated with TNF-alpha and pro-MMP-2 production. However, inhibition with specific neutralizing antibodies against TNF-alpha or PTHrP, or with a PTHrP antagonist, showed that these factors did not regulate each other. In conclusion, breast fibroblasts are constitutive PTHrP-producing cells with the potential for autocrine signaling through the PTH/PTHrP receptor.

Antibodies↗
PubMedDate not supplied

Scleroderma fibroblasts constitutively express the long pentraxin PTX3.

OBJECTIVE: PTX3 is a secreted molecule which consists of a C-terminal domain similar to classical pentraxins (e.g. C-reactive protein) and of an unrelated N-terminal domain. Unlike the classical pentraxins, PTX3 is expressed in response to IL-1beta and TNF-alpha but not to IL-6. The present study was designed to investigate the expression of PTX3 in normal and scleroderma fibroblasts. METHODS: Normal and SSc fibroblasts were cultured in the presence and absence of inflammatory cytokines. PTX3 mRNA expression in fibroblasts was evaluated by Northern analysis. PTX3 protein levels in fibroblast culture medium were estimated by ELISA. RESULTS: Normal fibroblasts were induced to express high levels of P7X3 mRNA by IL-1beta and TNF-alpha but not by other cytokines or growth factors. Scleroderma fibroblasts, unlike normal fibroblasts, constitutively expressed high levels of PTX3 in the absence of deliberate stimulation. The constitutive expression of PTX3 in SSc fibroblasts was not modified by anti-TNF-alpha antibodies or IL-1 receptor antagonist. In contrast, IFN-gamma and TGF-beta inhibited the constitutive but not the stimulated expression of PTX3 in SSc fibroblasts. CONCLUSIONS: PTX3 is a main feature of activated scleroderma fibroblasts.

C-Reactive Protein↗
PubMed2005

[Effect of prostaglandin E2 and the tumor necrosis factor-alpha on second messenger of lung fibroblast].

OBJECTIVE: To Study the effect of prostaglandin E(2) (PGE(2)) combined with tumor necrosis factor-alpha (TNF-alpha) on the second messenger of mouse lung fibroblast in order to research new target point for cytokine to treatment of pneumoconiosis. METHODS: The lung fibroblasts of breed mouse were primarily cultured. 10 ng/ml TNF-alpha and 10 ng/ml TNF-alpha combined with PGE(2) of different doses were added to culture medium of fibroblasts. The gamma value and CPM value were respectively measured for second messenger of cAMP, cGMP and IP(3) of fibroblast by immune-radio assay at different observation time. RESULTS: When treated with 10 ng/ml TNF-alpha + 1,000 pg/ml PGE(2) at 120 s time points, the CPM value of IP(3) of fibroblast was the maximal value [(76.33 +/- 7.10) CPM]; when cAMP/cGMP ratio declined to 4.29 at 24 h time point, the effect of fibroblast proliferation was the strongest; when 10 ng/ml TNF-alpha + 500 pg/ml PGE(2), and 2 000 pg/ml PGE(2), the CPM value of IP(3) of fibroblast were 27.00 +/- 3.00 and 61.00 +/- 2.65 respectively at 120 s time point, cAMP/cGMP value were 3.50 and 9.83 respectively at 24 h, the effect on fibroblast proliferation were obviously lower. CONCLUSION: Certain dose of PGE(2) could raise the ratio of cAMP/cGMP of fibroblast, and antagonize the proliferation effect of TNF-alpha on fibroblast.

Animals↗
PubMed1992

Enhanced induction of a 72 kDa heat shock protein in cultured retroocular fibroblasts.

Fibroblasts in the retroocular connective tissue appear to play a central role in the pathogenesis of Graves' ophthalmopathy (GO). We hypothesize that specific attributes, differentiating normal from GO retroocular fibroblasts, may render the latter more vulnerable to the ongoing immunopathological process in GO. We investigated whether GO fibroblasts differ from normal fibroblasts with respect to their sensitivity and capacity to express the inducible 72 kDa heat shock protein (HSP) in response to stressful environmental stimuli. Cultured retroocular fibroblasts derived from patients with GO and normal individuals were exposed to various changes in the culture medium that may simulate conditions in the affected retroocular space in vivo. HSP 72 reactivity was determined using sodium dodecyl sulfate polyacrylamide-gel electrophoresis of cellular extracts, followed by immunoblotting with a mouse monoclonal anti-HSP 72 antibody and quantitative scanning densitometry. In addition, indirect immunofluorescence was performed on parallel fibroblast monolayers. Following exposure to heat and acidic pH, deprivation from nutrients, and high monolayer density, GO fibroblasts expressed HSP 72 with significantly greater sensitivity and in significantly higher abundance than did normal fibroblasts. These results demonstrate that changes in the physiological environment induce HSP 72 expression in cultured fibroblasts. The enhanced sensitivity and capacity of GO retroocular fibroblasts to express the inducible HSP 72 in response to stressful stimuli may play a role in the autoimmune process affecting the retroocular space in GO.

