Modulate Fibroblast Proliferation Research Articles

Mini review on collagens in normal skin and pathological scars: current understanding and future perspective.

Pathological scar tissues are characterized by the presence of overabundant collagens whose structure and organization are also different from those in unwounded skin. This causes scar tissues to lose some functions performed by normal skin, and currently, there are no effective measures to prevent scar formation. Inflammation has been shown to modulate fibroblast proliferation, differentiation, and function, hence collagen production and organization. In this minireview, we provide an overview of the current understanding of collagen, specifically collagen type I and III which are main collagens in skin, structure and fibre formation and highlight their differences between normal skin and pathological scars. We discuss the role that cytokines play in modulating fibroblast function. We also identify some potential research directions which could help to further our understanding of the complex and dynamic wound healing and scar formation process.

Laminin α5 modulates fibroblast proliferation in epidural fibrosis through the PI3K/AKT/mTOR signaling pathway

Lumbar laminectomy is commonly deemed as the most valid surgery for a series of lumbar illnesses, such as lumbar disc herniation, which could lead to spinal canal stenosis. However, epidural fibrosis is one of the most common complications that limits the application of lumbar laminectomy, which is mainly caused by proliferation of local fibroblasts. Laminins are glycoproteins that consist of α, β and γ chains, which serve a crucial role in biological cell behaviors, such as adhesion, differentiation, migration and proliferation, especially the isoform with the fifth α chain-laminin α5. The PI3K/AKT/mTOR signaling pathway was demonstrated to be associated with various biological functions in cells. The aim of the present study was to explore whether laminin α5 is an important factor in epidural fibrosis by modulating the proliferation of fibroblasts through the activation of PI3K/AKT/mTOR signaling pathway. In the animal model, the results of the hematoxylin-eosin staining, cell counting, Masson's trichrome staining and immunohistochemical staining showed laminin α5 to be positively associated with epidural fibrosis. Furthermore, to verify the assumption that laminin α5 could modulate fibroblast proliferation through the PI3K/AKT/mTOR signal pathway, fibroblasts were transfected with laminin α5-small interfering (si)RNA. The results of western blotting (proliferating cell nuclear antigen and cyclin D1), the Cell Counting Kit-8 and EdU incorporation assays indicated that the proliferative level of fibroblasts decreased, and the expression of phosphorylated (p)-focal adhesion kinase 1, p-AKT and p-mTOR was reduced. Subsequently, laminin α5 was overexpressed and the change in cell proliferation and expression of associated proteins contrasted with that observed in siRNA. The results demonstrated that laminin α5 could interfere the activation of the PI3K/AKT/mTOR signaling pathway. Finally, the inhibition of the PI3K/AKT/mTOR signaling pathway by LY294002 resulted in decreased fibroblast proliferation. In conclusion, laminin α5 could modulate fibroblast proliferation in epidural fibrosis through the PI3K/AKT/mTOR signaling pathway.

Aberrant Caveolin-1-Mediated Smad Signaling and Proliferation Identified by Analysis of Adenine 474 Deletion Mutation (c.474delA) in Patient Fibroblasts: A New Perspective in the Mechanism of Pulmonary Hypertension.

A heterozygous Caveolin-1 c.474delA mutation has been identified in a family with heritable pulmonary arterial hypertension (PAH). This frameshift mutation leads to caveolin-1 protein that contains all known functional domains but has a change only in the final 20 amino acids of the C terminus. Here we studied how this mutation alters caveolin-1 function using patient-derived fibroblasts. Transmission electron microscopy showed that fibroblasts carrying the c.474delA mutation formed typical caveolae. Expression of mutated caveolin-1 in caveolin-1-null mouse fibroblasts failed to induce formation of caveolae due to retention of the mutated protein in the endoplasmic reticulum. However, co-expression of wild type caveolin-1 with mutated caveolin-1 restored the ability to form caveolae. Importantly, fibroblasts carrying the mutation showed 2-fold increase in proliferation rate associated with hyper-phosphorylation of Smad1/5/8. This mutation impaired the anti-proliferative function of caveolin-1. Inhibition of type I TGFβ receptors ALK1/2/3/6 responsible for phosphorylation of Smad1/5/8 reduced the hyper-proliferation seen in c.474delA fibroblasts. These results demonstrate the critical role of the final 20 amino acids of caveolin-1 in modulating fibroblast proliferation through dampening Smad signaling, and suggest that augmented Smad signaling and fibroblast hyper-proliferation are contributing factors in the pathogenesis of PAH in patients with caveolin-1 c.474delA mutation.

