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G Gabbiani

Publications and source records attributed to G Gabbiani.

At least 55 records · Page 3Linked to original sources
PubMed1999

Interferon-gamma inhibits experimental renal fibrosis.

UNLABELLED: Interferon-gamma inhibits experimental renal fibrosis. BACKGROUND: Recent evidence has implicated myofibroblasts as a cell type responsible for the laying down of extracellular matrix components during fibrosis in a number of organs. In this study, we examined the capacity of interferon-gamma (IFN-gamma) to inhibit the activation of fibroblasts to the myofibroblastic phenotype and hence reduce the extent of renal scarring in the rat subtotal nephrectomy (SNx) model using a novel method of intrarenal delivery. METHODS: Rats were divided into four groups: sham, SNx (group 1), SNx + drug vehicle (group 2) and SNx + IFN-gamma (400 units/day; group 3) for 30 days. Rats were sacrificed on days 15, 30, 45, and 90 following SNx. RESULTS: Clinical data showed a marked reduction in proteinuria in the group treated with IFN-gamma (161 vs. 280 mg/24 hr by day 45, P < 0.01) and a preservation of the creatinine clearance (1.16 vs. 0. 84 ml/min by day 45, P < 0.05) when compared to the SNx or SNx + vehicle groups throughout the time course. Immunohistochemical staining for alpha-smooth muscle actin (alpha-SMA) revealed a reduction in myofibroblastic cell types (6.5 +/- 3.1% glomerular alpha-SMA in group 3 compared with 14.8 +/- 4.2% glomerular alpha-SMA in group 2, P < 0.05, 3.8 +/- 1.4% tubulointerstitial alpha-SMA in group 3 compared with 8.8 +/- 2.0% tubulointerstitial alpha-SMA in group 2 on day 45, P < 0.05). There was also a reduction in immunostaining for collagens III and IV in the IFN-gamma-treated group. Scoring for both glomerulosclerosis and tubulointerstitial fibrosis in the IFN-gamma group (group 3) was lower than the other two operated groups. CONCLUSIONS: We conclude that IFN-gamma, administered at a dose of 400 units/day, has a strong inhibitory effect on myofibroblasts and that as a possible result of this action, renal fibrosis is reduced and renal function is preserved in the rat SNx model. The IFN-gamma renoprotective effect lasted only for the extent of its administration and subsided when discontinued.

Actins↗
PubMed1999

Augmented expression of inducible NO synthase in vascular smooth muscle cells during aging is associated with enhanced NF-kappaB activation.

Vascular smooth muscle cells (SMCs) are important targets for endothelium-derived nitric oxide (NO), but this production is attenuated in injured and diseased arteries and during aging. However, SMCs can produce NO themselves by expressing an inducible form of NO synthase (iNOS) under inflammatory conditions and in the repair process after arterial injury. We examined iNOS expression in SMCs derived from the aortic media of newborn, young adult, and old rats. Our results show that SMCs from newborn rats cannot produce significant amounts of NO on stimulation with interferon-gamma plus lipopolysaccharide or interleukin-1beta. In contrast, SMCs from old rats exhibit markedly enhanced iNOS activity. The difference in iNOS activity between the newborn and the old SMCs was closely correlated with levels of iNOS protein, mRNA, and gene promoter activity. Similarly, intercellular adhesion molecule-1 (ICAM-1) was also expressed more abundantly in the old than in the newborn SMCs in response to cytokines. Both iNOS and ICAM-1 are transcriptionally regulated by nuclear factor kappaB (NF-kappaB). Our data demonstrate an intense transactivation of NF-kappaB in old SMCs on tumor necrosis factor-alpha stimulation but only a weak one in newborn SMCs. The difference in the NF-kappaB activation could be explained by a much faster and more extensive IkappaBalpha degradation in old than in newborn SMCs. These data indicate that the capability to respond to proinflammatory stimuli by activating NF-kappaB differs between SMCs at different stages of development. This results in differential capability to express NF-kappaB-dependent genes such as iNOS and ICAM-1, which could have implications for host defense and the pathogenesis of vascular diseases.