Autoimmune Diseases↗
PubMed1986

[Effect of a medium with a low serum content on protein synthesis and secretion and RNA and DNA synthesis in a skin fibroblast culture from patients with rheumatoid arthritis and systemic scleroderma].

Synthesis and secretion of protein, as well as synthesis of RNA and DNA by skin fibroblasts of patients with systemic sclerodermia (SSD) and rheumatoid arthritis (RA) upon prolonged culturing of fibroblasts in the medium with low (0.5-1%) serum content differ markedly in their direction and intensity from protein, RNA and DNA synthesis by skin fibroblasts of healthy donors (HD) and by fetal fibroblasts. It has been found that skin fibroblasts of patients with RA and SSD, as well as those of HD, secrete 75-80% of protein synthesized by fibroblasts de novo upon their culturing in DMEM medium with 1% human serum. Under the same conditions, on days 2-5 of culturing, RNA synthesis in the fibroblasts of patients with RA and SSD was increased 3-4-fold, while DNA synthesis was increased 2-3-fold. Collagenolytic and caseinolytic activity in the culture medium of skin fibroblasts from HD and patients with RA reached maximal levels on days 3-5. High protein secretion was observed in DMEM serum-free medium in the presence of vitamin mixture upon culturing skin fibroblasts of patients with SSD. The results obtained show that skin fibroblasts from HD differ in their functional activity from those of patients with rheumatic disorders. It might be suggested, therefore, that the mechanism of protein secretion plays an important role in the maintenance of constant intracellular protein levels in resting cells.

Arthritis, Rheumatoid↗
PubMed1989

DNA synthesis in prolyl 4-hydroxylase positive fibroblasts in situ in synovial tissue. An autoradiography-immunoperoxidase double labeling study.

DNA synthesis in prolyl 4-hydroxylase (PH; EC 1.14.11.2) positive fibroblasts in situ in synovial tissue was studied using an autoradiography-avidin-biotin-peroxidase complex (ABC) double labeling. Fibroblasts in monolayer culture and in situ in synovial tissue were PH positive, whereas freshly isolated peripheral blood lymphocytes, monocytes, dendritic cells and granulocytes were PH negative. In rheumatoid arthritis (RA) 37 +/- 3 (22-56)% of all DNA synthesizing cells in situ were PH containing fibroblasts, whereas all DNA synthesizing cells in patients with meniscus lesion were PH positive. In both conditions, more than half of the self-replicating fibroblasts were located in the lining cell layer. This is probably not an artifact caused by insufficient penetration of 3H-thymidine because most of the DNA synthesizing lymphocytes were deep down in the synovial stroma. In RA 51 +/- 8 (17-88) PH positive fibroblasts in the S phase of the cell cycle were observed/3 mm2 synovial tissue, whereas the corresponding figure in meniscus patients was only 1 +/- 1 (0-5) (p less than 0.01). This suggests that the local fibroblasts in RA are activated, probably as a result of various fibroblast growth factors produced locally as a result of the inflammatory synovitis. In RA however, less than 1% of all local fibroblasts were self-replicating in situ, whereas labeling indices over 5% were not uncommon in RA synovial fibroblast cultures. This finding suggests that uncontrolled fibroblast proliferation is regulated in vivo by negative feedback mechanisms.

Arthritis, Rheumatoid↗
PubMed1985

[Protein, RNA and DNA synthesis in skin fibroblast cultures from healthy donors and patients with rheumatic diseases].