Aberrant caveolin-1-mediated Smad signaling and proliferation identified by analysis of adenine 474 deletion mutation (c.474delA) in patient fibroblasts: a new perspective on the mechanism of pulmonary hypertension.

A heterozygous caveolin-1 c.474delA mutation has been identified in a family with heritable pulmonary arterial hypertension (PAH). This frameshift mutation leads to a caveolin-1 protein that contains all known functional domains but has a change in only the final 20 amino acids of the C-terminus. Here we studied how this mutation alters caveolin-1 function, using patient-derived fibroblasts. Transmission electron microscopy showed that fibroblasts carrying the c.474delA mutation form typical caveolae. Expression of mutated caveolin-1 in caveolin-1-null mouse fibroblasts failed to induce formation of caveolae due to retention of the mutated protein in the endoplasmic reticulum. However, coexpression of wild-type caveolin-1 with mutated caveolin-1 restored the ability to form caveolae. Importantly, fibroblasts carrying the mutation showed twofold increase in proliferation rate associated with hyperphosphorylation of Smad1/5/8. This mutation impaired the antiproliferative function of caveolin-1. Inhibition of type I TGFβ receptors ALK1/2/3/6 responsible for phosphorylation of Smad1/5/8 reduced the hyperproliferation seen in c.474delA fibroblasts. These results demonstrate the critical role of the final 20 amino acids of caveolin-1 in modulating fibroblast proliferation by dampening Smad signaling and suggest that augmented Smad signaling and fibroblast hyperproliferation are contributing factors in the pathogenesis of PAH in patients with caveolin-1 c.474delA mutation.

Contractile function recovery in severely injured gastrocnemius muscle of rats treated with either oleic or linoleic acid.

What is the central question of this study? Oleic and linoleic acids modulate fibroblast proliferation and myogenic differentiation in vitro. However, their in vivo effects on muscle regeneration have not yet been examined. We investigated the effects of either oleic or linoleic acid on a well-established model of muscle regeneration after severe laceration. What is the main finding and its importance? We found that linoleic acid increases fibrous tissue deposition and impairs muscle regeneration and recovery of contractile function, whereas oleic acid has the opposite effects in severely injured gastrocnemius muscle, suggesting that linoleic acid has a harmful effect and oleic acid a potential therapeutic effect on muscle regeneration. Oleic and linoleic acids control fibroblast proliferation and myogenic differentiation in vitro; however, there was no study in skeletal muscle in vivo. The aim of this study was to evaluate the effects of either oleic or linoleic acid on the fibrous tissue content (collagen deposition) of muscle and recovery of contractile function in rat gastrocnemius muscle after being severely injured by laceration. Rats were supplemented with either oleic or linoleic acid for 4weeks after laceration [0.44 g(kg body weight)-1 day-1 ]. Muscle injury led to an increase in oleic-to-stearic acid and palmitoleic-to-palmitic acid ratios, suggesting an increase in Δ9 desaturase activity. Increased fibrous tissue deposition and reduced isotonic and tetanic specific forces and resistance to fatigue were observed in the injured muscle. Supplementation with linoleic acid increased the content of eicosadienoic (20:2, n-6) and arachidonic (20:4, n-6) acids, reduced muscle mass and fibre cross-sectional areas, increased fibrous tissue deposition and further reduced the isotonic and tetanic specific forces and resistance to fatigue induced by laceration. Supplementation with oleic acid increased the content of docosahexaenoic acid (22:6, n-3) and abolished the increase in fibrous tissue area and the decrease in isotonic and tetanic specific forces and resistance to fatigue induced by muscle injury. We concluded that supplementation with linoleic acid impairs muscle regeneration and increases fibrous tissue deposition, resulting in impaired recovery of contractile function. Oleic acid supplementation reduced fibrous tissue deposition and improved recovery of contractile function, attenuating the tissue damage caused by muscle injury.