Aging↗
PubMed1999

Retinoic acid regulates arterial smooth muscle cell proliferation and phenotypic features in vivo and in vitro through an RARalpha-dependent signaling pathway.

We have recently shown that all-trans retinoic acid (tRA) modulates arterial smooth muscle cell (SMC) morphologic features and biochemical composition in vitro. It has been proposed that different SMC phenotypes coexist in arteries, which may be retrieved in culture: hence, a differential action of tRA on distinct SMC subsets is conceivable. We have examined the effect of tRA on SMC proliferation, migration, plasminogen activator activity, and alpha-smooth muscle actin expression in 2 phenotypically different rat SMC populations, cultured respectively from the normal aortic media and from the intimal thickening (IT) after endothelial injury. tRA inhibited proliferation and increased migration and tissue-type plasminogen activator activity in both SMC populations, but decreased alpha-smooth muscle actin only in SMC cultured from the IT. The action of tRA is mediated by 2 families of nuclear receptors, RAR and RXR, each containing 3 isoforms, alpha, beta, and gamma. RAR and RAR-alpha agonists, but not RXR agonists, inhibited SMC proliferation in both cell populations and alpha-smooth muscle actin expression only in IT SMC. When administered intraperitoneally to balloon-injured rats, tRA and RAR-alpha agonists reduced the intimal hyperplasia in the carotid artery. Our results show that tRA and synthetic retinoids can affect the proliferation, migration, and differentiation of SMC in vitro. Furthermore, retinoids are able to reduce the IT induced by endothelial injury in vivo.

Animals↗
PubMed1999

Effect of hyaluronic acid on migration, proliferation and alpha-smooth muscle actin expression by cultured rat and human fibroblasts.

Hyaluronic acid represents a major constituent of the extracellular matrix and influences numerous cellular processes. We have studied the effect of this molecule on migration, proliferation and alpha-smooth muscle actin expression by rat and human cultured fibroblasts. Depending on the tissue origin of the fibroblasts, hyaluronic acid induced, inhibited or did not affect fibroblast migration, whereas proliferation was never affected. Alpha-smooth muscle actin expression was inhibited by hyaluronic acid in all types of human and rat fibroblasts studied, but the inhibition was stronger and dose dependent for subcutaneous fibroblasts and for those recovered from Dupuytren's nodules. This property of inhibiting alpha-smooth muscle actin expression makes hyaluronic acid a good candidate to exert antifibrotic activity.

Actins↗
PubMed1998

The fibronectin domain ED-A is crucial for myofibroblastic phenotype induction by transforming growth factor-beta1.

Transforming growth factor-beta1 (TGFbeta1), a major promoter of myofibroblast differentiation, induces alpha-smooth muscle (sn) actin, modulates the expression of adhesive receptors, and enhances the synthesis of extracellular matrix (ECM) molecules including ED-A fibronectin (FN), an isoform de novo expressed during wound healing and fibrotic changes. We report here that ED-A FN deposition precedes alpha-SM actin expression by fibroblasts during granulation tissue evolution in vivo and after TGFbeta1 stimulation in vitro. Moreover, there is a correlation between in vitro expression of alpha-SM actin and ED-A FN in different fibroblastic populations. Seeding fibroblasts on ED-A FN does not elicit per se alpha-SM actin expression; however, incubation of fibroblasts with the anti-ED-A monoclonal antibody IST-9 specifically blocks the TGFbeta1-triggered enhancement of alpha-SM actin and collagen type I, but not that of plasminogen activator inhibitor-1 mRNA. Interestingly, the same inhibiting action is exerted by the soluble recombinant domain ED-A, but neither of these inhibitory agents alter FN matrix assembly. Our findings indicate that ED-A-containing polymerized FN is necessary for the induction of the myofibroblastic phenotype by TGFbeta1 and identify a hitherto unknown mechanism of cytokine-determined gene stimulation based on the generation of an ECM-derived permissive outside in signaling, under the control of the cytokine itself.