Synthesis of protein, RNA and DNA was studied in skin fibroblast cultures of healthy donors and patients with systemic scleroderma (SSD) and in those with rheumatoid arthritis (RA) with the use of 14C-protein hydrolyzate, 14C-uridine and 14C-thymidine, respectively. A study was also made of the stimulation of 14C-proline incorporation in protein fibroblasts upon addition to serum-free media of 5% bovine embryonic serum. The stability of RNA in fibroblasts was tested. It was shown that the rate of protein synthesis was 11 times higher in fibroblasts of RA patients and 6 times higher in those of SSD patients as compared to the rate of protein synthesis in fibroblasts of normal subjects. The rate of DNA synthesis in skin fibroblasts of RA patients was 15 times higher and in those of SSD patients 4 times higher than normal. In both RA and SSD patients, the synthesis of short-labeled RNA was 2-3 times higher than normal. The addition of embryonic serum increased 2-3 times the incorporation of 14C-proline in protein skin fibroblasts of SSD patients. It was found that all RNA in skin fibroblasts was represented by long-living molecules and that 30-40% of short-labeled RNA in skin fibroblasts of healthy donors and SSD patients underwent degradation within 1-2 hours. The data obtained indicate that fibroblasts of the two pathologies under study are characterized by considerable differences in the synthesis of DNA and the activity of the protein-synthesizing system.

Arthritis, Rheumatoid↗
PubMed1989

Fibroblast matrix and surface components that mediate cell-to-cell interaction with lymphocytes.

The interaction between lymphocytes and fibroblasts in vitro has been examined using a quantitative ELISA assay to measure the binding of T and B cells to monolayer cultures of human dermal fibroblasts. This was carried out on microtiter culture plates, using an anti-Thy-1 monoclonal antibody, to determine the attachment of murine T lymphocytes and an affinity-purified polyclonal anti-IgM antibody to measure B cell binding. Both types of lymphocyte were found to adhere strongly to intact human fibroblasts, and also had high levels of attachment to purified fibroblast plasma membranes and extracts of the fibroblast extracellular matrix. Attachment, particularly of B lymphocytes, also took place onto plastic surfaces coated with fibronectin, but not to collagens or to intact fibroblasts that had been fixed with a low concentration of paraformaldehyde. Lymphocyte binding to fibroblasts was partially prevented by a monoclonal antibody against fibroblast MHC class II antigens, but not against the class I membrane complex, or by polyclonal antiserum to the cell surface mannose 6-phosphate receptor. In addition, although both lymphocyte types were able to adhere to fibro-nectin, the presence of antibody against fibronectin or the synthetic peptide Arg-Gly-Asp-Ser, had no effect on their attachment to fibroblasts. Thus, lymphocyte adhesion may occur by fibronectin, but other types of interactions with fibroblasts also appear to take place.

Animals↗
PubMed1985

Stimulation of normal rat bone marrow fibroblast proliferation by sera from leukemic Fischer rats.

Fibroblast hyperplasia and accumulation of fibrous material in the bone marrow of patients with idiopathic (primary) or secondary myelofibrosis (MF) is believed to result from a reaction by marrow fibroblasts to an altered marrow microenvironment, the alteration being potentiated by abnormal hemic cells. We investigated the hypothesis that humoral factors might contribute to fibroblast overgrowth in MF by using an animal model, aged Fischer rats, where MF frequently occurs with leukemia. Sera from leukemic rats and leukemic cell conditioned media (CM) were assayed for enhancement of normal rat marrow fibroblast proliferation in a culture system where fibroblasts form discrete, adherent colonies. Our results demonstrated that: leukemic sera induced a 170% increase in total fibroblast colony numbers and a 325% increase in colonies containing more than 80 cells, stimulation of fibroblast growth was leukemia related since sera from rats with transplanted leukemia enhanced marrow fibroblast proliferation, leukemic cell CM did not contain a growth factor for marrow fibroblasts, sera from leukemic rats and 2-mercaptoethanol were additive in enhancing marrow fibroblast proliferation and probably act by different mechanisms, and leukemic rat sera was less effective as a colony-stimulating factor than normal rat sera, a condition mimicked when leukemic and normal spleen CM were compared. This is the first time that a serum component has been implicated in the pathogenesis of MF; our work may contribute to understanding the mechanism involved when MF occurs as a complication of leukemia.

Animals↗
PubMed1981

Interferon-induced resistance of fibroblasts to cytolysis mediated by natural killer cells: specificity and mechanism.

Interferons (IF) have been shown to have a complex regulatory effect on cell-mediated cytotoxicity. In this paper, we described our analysis of the protective effect of IF on human fibroblast target cells. Fibroblasts that have been preincubated with IF are not lysed by natural killer (NK) cells. This IF-induced protection is not observed in antibody-dependent cell-mediated cytotoxicity nor in complement-dependent lysis. Both types of leukocyte IF, viral and immune, and fibroblast IF protect fibroblasts from NK cell-mediated lysis. Electrophoretically purified viral type leukocyte IF also induces protection. Experiments of absorption of NK cells on target cell monolayers and analysis of cytotoxicity at the single-cell level in semisolid medium demonstrate that NK cells bind to both IF-treated and untreated fibroblasts but lyse only the latter. The cytotoxic ability of NK cells is inactivated after interaction with fibroblasts but not with IF-treated fibroblasts. Unlabeled normal fibroblasts, but not IF-treated fibroblasts, compete for the cytotoxicity mediated by NK cells on 51Cr-labeled target fibroblasts. Based on these findings, we propose a possible interpretation of the paradoxic existence of cells that can kill normal cells in the organism and still function in cell-mediated immunity to protect against virus-infected or tumor cells.