Effect of ceria on the organization and bio-ability of anatase fullerene-like crystals

The presence of Ce atoms induce the formation of fullerene-like structures and increase the oxygen storage capacity of the anatase. It was demonstrated that such special effects can be exploited to modulate fibroblast proliferation.

17 Cardiomyocytes Influence Fibroblast Proliferation and α-Smooth Muscle Actin Expression via the Secretion of Paracrine Mediators

IntroductionCardiac fibroblasts are known to modulate cardiomyocyte structure and function through soluble mediators. While less studied, the ability of cardiomyocytes to influence fibroblast properties, such as proliferation and activation, is...

Role of keratinocytes in wound contraction: an impact assessment using a model of collagen matrix populated with fibroblasts

BACKGROUND: The possible participation of keratinocytes in wound remodeling has been widely studied. This study investigated the impact of keratinocytes in wound contraction. METHODS: Murine type I collagen gels populated by human fibroblasts and seeded with human keratinocytes on the surface to form a dermo-epidermal equivalent were used as the study group. Collagen gels populated by only fibroblasts were used as the control group. The criteria for the preparation and storage of gels were similar for both groups. RESULTS: An evident and statistically significant increase in gel contraction was observed in samples populated by keratinocytes compared to the control group. CONCLUSIONS: These results suggest that keratinocytes not only modulate fibroblast proliferation but also play an active role in wound contraction per se. Further research on the mechanisms involved in the communication pathways between cells and between cells and the matrix shall be assessed from the perspective of keratinocyte participation in wound healing and pathologic scarring.

Mitogenic Activity of S100A9 (MRP-14)

S100A9 is a calcium binding protein found in high amounts in granulocytes and monocytes. We have shown that S100A9 stimulated the proliferation of fibroblasts, but its mechanism remains unknown. In this report, S100A9 is shown to be mitogenic and to stimulate fibroblast proliferation without other growth factors in the serum. Although an S100A8/S100A9 heteropolymer inhibited the growth of fibroblasts by chelating zinc ions, these ions had no effect on the growth-stimulating activity of S100A9. The effects of serum and S100A9 on fibroblast growth were additive, and S100A9 stimulated the growth without serum. Furthermore, S100A9 stimulated the incorporation of bromodeoxyuridine in fibroblasts. However, the effect of S100A9 on the activation of extracellular signal regulated protein kinases (ERK) was small. These results suggest that S100A9 is involved in the regulation of inflammatory processes by modulating fibroblast proliferation.

Interferon-gamma upregulates the stromelysin-1 gene expression by human skin fibroblasts in culture.

The equilibrium between deposition and degradation of extracellular matrix(ECM) is essential to normal tissue development and repair of wound or inflammatory responses. It has recently become apparent that several cytokines and growth factors are capable of modulating fibroblast proliferation and biosynthetic activity. To understand the role of these factors in connective tissue regulation, we examined the effect of interferon-gamma (IFN-gamma) on stromelysin-1 gene expression in cultured human dermal fibroblasts. The steady-state levels of stromelysin-1 mRNA were increased in IFN-gamma treated cultured dermal fibroblasts. In the CAT assay, the stromelysin-1 promoter activity was increased 2.8-fold compared with untreated control. Therefore IFN-gamma stimulates the stromelysin-1 promoter activity, resulting in transcriptional enhancement of gene expression. Transforming growth factor-beta (TGF-beta) showed the antagonistic action to the effects of IFN-gamma in cultured dermal fibroblasts. Furthermore, gel mobility shift assays demonstrated enhanced AP-1 binding activities in nuclear extracts from cells incubated with IFN-gamma. These data suggest that IFN-gamma is an up-regulator and TGF-beta is a down regulator on the stromelysin-1 gene expression, respectively, and the AP-1 binding site may be necessary for gene response.