Actins↗
PubMed1998

Plasminogen activator expression in rat arterial smooth muscle cells depends on their phenotype and is modulated by cytokines.

Cultured rat aortic smooth muscle cells (SMCs) exhibit at least 2 phenotypic variants: (1) a spindle-shaped phenotype, obtained from normal adult media, and (2) an epithelioid phenotype, obtained from intimal thickening 15 days after endothelial injury. Both phenotypes can be cloned from each location, with normal media yielding a majority of spindle-shaped clones and intimal thickening yielding a majority of epithelioid clones. These findings suggest that intimal thickening develops essentially from a subpopulation of medial SMCs exhibiting epithelioid features in vitro. Using zymographic and Northern blot analyses, we have studied plasminogen activator (PA) expression by these SMCs. Our results show that epithelioid SMCs, cultured as whole SMC populations or as clones, display higher PA activity than do spindle-shaped SMCs, irrespective of their origin. This is mainly due to differences in the expression of tissue PA and, to a lesser extent, urokinase PA and is accompanied by a decrease in PA inhibitor 1. Tissue PA activity is increased by basic fibroblast growth factor and platelet-derived growth factor-BB, particularly in epithelioid SMCs. Taken together, these results indicate that SMCs are heterogeneous with respect to their proteolytic profile, at least as far as the PA system is concerned. Proteolytic activity of the different SMC populations is modulated by cytokines that play a role in intimal thickening. Our results are in agreement with the suggestion that epithelioid SMCs are mainly responsible for intimal thickening.

Animals↗
PubMed1998

Multiple isoform recovery (MIR)-PCR: a simple method for the isolation of related mRNA isoforms.

We present a rapid and efficient method for the detection of related transcripts with different expression levels. This approach combines the rapid amplification of cDNA ends (RACE) method with a cDNA subtractive technique. The strategy is based on successive subtractions of prevalent isoforms resulting in enrichment of less expressed transcripts. For each subtraction, a biotinylated primer specific for the prevalent isoform is hybridized on the total cDNA and the hybrid is retained on a streptavidin affinity column. The unbound cDNA serves as a template for subsequent isoform identification. To illustrate its application we describe the isolation of three new actin cDNA isoforms in the freshwater planarian Dugesia (S) polychroa.

Actins↗
PubMed1998

Rat fibroblasts cultured from various organs exhibit differences in alpha-smooth muscle actin expression, cytoskeletal pattern, and adhesive structure organization.

In vivo, alpha-smooth muscle actin (SMA) is expressed de novo and temporarily by fibroblastic cells during wound healing and correlates particularly with wound contraction. In culture, the presence of varying proportions of cells expressing and not expressing this actin isoform (alpha-SMA-positive and alpha-SMA-negative cells) is characteristic of fibroblastic populations from different tissues. It is possible that mechanisms controlling the expression of actin isoforms, and thus modulating cytoskeleton-related functions, play a major role in the organization of cell shape and motility. We have compared the cell shape as well as the cytoskeleton and focal contact organization in alpha-SMA-positive and alpha-SMA-negative rat fibroblasts from various organs (i.e., skeletal muscle, dermis, subcutaneous tissue, and lung). Within each category, i.e., alpha-SMA-positive or alpha-SMA-negative fibroblasts, no significant morphological differences were seen among populations derived from different tissues. In contrast, alpha-SMA-positive and alpha-SMA-negative fibroblasts were significantly different, independently of their origin: alpha-SMA-positive cells had larger average areas, higher numbers of narrow extensions at the edges, larger focal adhesions with the substratum, and a more important network of cellular fibronectin than alpha-SMA-negative cells. Thus, alpha-SMA-positive and alpha-SMA-negative variants naturally present in fibroblastic populations exhibit important phenotypic differences probably associated with distinct functional activities.

Actins↗
PubMed1998

The expression of alpha 2 beta 1 integrin and alpha smooth muscle actin in fibroblasts grown on collagen.

