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PubMed1990

Characterization of two types of human oral fibroblast with a potential application to cellular toxicity studies: tooth pulp fibroblasts and buccal mucosa fibroblasts.

Cultures of human oral buccal mucosa fibroblasts and human tooth pulp fibroblasts were established and grown under standard routine conditions. The biological characteristics of cell proliferation, growth pattern, cell morphology and enzyme release (lactate dehydrogenase and acid phosphatase) were studied. Under the same standard in vitro conditions the two cell types demonstrated different growth patterns and different levels of enzyme activity. It is suggested that differences in biological characteristics should be considered when selecting appropriate cells for toxicological studies of dental materials.

Acid Phosphatase↗

PubMed2026

Atherosclerotic plaque fibroblasts derive from adventitial and medial Pdgfra-lineage-positive cells and predominantly maintain fibroblast identity.

AIMS: Fibroblasts are mesenchymal cells in the healthy vascular adventitia. In atherosclerosis, single-cell sequencing datasets suggest fibroblasts are abundant in plaques. However, their identity, origin, and fate during plaque progression remain unclear, which we aim to unravel here. APPROACH AND RESULTS: To robustly define fibroblast identity, origin, and fate, we employed meta-analyses of 54 single-cell RNA sequencing libraries, including murine smooth muscle cell (Myh11) and endothelial cell (EC) (Cdh5) lineage reporter mice with and without atherosclerosis; human control and atherosclerotic arteries; and murine adventitia and atherosclerotic plaques processed separately from low-density lipoprotein (LDL) receptor knockout (Ldlr-/-) mice. These meta-analyses showed that murine and human plaque fibroblast identity was robustly defined by Pdgfra, Pi16, Cygb, and Serpinf1 mRNA. Ninety-five percent of plaque fibroblasts do not derive from the Myh11 lineage, while no Cdh5-lineage-positive cells were present in the fibroblast cluster. We identified five murine arterial fibroblast subsets in atherosclerotic murine aorta: progenitor fibroblasts, matrix fibroblasts, inflammatory fibroblasts, an EC-like fibroblast subset, detected in both adventitia and plaques, and Col5a3+ fibroblasts, unique to the adventitia. We next studied fibroblast identity, origin, and fate using pseudotime analysis and Pdgfra-CreERT2/tdTomato lineage reporter mice (Pdgfra Lin+). Healthy Pdgfra Lin+ reporter mice showed predominant adventitial tdTomato expression, and infrequent medial and intimal Pdgfra Lin+ cells co-expressing MYH11 and PECAM1, respectively. The Pdgfra Lin+ plaque area increased with diet duration. Pdgfra Lin+ cells largely maintain fibroblast identity in the plaque, while <10% co-express SMC markers (MYH11, SM22α), or contribute to ACTA2+ cap cells. ECs gaining mesenchymal markers are transcriptionally distinct from Cdh5-lineage-negative fibroblasts gaining EC markers. Plaque-resident EC-like fibroblasts displayed a mesenchymal-to-endothelial transition transcriptome, which was induced in human primary fibroblasts in vitro by starvation, and dampened or reversed by IL1B, TGFB1, TGFB3, and oxidized LDL. Cross-species integration showed that all murine plaque fibroblasts were conserved in human atherosclerosis, with one additional subset partially resembling murine subsets, and three human-specific subsets. Importantly, human fibroblast subsets differentially correlated to human plaque traits, with EC-like fibroblasts correlating to plaque instability. CONCLUSION: Our results indicate that 95% of plaque-residing fibroblasts are Myh11 Lin- Plaque fibroblasts have a dual origin, predominantly adventitial Pdgfra Lin+ progenitor fibroblasts, with a minor contribution from medial Pdgfra Lin+  Myh11+ SMCs. Most plaque fibroblasts maintain fibroblast identity. Murine plaque fibroblast subsets were conserved in human atherosclerosis. EC-like fibroblasts are linked to human plaque instability. Intervening in progenitor-to-specific fibroblast transitions could present a new avenue to promote plaque stability in atherosclerosis.