Chloride conduction of nasal fibroblasts in polyposis patients with cystic fibrosis and in patients without cystic fibrosis. Relevance for the ENT physician

Cystic fibrosis (CF) is a complex systemic disease that has pathological alterations in the upper airways, including the recurrent formation of nasal polyps. Although the fibroblast is the predominant cell type in nasal stroma and nasal polyps, little is known about the electrophysiological properties of nasal fibroblasts. We investigated whether fibroblasts possess a cAMP-regulated chloride conductance which is impaired in patients with CF. Thus far the few studies concerning conductance in fibroblasts have been performed on skin fibroblasts using indirect methods and have yielded conflicting results. Therefore we studied chloride conductance in fused nasal fibroblasts by employing conventional microelectrodes. We have demonstrated that a cAMP-regulated chloride conductance is present in fibroblasts. However, this chloride conductance cannot be activated in fibroblasts from CF-patients. Thus, we present direct evidence that the impairment of the cAMP-regulated chloride conductance in CF is not confined to epithelial cells but also affects the fibroblast. We discuss how this conductance might modulate fibroblast proliferation to produce polyp formation.

Effects of Particles on Fibroblast Proliferation and Bone Resorption In Vitro

An in vitro study was conducted to determine the ability of particle challenged human peripheral monocytes to modulate fibroblast proliferation and bone resorption. The effects of commercially pure titanium, titanium-aluminum-vanadium, and ultrahigh molecular weight polyethylene wear debris, either fabricated or retrieved from patients with failed total hip arthroplasties, were examined as a function of the composition, size, and dose of particles. In vitro generated particles were selected to be matched closely in particle size distribution to that found in vivo. Dosages were controlled by standardizing the ratio of particle surface area to mean monocyte surface area. The results support the hypothesis that, in vitro, challenge of monocytes by particulate wear debris results in a biphasic dose response. For the metal particles, fibrogenesis was observed over the range of 1x to 10x surface area ratio (the surface area of particles to the surface area of cells), although for metallic and polyethylene particles, saturated doses of 10x surface area ratio were required to stimulate bone resorption. In addition, metallic particles were able to stimulate fibrogenesis at doses at which simulated and retrieved polyethylene were ineffective. Although there may be a nonosteolytic chronically tolerable annual dose of ultrahigh molecular weight polyethylene wear debris corresponding to approximately 1x surface area ratio, lower doses, especially of metallic debris, may produce reactive fibroblast proliferation and fibroplasia that may contribute to implant loosening and failure.

Mechanisms of tubulo-interstitial injury in progressive renal diseases.

A vast amount of evidence, based upon human renal biopsy material, indicates that the presence of tubular atrophy and interstitial fibrosis is a better indicator of outcome of renal function than is the extent of glomerular sclerosis. The pathophysiological basis for this surprising fact has not been adequately addressed. In this review we point out that the systemic hypertension which accompanies most forms of chronic renal disease could impact adversely upon the vasodilated interstitial vascular compartment which, together with a component of primary capillary injury related to the disease process, could cause progressive obliteration of particular capillaries. This would initiate a process of chronic tubular ischaemia ultimately leading to tubular atrophy. Since tubular cells have been shown to produce an array of cytokines and growth factors which modulate fibroblast proliferation, extracellular matrix production and chemo-attracts for infiltrating cells, it is further proposed that it is the tubular injury which initiates the deleterious cascade of events. Tubular injury may be aggravated by the filtration of potentially 'noxious' molecules through the diseased glomerulus and by infiltrating cells. As the vascular bed into which glomerular blood flow empties is progressively obliterated, glomerular function declines and renal failure advances in relation to the degree of tubulo-interstitial fibrosis.