The maturation of connective tissue involves the organization of collagen fibres by resident fibroblasts. Fibroblast attachment to collagen has been demonstrated to involve cell surface receptors, integrins of the beta 1 family. Integrins are associated with cytoplasmic actin of microfilaments either directly or through focal adhesions. The major actin isoform of fibroblast microfilaments is beta actin and to a lesser extent alpha smooth muscle (alpha SM) actin. Cultured human dermal fibroblasts derived from adult dermis, newborn foreskin or keloid scar were grown on either uncoated or collagen-coated surfaces. The expression and synthesis of both alpha 2 beta 1 integrin and alpha SM actin were followed by immunohistology and immunoprecipitation. Fibroblasts on uncoated surfaces expressed little alpha 2 beta 1 integrin on their surface, while 20 per cent of them demonstrated alpha SM actin within microfilaments. Fibroblasts grown on a collagen-coated surface minimally expressed alpha SM actin in microfilament structures and a majority of the cells were positive for alpha 2 beta 1 integrin on their membranes. Using [35S]-methionine incorporation and immunoprecipitation, it was shown that fibroblasts grown in uncoated dishes synthesized more alpha SM actin than fibroblasts grown on collagen-coated dishes. In contrast, fibroblasts grown on collagen coated dishes synthesized more alpha 2 beta 1 integrin compared to the same cells grown on uncoated dishes. Fibroblasts maintained on a type I collagen upregulate the expression and synthesis of alpha 2 beta 1 integrin, and downregulate the expression and synthesis of alpha SM actin.

Actins↗
PubMed1998

Actin isoforms in amphioxus Branchiostoma lanceolatum.

Actin is a highly conserved cytoskeletal protein that is ubiquitous in all eukaryotes. Little is known about actin expression in amphioxus, the closest living relative of the vertebrates. In the present study, involving Western blotting and indirect immunofluorescence, we report the characterization and localization of various actin isoforms in amphioxus (Branchiostoma lanceolatum) tissues. Three antibodies against vertebrate actins were used: a polyclonal antibody recognizing beta-cytoplasmic actin (anti-beta actin), a monoclonal antibody against sarcomeric actins (anti-alphaSR-1), and a monoclonal antibody specific for alpha-smooth actin (anti-alphaSM-1). Western blot analysis of amphioxus extracts immunodecorated with these antibodies showed a 43-kDa-positive band co-migrating with respective controls. The amphioxus isoactin expression patterns recognized by these antibodies were similar to those of vertebrates, i.e., anti-beta actin showed positive staining mainly in non-muscle cells, anti-alphaSR-1 labelled dorsolateral myotomal muscles, and anti-alphaSM-1 stained ventral muscles. These results demonstrate that at least two muscle actins are present in amphioxus, suggesting that muscle actin gene duplication events began before vertebrate divergence from the amphioxus lineage.

Actins↗
PubMed1998

[Myofibroblasts and retinal fibrovascular membranes].

INTRODUCTION: Epiretinal tissue proliferations occurring during the evolution of ischemic microangiopathies or preretinal diseases are tough to cause retinal detachment by traction mechanisms. Cellular migration/proliferations and finally contraction are tough to be the pathogenic element. Myofibroblasts are contractile cells having features intermediate between those of the fibroblasts and smooth muscle. We conducted a study to explore whether such cells are present in preretinal membranes. MATERIALS AND METHODS: 8 membranes, preelevated during vitrectomy for proliferative vitreoretinopathy or diabetic proliferative vitreoretinopathy, were analysed with immunostaining technique searching for alpha-actine smooth muscle, desmine, which are specific markers for myofibroblasts and TGF-beta1, that is considered as the mean factor promoting the transformation of fibroblasts into myofibroblasts. RESULTS: All the histological preparation showed abundant staining with antibody against alpha-actine smooth muscle, desmine and TGF-beta1. CONCLUSIONS: Myofibroblasts are one of the major cellular element of preretinal membranes. They are scattered throughout the membrane and seem to account for their contractile properties.

Actins↗
PubMed1998

Blood pressure-independent cardiac hypertrophy induced by locally activated renin-angiotensin system.