Atherosclerosis↗

PubMed1992

Collagen synthesis in fibroblasts from human colon: regulatory aspects and differences with skin fibroblasts.

The purpose of this study was to examine regulation of collagen synthesis in human colon fibroblasts and compare the results from colon fibroblasts with those obtained in fibroblasts from human skin. The effects of interleukin-1 beta, tumour necrosis factor alpha, interferon gamma, transforming growth factor-beta, dexamethasone, and the calcium ionophore A23187 were investigated. All compounds were tested both in the absence and in the presence of fetal calf serum in the culture medium. The process of collagen synthesis in fibroblasts from colon and skin appears to be affected differently by these regulatory compounds. The most pronounced differences were that the relative collagen synthesis increased in dermal fibroblasts and decreased in colon fibroblasts upon addition of serum. In the presence of serum, interleukin-1 beta inhibited collagen synthesis in skin fibroblasts but not in colon fibroblasts. Dexamethasone suppressed the relative collagen synthesis in skin fibroblasts but not in colon fibroblasts. Transforming growth factor-beta stimulated the collagen synthesis in dermal fibroblasts in the presence of serum, but inhibited the process in colon fibroblasts. Because fibroblasts are the primary sources of collagen needed during wound repair, these results may offer (part of) the explanation why wounds in skin and intestine appear to behave differently under certain conditions.

Calcimycin↗

PubMed1983

Bleomycin-induced pulmonary fibrosis in hamsters. An alveolar macrophage product increases fibroblast prostaglandin E2 and cyclic adenosine monophosphate and suppresses fibroblast proliferation and collagen production.

Bleomycin-induced pulmonary fibrosis in hamsters is associated with collagen accumulation that results from increased lung collagen synthesis rates. However, 1-2 wk after intratracheal instillation of bleomycin, lung collagen synthesis rates decline toward control values. To evaluate the potential role of the bronchoalveolar macrophage in regulating lung collagen production, we studied the effects of macrophages from normal and bleomycin-treated hamsters upon fibroblasts in vitro. We observed: (a) Medium from macrophage cultures decreased fibroblast [3H]thymidine incorporation and nondialyzable [3H]hydroxyproline production in a dose-dependent manner. Fibroblast cell counts were decreased in exposed cultures, and fibroblast viability was unchanged. Procollagen prolyl hydroxylation and prolyl-transfer RNA-specific activity were not altered by macrophage medium; this indicates that [3H]hydroxyproline reflects collagen production rate under the experimental conditions. (b) The suppressive effect of macrophage medium was selective for collagen since collagen production decreased more than noncollagen protein production. (c) Medium from bleomycin-treated hamster macrophages suppressed fibroblast proliferation and collagen production to a greater degree than medium from normal hamster macrophages. (d) Fibroblast suppression by macrophage medium was associated with increased fibroblast endogenous prostaglandin E2 production and intracellular cyclic AMP (cAMP). (e) Incubation of fibroblasts with indomethacin before exposure completely inhibited prostaglandin E2 production and increases in cAMP, and eliminated suppression of fibroblast proliferation and collagen production. The macrophage-derived suppressive factor has an apparent molecular weight of 20,000-30,000 and is heat stable. It does not appear to be species restricted since both hamster and human lung fibroblasts are similarly suppressed. It is at least in part preformed in macrophages obtained by lavage, but its production can also be stimulated in vitro. We concluded that alveolar macrophages release a product that stimulates endogenous fibroblast prostaglandin E2 production and cAMP formation with resultant suppression of fibroblast proliferation and collagen production. Enhanced release of suppressive factor by macrophages during a time when lung collagen production is declining in bleomycin-induced pulmonary fibrosis suggests that macrophages may limit collagen accumulation in pulmonary fibrosis.