Absorption↗
PubMed1994

[Proliferation of pulmonary fibroblasts obtained from rats with bleomycin-induced pulmonary fibrosis and effects of rat rIL-1 alpha on this proliferation].

To clarify the mechanism of pulmonary fibrosis, the growth rate of fibroblasts obtained from bleomycin (BLM)-treated rats was compared with that of fibroblasts obtained from control rats. Proliferation of fibroblasts obtained from rats at 4 days after BLM instillation (BRF) was significantly more rapid than that of fibroblasts obtained from rats at 4 days after saline instillation (CRF), as assessed by cell counting and 3H-TdR incorporation. CRF and BRF were separated by the method of discontinuous Percoll gradients. Three fibroblast fractions of specific gravity (S.G.), S.G. < 1.036, 1.036 < or = S.G. < 1.050, 1.050 < or = S.G. < 1.062, were obtained. Percentages of the densest fibroblast fraction were 46.2 +/- 5.2 in BRF and 6.4 +/- 2.8 in CRF. The densest fibroblasts in BRF proliferated more rapidly than the least dense fibroblasts. Rat rIL-1 alpha suppressed the proliferation of CRF and BRF, dose dependently. These results suggest that an increase in the densest fibroblasts in the lung might correlate with BLM-induced pulmonary fibrosis and that IL-1 alpha suppresses the proliferation of pulmonary fibroblasts.

Animals↗
PubMed1995

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

The nontumorigenic HeLa x skin fibroblast hybrid cell line, CGL1, can be induced to re-express HeLa tumor-associated cell surface antigen, p75-IAP (intestinal alkaline phosphatase), with resulting neoplastic transformation, by exposure to gamma radiation. This has allowed the human hybrid system to be developed into a quantitative in vitro model for radiation-induced neoplastic transformation of human cells. Recently, several gamma-ray-induced IAP-expressing mutants (GIMs) of the nontumorigenic HeLa x skin fibroblast hybrid CGL1 were isolated and all were tumorigenic when injected subcutaneously into nude mice (Mendonca et al., Cancer Res. 51, 4455-4462, 1991). Control cell lines which were negative for p75-IAP (CONs) were also isolated from irradiated populations, and none were found to be tumorigenic. We have now begun to investigate the molecular basis of radiation-induced neoplastic transformation in this system by studying the potential genetic linkage between p75/IAP expression, tumorigenicity and damage to a putative tumor suppressor locus on fibroblast chromosome 11. Previous analysis of rare spontaneous segregants has indicated that this locus is involved in the regulation of tumorigenicity and in the expression of the HeLa tumor-associated cell surface marker intestinal alkaline phosphatase (p75-IAP) in this system. Therefore, analysis by restriction fragment length polymorphism and chromosome painting have been performed for chromosome 11, and for chromosome 13 as a control, for the p75/IAP-positive GIM and p75/IAP-negative CON cell lines. We report that in five of eight of the GIMs large-scale damage to the fibroblast chromosome 11's is evident (four GIMs have lost one complete copy of a fibroblast chromosome 11 and one GIM has both copies of fibroblast chromosome 11 heavily damaged). None of the CONs, however (0/5), have lost a complete copy of either fibroblast chromosome 11. No large-scale damage to the control chromosome 13's was detected in the GIMs or CONs. The data further suggest that both copies of fibroblast chromosome 11 contain an active locus and that radiation-induced loss of either fibroblast chromosome 11 will result in neoplastic transformation in this system. We conclude that it is the loss of a putative tumor suppressor locus on fibroblast chromosome 11 which is responsible at least in part for radiation-induced neoplastic transformation of these human hybrid cells.

Cell Transformation, Neoplastic↗
PubMed1995

Increased proliferation of bone marrow-derived fibroblasts in primitive hypertrophic osteoarthropathy with severe myelofibrosis.