Interferon‐γ reverses the stimulation of collagen but not fibronectin gene expression by transforming growth factor‐β in normal human fibroblasts

Abstract. It has recently become apparent that several cytokines and growth factors are capable of modulating fibroblast proliferation and biosynthetic activity. To understand the role of these factors in connective tissue regulation, we examined the effects of the simultaneous addition of interferon‐γ (IFN‐γ) and transforming growth factor‐β (TGF‐β) on normal human dermal fibroblast collagen and fibronectin production. In addition, in vitro transcription rates and steady‐state mRNA levels for these molecules were determined by nuclear run‐off assays and Northern and dot‐blot hybridization using specific human cDNA probes. Treatment of cultures with TGF‐β caused stimulation of collagen and fibronectin production. Addition of IFN‐γ to the TGF‐γ‐treated cultures abrogated the stimulatory effects of TGF‐γ on collagen production in a dose‐dependent manner and resulted in a net inhibition of collagen production. In contrast, the increase in fibronectin synthesis induced by TGF‐γ was augmented further by IFN‐γ. These changes in collagen and fibronectin production were accompanied by parallel changes in the steady‐state mRNA levels for these proteins. The effects of TGF‐γ plus IFN‐γ on fibronectin gene expression appeared to be mediated entirely by transcriptional mechanisms, whereas the effects on collagen gene expression resulted from a combination of transcriptional and post‐transcriptional events.

Interferon‐γ reverses the stimulation of collagen but not fibronectin gene expression by transforming growth factor‐ β in normal human fibroblasts

It has recently become apparent that several cytokines and growth factors are capable of modulating fibroblast proliferation and biosynthetic activity. To understand the role of these factors in connective tissue regulation, we examined the effects of the simultaneous addition of interferon-gamma (IFN-gamma) and transforming growth factor-beta (TGF-beta) on normal human dermal fibroblast collagen and fibronectin production. In addition, in vitro transcription rates and steady-state mRNA levels for these molecules were determined by nuclear run-off assays and Northern and dot-blot hybridization using specific human cDNA probes. Treatment of cultures with TGF-beta caused stimulation of collagen and fibronectin production. Addition of IFN-gamma to the TGF-beta-treated cultures abrogated the stimulatory effects of TGF-beta on collagen production in a dose-dependent manner and resulted in a net inhibition of collagen production. In contrast, the increase in fibronectin synthesis induced by TGF-beta was augmented further by IFN-gamma. These changes in collagen and fibronectin production were accompanied by parallel changes in the steady-state mRNA levels for these proteins. The effects of TGF-beta plus IFN-gamma on fibronectin gene expression appeared to be mediated entirely by transcriptional mechanisms, whereas the effects on collagen gene expression resulted from a combination of transcriptional and post-transcriptional events.

Stimulation of fibroblast proliferation and prostaglandin production by purified recombinant murine interleukin 1

Recombinant murine interleukin 1 (IL-1) obtained from a clone of Escherichia coli containing an IL-1 expression plasmid was purified to homogeneity using a sequential extraction procedure and gel filtration chromatography. The purified recombinant IL-1 exhibited a pI of approximately 5.2 and a sp act of 6 × 10 6 units/mg. These values are in agreement with those obtained with natural murine IL-1. The purified recombinant IL-1 enhanced the proliferation of human HEL and WI-38 fibroblasts in a serum-free medium. In addition, IL-1 stimulated fibroblast PGE 2 5- to 30-fold over a 24-hr period. The effects of IL-1 on fibroblast activation were obtained with the same concentrations of IL-1 that are effective in the mouse thymocyte assay. These results unequivocally establish the ability of IL-1 to modulate fibroblast proliferation and function.

Ultrastructural and metabolic effects of products of activated mononuclear leucocytes on cultured human dermal fibroblasts

Products of antigen- or mitogen-activated mononuclear leucocytes (ML) are known to modulate fibroblast proliferation and collagen production in vitro. In tissue, ML accumulate at sites of inflammation and are probably involved in the process of fibrosis. We have established that supernatants (SN) of concanavalin A-activated ML increase synthesis of glycosaminoglycan (GAG) in human dermal fibroblast (DF) cultures). When explants of normal human skin were cultured in CMRL 1066 medium supplemented with 15% pooled human serum and containing MLSN the outgrowing DF acquired a distinctive phenotype. In comparison to control DF, the cells treated with MLSN exhibited a marked increase in the number of intracytoplasmic organelles especially dilated cisternae of RER filled with electrondense material, abundant lysosomes, prominent Golgi apparatus and bundles of microfilaments often extending beyond the cell boundaries. Cell surfaces were shaggy and floccular material accumulated in patches along the plasma membranes.