Animals↗

PubMed1985

Monocyte inhibition of lung fibroblast growth: relationship to fibroblast prostaglandin production and density-defined monocyte subpopulations.

Mononuclear cell fibroblast interactions in the normal human lung are poorly understood. Mononuclear cells can regulate fibroblast function and blood monocytes are known to migrate to the lung and participate in pulmonary inflammation. Thus, to clarify mononuclear cell-fibroblast interactions in the normal lung, we obtained supernatants from adherent monocytes and characterized their effect on the log phase growth of human lung fibroblasts. Monocyte supernatants inhibited fibroblast growth in a dose-dependent fashion. The inhibition was the result of an approximately 16,000 MW soluble factor(s) that was heat stable, trypsin sensitive, and chymotrypsin resistant. Elaboration of the factor(s) required monocyte protein synthesis and was not restricted to a density-defined monocyte subpopulation. The inhibitory capacity of a monocyte supernatant was directly related to its ability to stimulate fibroblast prostaglandin production. Blocking fibroblast prostaglandin production reversed the inhibition of fibroblast growth caused by monocyte supernatants. Thus, monocyte inhibition of fibroblast growth may be mediated by fibroblast prostaglandin production. Recruitment of monocytes to the lung and subsequent monocyte inhibition of fibroblast growth may be important in regulating pulmonary fibrosis.

Cell Division↗

PubMed1990

Re-evaluation of fibroblasts and fibroblast-like cells.

Scanning electron microscopic observations of connective tissue cells show a new aspect of the nature of fibroblasts, and the subsequent broad survey of references makes clear that fibroblasts of many tissues have various features which are regarded as atypical of fibroblasts, and at the same time that various connective tissue cells in different organs have features typical of fibroblasts. Both morphological and functional features of fibroblasts are more or less common to those of fibroblast-like cells, and differences among these cells are quantitative rather than qualitative. Therefore, it is almost impossible to set clear-cut criteria for distinguishing genuine fibroblasts from a large population of fibroblast-like cells. The majority of cells sharing features of fibroblasts, if not all, seem to belong to the same population of cells. They are probably adapted to special functional needs in their own micro-environment that are peculiar to local or pathological or experimental conditions. It is proposed to categorize these cells into subtypes depending on their main functions: 1, fibrogenesis; 2, tissue skeleton or barrier; 3, intercellular communication system; 4, gentle contractile machinery; 5, endocrine activity; and 6, vitamin A-storing. Re-evaluation of fibroblasts and fibroblast-like cells is required to facilitate their better understanding.

Animals↗

PubMed1988

Tumor necrosis factor interacts with interleukin-1 and interferons to inhibit fibroblast proliferation via fibroblast prostaglandin-dependent and -independent mechanisms.

Mononuclear cells are important regulators of fibroblast function. However, the soluble factors mediating these effects and the importance of intercytokine interactions in these regulating events remain poorly understood. We analyzed the effect of recombinant (r) interleukin-1-alpha (IL-1), tumor necrosis factor (TNF), and gamma, alpha, and beta 1 interferons (IFN), alone and in combination, on the proliferation of normal human lung fibroblasts. rIL-1 and rTNF weakly stimulated and the rIFNs inhibited fibroblast proliferation. Importantly, when rTNF was combined with rIL-1 or rIFN, synergistic inhibition of fibroblast proliferation was noted. The inhibition caused by rIFN-gamma plus rTNF was largely independent of fibroblast prostaglandin (PG) production because fibroblast PG levels were unaltered in the presence of these cytokines and blocking fibroblast PG production did not alter their inhibitory effect. In contrast, the inhibition caused by rIL-1 plus rTNF appeared to be at least partially mediated by fibroblast PG production because these cytokines acted synergistically to stimulate fibroblast PG production and blocking fibroblast PG production reversed the inhibition that they caused. These studies demonstrate that TNF can interact with IL-1 or IFN to inhibit the proliferation of normal diploid fibroblasts and that the inhibitory effects of these cytokine combinations are mediated by different mechanisms.