Pachydermoperiostosis or primary hypertrophic osteoarthropathy (HOA) is a rare congenital growth disorder of connective tissue. We report a case of severe myelofibrosis in a patient with HOA. When cultured in vitro, patient bone marrow-derived fibroblasts displayed a high proliferative potential with a shortened doubling time (24 hours v 36 to 48 hours for normal fibroblasts). The role of platelet-derived growth factor (PDGF), previously implicated in the pathogenesis of secondary acquired myelofibrosis, was studied. HOA fibroblasts expressed an increased number of PDGF-BB binding sites (300,000 sites/cell v 200,000 sites/cell for normal fibroblasts) without any modification of affinity. The increased expression of PDGF-R beta appeared to result from an accelerated rate of PDGF-R beta resynthesis with normal kinetics of endocytosis. As a consequence, a several-fold increase of PDGF-R beta tyrosine kinase activity was observed. No autocrine mechanism of growth was suspected as neither spontaneous PDGF-R beta autophosphorylation nor mitogenic activity in HOA fibroblast-conditioned medium was detected. Patient serum and platelet lysate were less potent than controls in inducing [3H]thymidine incorporation into HOA fibroblasts. This was inconsistent with a paracrine mechanism of growth. In vitro, human serum or PDGF-BB were not more mitogenic for HOA than normal fibroblasts. High levels of cyclin D1, a putative oncogene, were detected in serum-deprived HOA fibroblasts. Cyclin D1 overexpression could be implicated in the accelerated growth of these cells. Our results suggest that the mechanism of fibroblastic proliferation observed in this case of myelofibrosis might differ from those reported in other acquired myeloproliferative syndromes and could be associated with an intrinsic abnormality of HOA fibroblast growth.

Anemia↗
PubMed1994

Antigen-presenting capacity of rheumatoid synovial fibroblasts.

In normal, healthy joints, synovial fibroblasts do not express major histocompatibility complex (MHC) class II molecules. However, in inflamed joints of rheumatoid arthritis (RA) patients, synovial fibroblasts show an abundant expression of MHC class II. Does this increase in expression have functional consequences for antigen presentation to T cells? To date, the precise role of synovial fibroblasts in antigen presentation has not been documented. Here, we show by three different examples that cultured synovial fibroblasts with interferon-gamma (IFN-gamma)-induced MHC class II expression are capable of processing soluble protein for presentation to CD4+ T cells. First, the antigen-presenting cell (APC) function of synovial fibroblasts was studied in an autologous model. From synovial tissue of a RA patient both a fibroblast cell line and a tetanus toxoid (TT)-specific CD4+ T-cell line were generated. A dose-dependent TT response was observed only when TT was presented by IFN-gamma-pretreated synovial fibroblasts. As more direct evidence for MHC class II-restricted antigen presentation, the response of a Mycobacterium tuberculosis-specific CD4+ T-cell clone isolated from rheumatoid synovial fluid was demonstrated in the presence of synovial fibroblasts. The response was DR4Dw4-restricted and could be inhibited by monoclonal antibody (mAb) to HLA-DR. In addition, the lymphokine secretion pattern of the synovial T-cell clone did not differ qualitatively upon antigen-specific stimulation using peripheral blood mononuclear cells (PBMC) or synovial fibroblasts as APC. In order to provide evidence for intracellular antigen processing we next examined the response of a M. leprae-specific T-cell clone with known epitope specificity. Our data suggest that synovial fibroblasts are not passive bystanders, but can become active participants in the development and maintenance of chronic inflammation.

Antigen-Presenting Cells↗
PubMed1994

Fibroblast function in psoriatic arthritis. I. Alteration of cell kinetics and growth factor responses.

OBJECTIVE: To explore the potential role of fibroblasts in the pathogenesis of psoriasis and its related arthritis. Specifically, we analyzed the cell cycle of psoriatic fibroblasts obtained from skin and synovium by flow cytometry, and we also studied their response to several growth factors. METHODS: Fibroblast cultures were established from normal and psoriatic skin, uninvolved and involved, and synovium. NIH-3T3 cells were also used as indicator cells in some of the experiments. Fibroblasts DNA cell cycle analysis was performed by flow cytometry, and the data was analyzed by using the "Cytologic DNA applications software version 2." In addition, fibroblasts were stimulated with growth factors including epidermal growth factor, transforming growth factor-beta, and platelet derived growth factor. RESULTS: A significant increase of S and G2-M phase values in confluent cultures of psoriatic fibroblasts in both skin and synovium compared to normal fibroblasts was found. Psoriatic fibroblasts also exhibited a greater proliferative response to growth factors compared to normal fibroblasts. CONCLUSION: Data obtained clearly showed a significant intrinsic in vitro alteration in skin and synovium fibroblasts from patients with psoriasis.

3T3 Cells↗