Cardiac hypertrophy is frequent in chronic hypertension. The renin-angiotensin system, via its effector angiotensin II (Ang II), regulates blood pressure and participates in sustaining hypertension. In addition, a growing body of evidence indicates that Ang II acts also as a growth factor. However, it is still a matter of debate whether the trophic effect of Ang II can trigger cardiac hypertrophy in the absence of elevated blood pressure. To address this question, transgenic mice overexpressing the rat angiotensinogen gene, specifically in the heart, were generated to increase the local activity of the renin-angiotensin system and therefore Ang II production. These mice develop myocardial hypertrophy without signs of fibrosis independently from the presence of hypertension, demonstrating that local Ang II production is important in mediating the hypertrophic response in vivo.

Angiotensin II↗
PubMed1998

Early granulocyte-macrophage colony-stimulating factor expression by alveolar inflammatory cells during bleomycin-induced rat lung fibrosis.

Topical administration of granulocyte-macrophage colony-stimulating factor (GM-CSF) into the subcutaneous tissue or in the pulmonary alveoli of the rat induces a fibrotic reaction characterized by the presence of alpha-smooth muscle actin-rich myofibroblasts, suggesting that GM-CSF plays a role in the development of fibrotic changes. A high level of expression of GM-CSF also has been demonstrated in epidermal cells during human atopic dermatitis. It is accepted that transforming growth factor beta1 (TGF-beta1) plays a key role in the modulation from fibroblast into myofibroblast, although it is not known how TGF-beta1 activity is stimulated. Up until now, no evidence of early GM-CSF expression during development of fibrosis has been reported. Herein we have studied, using RT-competitive PCR, the expression of GM-CSF mRNA during the early steps of pulmonary fibrosis development after intra-alveolar instillation of bleomycin, a well-established experimental model of this lesion. GM-CSF mRNA was already increased in the total lung at 6 hours and maximal at 12 hours after bleomycin instillation and returned to basal levels at 24 hours. This was followed by an increase of TGF-beta1 and TGF-beta receptor type II (but not of types I and III) mRNAs. Analyses of macrophages and polymorphonuclear neutrophils isolated by bronchoalveolar lavage 12 hours after bleomycin instillation indicated that they were responsible, at least in part, for the accumulation of GM-CSF mRNA. Our results show for the first time that GM-CSF is expressed, very early and temporarily, by inflammatory cells accumulating in the alveolus after bleomycin administration and before the appearance of TGF-beta1. Moreover, we have shown that GM-CSF induces the expression of TGF-beta1 mRNA by alveolar macrophages. Our data support the possibility that GM-CSF participates in the initial steps of the chain of events leading to fibrosis, perhaps through a stimulation of TGF-beta1 production.

Actins↗
PubMed1997

Apoptosis during wound healing, fibrocontractive diseases and vascular wall injury.

Following injury, tissue repair involves inflammation, granulation tissue formation and scar constitution. Granulation tissue develops from the connective tissue surrounding the damaged or missing area and contains mainly small vessels, inflammatory cells, fibroblasts and myofibroblasts. As the wound closes and evolves into a scar, there is a striking decrease in cellularity, including disappearance of typical myofibroblasts. The question arises as to what process is responsible for granulation tissue cell disappearance. Our results (in cutaneous wounds) and results of other laboratories (particularly in lungs and kidney) suggest that apoptosis is the mechanism responsible for the evolution of granulation tissue into a scar. During excessive scarring (hypertrophic scar or fibrosis), it is conceivable that the process of apoptosis cannot take place. After experimental endothelial injury in an artery, accumulation of smooth muscle cells participates in the formation of intimal thickening. Apoptotic features have been observed in cells of intimal thickening and also within human atherosclerotic plaques. In the case of atherosclerosis, apoptosis could be detrimental: since smooth muscle cells participate in plaque stability, apoptosis could lead to weakening and rupture of the plaque. These results underline the fact that both increased cell survival or excessive cell death can be associated with pathological disorders. Specific therapies devised to enhance or decrease the susceptibility of individual cell types to apoptosis development could modify the evolution of a variety of human diseases.

Animals↗