Cell Division↗

PubMed1978

Contribution of human somatomedin activity to the serum growth requirement of human skin fibroblasts and chick embryo fibroblasts in culture.

Although purified human somatomedins and related peptides have been shown to stimulate growth-related processes in cultured human skin fibroblasts, it is unknown whether the serum macromolecules required for the routine growth of human fibroblasts in culture include the somatomedins. To evaluate this question we have obtained sera from five GH-deficient patients before and after GH treatment and have compared these pre- and post-GH sera for their ability to stimulate [3H]thymidine incorporation into DNA and cell multiplication in the patients' own fibroblasts in culture. Subconfluent human fibroblasts demonstrated a dose-dependent increase in thymidine incorporation and cell number when exposed to human sera. Pre- and post-GH sera were equipotent. Partial purification of somatomedin activity by boiling sera at pH 5.5 diminished the level of thymidine incorporation to 10-20% of the level achieved with unboiled sera but did not unmask a difference between pre- and post-GH sera. In contrast to the results in human fibroblasts, boiled post-GH sera were 2- to 5-fold more potent than pre-GH sera in stimulating thymidine incorporation in chick embryo fibroblasts. Mixing experiments failed to demonstrate inhibition of thymidine incorporation by boiled pre-GH sera. Boiled post-GH sera also were twice as active as boiled pre-GH sera in stimulating multiplication of chick embryo fibroblasts and confirmed that thymidine incorporation reflected DNA synthesis. GH added directly to chick embryo fibroblasts did not stimulate thymidine incorporation. We conclude that despite the presence of specific receptors for somatomedin-like peptides on human skin fibroblasts, somatomedins are not a major component of the serum macromolecules required for the routine growth of human fibroblasts in culture. In contrast, in chick embryo fibroblasts, somatomedins do appear to constitute an important part of the serum macromolecules supporting cell growth.

Animals↗

PubMed1988

Heterogeneous proliferative characteristics of human adult lung fibroblast lines and clonally derived fibroblasts from control and fibrotic tissue.

Pulmonary fibrosis results from an altered deposition of collagen within the lung parenchyma. This alteration is likely the result of both increased fibroblast proliferation and abnormalities in fibroblast collagen metabolism. Although the development of pulmonary fibrosis is preceded by inflammatory events in the lung, it is unclear whether altered fibroblast behavior requires continuous exposure to inflammatory mediators or alternatively results from the emergence in the lung of fibroblast populations possessing characteristics such as to explain the abnormalities seen in pulmonary fibrosis. To examine the latter hypothesis, we have established a number of fibroblast cell lines from control (C) lung tissue as well as from tissue from patients with active pulmonary fibrosis (PF), and have examined their in vitro proliferative characteristics. Our data show that PF fibroblasts proliferate significantly faster compared to C fibroblasts under standard culture conditions. We have also examined the in vitro proliferative characteristics of a substantial number of clonally derived fibroblasts. We report that a marked heterogeneity exists in terms of proliferation and also that a small but significant number of fast-growing clones are present in panels of clones derived from fibrotic tissue. These data suggest that there exists in fibrotic tissue, clones of fibroblasts with intrinsic growth characteristics which could in itself explain the increased fibroblast proliferation seen in pulmonary fibrosis. The fibrotic clones may emerge as dominant in the fibrotic lung under conditions of injury and repair likely to favor the expansion of this phenotype.

Adult↗

PubMed1978

Binding of chemotactic collagen-derived peptides to fibroblasts. The relationship to fibroblast chemotaxis.

We previously showed that collagen, alpha-chains, and collagen-derived peptide fragments induce chemotactic migration of human fibroblasts in vitro. We now describe biochemical and immunological evidence showing there are binding sites for collagen peptides on fibroblast membranes.By the use of (14)C-labeled alpha1(I) chain, binding to intact fibroblasts was demonstrated. The process was reversible, and time- and fibroblast concentration-dependent. Scatchard plot analyses of the data obtained for the binding of alpha1(I) suggested that there are congruent with 16 x 10(6) binding sites per fibroblast with an association constant of 1.1 x 10(7)/M for alpha1(I). Dissociation of the bound radioactivity and subsequent chromatographic analysis on agarose A-1.5 m revealed that the alpha1 was unaltered. The binding of (14)C-labeled alpha1 was inhibited by each of the CNBr peptides derived from alpha1 chain of chick skin collagen and CNBr peptide mixtures of various genetic types of collagen chains. Immunofluorescence studies with anti-alpha1 antibody showed that alpha1-treated fibroblasts exhibited strong immunofluorescence. The intensity of fluorescence was markedly diminished by prior absorption of the antibody with alpha1. The alpha1-treated cells stained with preimmune sera did not show significant fluorescence.Dose-response curves of fibroblast chemotaxis induced by alpha1 and the binding of alpha1 by fibroblasts correlate closely. Furthermore, the potency of alpha1-CNBr peptides as chemotactic agents correlates with their ability to inhibit the binding of labeled alpha1(I). These data suggest the hypothesis that collagenderived peptides cause fibroblast chemotactic migration by acting on fibroblast membranes.

Binding Sites↗

PubMed1988

Coculture of mouse IL-3-dependent mast cells with 3T3 fibroblasts stimulates synthesis of globopentaosylceramide (Forssman glycolipid) by fibroblasts and surface expression on both populations.

Mouse IL-3-dependent bone marrow culture-derived mast cells (BMMC) and mouse 3T3 fibroblasts, cultured separately or together, were examined for their cell surface expression and biosynthesis of globopentaosylceramide, a marker of the mouse serosal mast cell. As assessed by flow cytometric analysis, BMMC cultured for up to 7 wk in 50% WEHI 3-conditioned medium containing IL-3 did not bind the B1.1 anti-globopentaosylceramide mAb (six experiments). A total of 10 +/- 4% (mean +/- SD, three experiments) of 3T3 fibroblasts that had reached confluence in medium without IL-3 bound B1.1 antibody and, after an additional approximately 28 days of culture in that medium or in 50% WEHI 3-conditioned medium, 12 +/- 3% (mean +/- SD, five experiments) and 16 +/- 7% (mean +/- SD, three experiments) of the cells, respectively, bound the antibody. After coculture of BMMC and confluent 3T3 fibroblasts for 28 days in 50% WEHI 3-conditioned medium, followed by dispersal and purification of the cells, 92 +/- 18% of the mast cells and 92 +/- 16% (mean +/- SD, seven experiments) of the fibroblasts were B1.1+. Whereas the increase in the expression of the epitope bound by B1.1 antibody on fibroblasts was noted by day 14 of coculture, expression of the epitope on mast cells did not occur until day 21 (three experiments). Biosynthesis of globopentaosylceramide was assessed by intrinsic radiolabeling of each cell population and identification of the extracted neutral glycosphingolipids by TLC and autoradiography. Synthesis of globopentaosylceramide was not detected in extracts of 9 x 10(6) BMMC, 1 x 10(6) confluent 3T3 fibroblasts cultured alone for 28 days, or 9 x 10(6) mast cells purified from 28-day cocultures but was readily detected in extracts of 3 x 10(5) fibroblasts purified from the same cocultures. These findings indicate that BMMC stimulate an increase in the synthesis and expression of globopentaosylceramide on 3T3 fibroblasts and suggest that the subsequent appearance of this neutral glycosphingolipid on the surface of the mast cells is due to its secretion by fibroblasts and adsorption to the mast cell surface. Thus, the interactions between mast cells and fibroblasts during coculture alter the biochemical and Ag phenotypes of both populations.

Animals↗

PubMed1989

Skin fibroblasts from patients with the genetic disorder hereditary hemorrhagic telangiectasia compared with ataxia-telangiectasia fibroblasts in their response to the radiomimetic drug neocarzinostatin.

Skin fibroblasts from patients with hereditary hemorrhagic telangiectasia (HHT) were compared with fibroblasts from patients with the genetic disorder ataxia-telangiectasia (A-T), and with SV40-transformed A-T fibroblasts, regarding their sensitivity to the radiomimetic drug neocarzinostatin (NCS). Whereas A-T fibroblasts were found to be hypersensitive to NCS at low concentrations (as measured by the cellular survival test), as previously reported, the HHT fibroblasts were more resistant to NCS and behaved as normal fibroblasts. The SV40-transformed A-T cells also resembled normal fibroblasts in their response to NCS in the colony formation test. In the DNA synthesis test, HHT strains of fibroblasts did not fully resemble untransformed and SV40-transformed A-T cells that continued to synthesize DNA following NCS treatment, since NCS inhibited DNA synthesis in HHT fibroblasts by 5 to 15%; whereas the same treatment in normal fibroblasts reduced DNA synthesis by 40%.

Antibiotics, Antineoplastic↗

PubMed1985

Cell culture of embryonic chick duodenal cells: preparation of epithelial-fibroblast bilayers and homotypic cultures of fibroblasts and epithelial cells.

A method for the primary cell culture of trypsin-dissociated embryonic chick duodenum is described. Both heterotypic (epithelial cells and fibroblasts together) and homotypic (highly enriched cultures of epithelial cells or fibroblasts alone) cell cultures were established. Dispersed duodenal epithelial cells and fibroblasts grown in 10% fetal bovine serum (FBS) spontaneously aggregated and proliferated as a bilayer of cells with the epithelial cells growing on top of the fibroblasts. Changing the serum supplement to 6% chicken serum (CS) and 4% FBS when the fibroblast monolayer reached confluence resulted in epithelial cell proliferation. Homotypic cultures of epithelial cells and fibroblasts were prepared and analyzed by scanning electron microscopy and transmission electron microscopy. Fibroblasts, isolated by differential adhesion and grown in 10% FBS, did not demonstrate measurable alkaline phosphatase activity. Homotypic epithelial cell cultures, isolated by floating them off the fibroblasts with collagenase, and maintained on collagen in 6% CS/4% FBS, demonstrated higher alkaline phosphatase-specific activity (16.1 +/- 2.3 U/mg protein) compared with epithelial-fibroblast bilayer cell cultures (12.1 +/- 1.3 U/mg protein).

Alkaline Phosphatase↗

PubMed1991

Constitutive expression of a 92-kD gelatinase (type V collagenase) by rheumatoid synovial fibroblasts and its induction in normal human fibroblasts by inflammatory cytokines.

Synovial fibroblasts freshly isolated from the rheumatoid joint are characterized by their marked connective tissue degradative ability. This phenotype includes the ability to secrete large amounts of the matrix-degrading metalloproteinases, collagenase, and stromelysin. We have found that another aspect of this phenotype is the constitutive expression at both protein and mRNA levels of a 92-kD gelatinolytic metalloproteinase, which is not secreted by normal dermal or lung fibroblasts and is immunologically cross-reactive with a type V collagenase expressed by activated macrophages and neutrophils. Expression of this 92-kD metalloproteinase confers upon the fibroblasts the capacity to degrade collagenase- and stromelysin-resistant interstitial elements, such as collagen types IV, V and XI. In contrast to the 92-kD metalloproteinase, a 68-kD gelatinase (type IV collagenase) was expressed by all fibroblast types studied, indicating that its regulation is distinct from that of the 92-kD gelatinase. To identify what cytokines may be important in the induction of the rheumatoid synovial phenotype, including expression of the 92-kD gelatinase, we exposed normal dermal fibroblasts to a number of cytokines including many known or considered likely to be present in rheumatoid synovial fluid and tissue. Although IL-1 beta, tumor necrosis factor-alpha, lymphotoxin, platelet-derived growth factor, and basic fibroblast growth factor were capable of stimulating fibroblasts to secrete collagenase, only tumor necrosis factor-alpha, lymphotoxin, and IL-1 beta were able to induce expression of the 92-kD gelatinase, demonstrating discordant regulation of the two metalloproteinases. Expression of the 68-kD gelatinase was independent of that of the 92-kD gelatinase, as demonstrated at the protein and mRNA levels. Late passage rheumatoid synovial fibroblasts, which no longer constitutively expressed the 92-kD gelatinase, displayed an accentuated response to IL-1 beta when compared to normal dermal fibroblasts. Thus, in addition to IL-1 beta, tumor necrosis factor-alpha or lymphotoxin may contribute to the expression of a specific rheumatoid synovial phenotype in vivo that is associated with progressive matrix destruction.

Arthritis, Rheumatoid↗

PubMed1992

Cytokine regulation of human lung fibroblast hyaluronan (hyaluronic acid) production. Evidence for cytokine-regulated hyaluronan (hyaluronic acid) degradation and human lung fibroblast-derived hyaluronidase.

We characterized the mechanisms by which recombinant (r) tumor necrosis factor (TNF), IFN-gamma, and IL-1, alone and in combination, regulate human lung fibroblast hyaluronic acid (HA) production. Each cytokine stimulated fibroblast HA production. The combination of rTNF and rIFN-gamma resulted in a synergistic increase in the production of high molecular weight HA. This was due to a synergistic increase in hyaluronate synthetase activity and a simultaneous decrease in HA degradation. In contrast, when rTNF and rIL-1 were combined, an additive increase in low molecular weight HA was noted. This was due to a synergistic increase in hyaluronate synthetase activity and a simultaneous increase in HA degradation. Human lung fibroblasts contained a hyaluronidase that, at pH 3.7, depolymerized high molecular weight HA to 10-40 kD end products of digestion. However, hyaluronidase activity did not correlate with fibroblast HA degradation. Instead, HA degradation correlated with fibroblast-HA binding, which was increased by rIL-1 plus rTNF and decreased by rIFN-gamma plus rTNF. Recombinant IL-1 and rTNF weakly stimulated and rIL-1 and rTNF in combination further augmented the levels of CD44 mRNA in lung fibroblasts. In contrast, rIFN-gamma did not significantly alter the levels of CD44 mRNA in unstimulated or rTNF stimulated cells. These studies demonstrate that rIL-1, rTNF, and rIFN-gamma have complex effects on biosynthesis and degradation which alter the quantity and molecular weight of the HA produced by lung fibroblasts. They also show that fibroblast HA degradation is mediated by a previously unrecognized lysosomal-type hyaluronidase whose function may be regulated by altering fibroblast-HA binding. Lastly, they suggest that the CD44 HA receptor may be involved in this process.

Adult↗

PubMed1991

Fibroblast proliferation factors in pulmonary granuloma induced by Trichosporon cutaneum in rabbits: presence of a lymphocyte-derived fibroblast proliferation factor and its functional specificity.

The fibroblast proliferation activity (FPA) in pulmonary granulomatous lesions in rabbits which were exposed once (primary response) or twice (secondary response) to Trichosporon cutaneum was examined using R9ab, a rabbit fibroblast cell line cell, and fibroblasts from the lesions of the primary and secondary responses. The FPA in lung extracts and cell-free culture supernatants of bronchoalveolar lavage cells from the secondary response was more evident than that from the primary response. FPA from the primary response were recovered at about 60, 18, and 4.5 kD and those from secondary response at about 60, 26, 18, and 4.5 kD on Sephadex G-75 gel filtration. Among the FPA, the activity with a molecular weight of 26 kD and a pI of 7.0 was derived from lymphocytes, whereas the other activities were derived from macrophages. The macrophage-derived fibroblast proliferation factors (FPF) enhanced proliferation of fibroblasts from the lesions of both primary and secondary responses, while the lymphocyte-derived FPF enhanced only proliferation from the secondary response. It was further found that lymphocyte-derived FPF could chemotactically attract fibroblasts from the secondary but not the primary response, indicating functional specificity of lymphocyte-derived FPF on fibroblasts in the secondary response. The present results suggest that this lymphokine with fibroblast proliferation and chemotactic activity plays an important role in the granuloma formation in the secondary response to T. cutaneum.

Animals↗

PubMed1990

Gene-expression and release of macrophage-colony stimulating factor in quiescent and proliferating fibroblasts. Effects of serum, fibroblast growth-promoting factors, and IL-1.

Macrophage (M)-CSF induces the proliferation and differentiation of macrophage-precursor cells, and is an important factor in the survival and activation of mature mononuclear phagocytes. Several effector cell populations such as mononuclear phagocytes, endothelial cells, and fibroblasts, have been reported to produce M-CSF at high levels. We investigated the gene-expression and release of M-CSF in fibroblasts. Quiescent, growth-arrested, fibroblasts showed little expression of M-CSF as demonstrated by Northern blot analysis, and little production of M-CSF as measured in the murine bone marrow bioassay. Initiation of proliferation of fibroblasts by fetal bovine serum was followed by an increase in M-CSF transcription and release of the protein. Similarly, single factors (platelet-derived growth factor) or combinations of factors (epidermal growth factor (EGF) + insulin, platelet-derived growth factor + EGF + insulin, or fibroblast growth factor + EGF + insulin) that induced proliferation of the fibroblasts, also induced increased expression of M-CSF. As soon as 1 h after the addition of fetal bovine serum, M-CSF expression was increased, and reached a plateau at 2 to 8 h after induction. IL-1 increased M-CSF expression both in quiescent and proliferating fibroblasts, and induced the expression and release of granulocyte-, and granulocyte/macrophage-CSF. These results show that expression of M-CSF in fibroblasts is not constitutively at a high level, but undergoes regulation depending on the cellular proliferative state and on further activation by acute response proteins such as IL-1.

Animals↗

PubMed1986

A monoclonal antibody to the carboxyterminal domain of procollagen type I visualizes collagen-synthesizing fibroblasts. Detection of an altered fibroblast phenotype in lungs of patients with pulmonary fibrosis.

Excessive collagen deposition plays a critical role in the development of fibrosis, and early or active fibrosis may be more susceptible to therapeutic intervention than later stages of scarring. However, at present there is no simple method for assessing the collagen-synthesizing and secreting activity of fibroblasts in human tissues. Type I procollagen carboxyterminal domains are proteolytically removed during collagen secretion. Thus, antibodies to these domains should stain fibroblasts synthesizing type I collagen but not extracellular collagen fibrils which could mask the signal from the cells. We developed and characterized a monoclonal antibody (Anti-pC) specific for the carboxyterminal propeptide of type I procollagen. To determine the relationship between Anti-pC staining and collagen synthesis, we stained embryonic and adult chicken tendon. Embryonic chick tendon fibroblasts actively synthesizing type I collagen stained heavily with Anti-pC, while quiescent adult tendon fibroblasts did not stain with Anti-pC. Wounded adult tendons developed fibroblasts that stained with Anti-pC at the wound site. Thus, Anti-pC specifically visualized fibroblasts actively synthesizing collagen. Lung biopsies from patients with fibrotic lung disease were stained with Anti-pC. Interstitial and intraalveolar fibroblasts in biopsies from patients with active fibrosis stained intensely with Anti-pC, while normal human lung was unstained. The absence of staining in normal lung supports the hypothesis that fibrosis is associated with an altered collagen-synthesizing phenotype of tissue fibroblasts. Anti-pC may provide a useful clinical tool for assessing fibrogenic activity at sites of tissue injury.

Antibodies, Monoclonal↗