Deposition Of Collagen Type Research Articles

NG2 modulates tight junction integrity in brain microvessels

In this work we addressed the putative role of NG2 in the control of endothelial tight junction (TJ) dismantling, blood-brain barrier (BBB) breakdown, and leukocyte transendothelial migration in inflammatory/degenerative neurological disorders. During CNS development two cell types are known to express the transmembrane proteoglycan NG2, namely oligodendrocyte precursors (OPCs) and microvessel-associated pericytes. In both cell types expression of NG2 is a hallmark of cell activation in response to signalling molecules, mainly growth factors, and via interaction with ECM components. According to our previous results in a chronic model of experimental autoimmune encephalomyelitis (EAE) induced in C57BL/6 mice, NG2-expressing OPCs react to demyelination by an increasing proliferation and this phenomenon seems to be promoted by the interactions of NG2 with naked axons. In this condition, a dismantled TJ-induced microvessel leakage is associated with an increased number of NG2-bearing pericytes and perivascular OPCs. By comparing these data with observations made in naive and EAE NG2 null mice, which show a constitutive alteration of the TJ staining pattern apparently restored during EAE, a key role is disclosed for NG2 in the regulation of the dynamics of TJ components and how these are remodelled during pathological conditions. Our evidence is that loss of NG2 causes a reduced deposition of collagen type IV of the vascular basement membrane and a parallel reduction in collagen type VI microfilaments associated with this membrane. This in turn causes a disruption of the pericyte-endothelial cell interactions that indirectly modulate TJ integrity.

Upregulation of Hic-5 in glomerulosclerosis and its regulation of mesangial cell apoptosis

Glomerulosclerosis is characterized by the loss of glomerular cells by apoptosis and deposition of collagen type I into the normal collagen IV-containing mesangial matrix. We sought to determine the alterations that might contribute to these changes by performing proteomic analysis of rat mesangial cell lysates comparing cells cultured on normal collagen type IV to those grown on abnormal collagen type I surfaces. Subculture on collagen type I was associated with changed expression of several proteins, including a significant upregulation of the paxillin-like LIM protein, hydrogen-peroxide-induced clone 5 (Hic-5), and increased the susceptibility of the cells to apoptosis in response to physiological triggers. When we knocked down Hic-5 (using siRNA), we found mesangial cells grown on collagen type I were protected from apoptosis to the same degree as untreated cells grown on collagen type IV. Further we found that the level of Hic-5 in vivo was almost undetectable in control rats but increased dramatically in the glomerular mesangium of remnant kidneys 90 and 120 days after subtotal nephrectomy. This induction of Hic-5 paralleled the upregulation of mesangial collagen type I expression and glomerular cell apoptosis. Our results suggest that Hic-5 is pivotal in mediating the response of mesangial cells to attachment on abnormal extracellular matrix during glomerular scarring.

Increased expression of collagens, transforming growth factor-β1, and -β3 in gluteal muscle contracture

BackgroudGluteal muscle contracture (GMC) is a multi-factor human chronic fibrotic disease of the gluteal muscle. Fibrotic tissue is characterized by excessive accumulation of collagen in the muscle's extracellular matrix. Transforming growth factor (TGF)-β1 and -β2 are thought to play an important role in fibrogenesis, while TGF-β3 is believed to have an anti-fibrotic function. We hypothesize that the expression of collagen and TGF-βs would be up-regulated in GMC patients.MethodsThe expression of collagen type I, type III and TGF-βs were studied in 23 fibrotic samples and 23 normal/control samples in GMC patients using immunohistochemistry, reverse transcription and polymerase chain reaction (RT-PCR) and western bolt analysis.ResultsCompared to the unaffected adjacent muscle, increased expression of TGF-β1 and -β3 was associated with deposition of collagen type I and type III in the fibrotic muscle of the GMC patients at the mRNA level. Strong up-regulation of these proteins in fibrotic muscle was confirmed by immunohistochemical staining and western blot analysis. TGF-β2 was not up-regulated in relation to GMC.ConclusionThis study confirmed our hypothesis that collagen types I, III, TGF-β1 and TGF-β3 were up-regulated in biopsy specimens obtained from patients with GMC. Complex interaction of TGF-β1 with profibrotic function and TGF-β3 with antifibrotic function may increase synthesis of collagens and thereby significantly contribute to the process of gluteal muscle scarring in patients with GMC.

TORAFIC study protocol: torasemide prolonged release versus furosemide in patients with chronic heart failure

Loop diuretics, such as torasemide and furosemide, are important agents in the treatment of chronic heart failure. Beneficial effects of torasemide immediate-release formulation beyond diuresis have been documented as the ability of this compound to inhibit myocardial synthesis and deposition of collagen type I in patients with chronic heart failure. In addition, torasemide-treated patients, but not furosemide-treated patients, showed decreased serum concentrations of the C-terminal propeptide of procollagen type I, a biochemical marker of myocardial fibrosis. The aim of the TORAFIC study is to test the efficacy of torasemide prolonged-release formulation (PR) in reducing myocardial fibrosis in chronic heart failure in a large, randomized clinical trial. Methods: This prospective, Phase IV, randomized, blinded end point, active-controlled clinical trial will randomize 142 patients with chronic heart failure in New York Heart Association functional class II–IV to 8 months treatment with either torasemide-PR (10–40 mg daily) or furosemide (40–160 mg daily). The primary objective is to test the hypothesis that torasemide-PR is superior to furosemide in reducing myocardial fibrosis. The primary outcome measure is the difference in the change of serum propeptide of procollagen type I concentration from the initial to the final visit between both study groups. Secondary outcome measures include all efficacy variables related to heart failure (signs and symptoms, ECG, echocardiogram and serum levels of N-terminal brain natriuretic propeptide). Secondary safety variables are heart rate, blood pressure, laboratory data, adverse events, cardiovascular events (hospital admission, emergency department visits) and quality of life (Minnesota questionnaire). Discussion: This trial will test whether torasemide-PR possesses antifibrotic properties, which may provide an additional benefit beyond diuresis in patients with chronic heart failure.

Growth hormone substitution in growth hormone-deficient adults: Effects on collagen type I synthesis and skin thickness

Growth hormone stimulates collagen type I synthesis. Collagen type I is a common matrix compound in a large number of connective tissues. The aim of our study was to prove whether a stimulation of collagen type I synthesis might be accompanied by a deposition of collagen type I in the skin (cutis). Twenty growth hormone-deficient hypopituitary patients were included in a randomised, double-blind, placebo controlled, prospective, twelve-month study (eighteen patients assessable at the end of the study). The patients were treated with recombinant human growth hormone 0.25 U/kg/week subdivided in daily subcutaneous injections beginning with half the dosage during the first four weeks. During the first six months half of the patients were treated with placebo. PICP, the indicator of collagen type I synthesis, was increased after six months of therapy when compared to placebo. Skin thickness measured by ultrasound at the forearm and mechanically at the dorsum of the hand with strong compression of the skin both increased significantly following growth hormone substitution. Our data indicate that the stimulation of collagen type I synthesis by growth hormone substitution is followed by a deposition of collagen type I in the skin.

Early expression of monocyte chemoattractant protein-1 correlates with the onset of isoproterenol-induced cardiac fibrosis in rats with distinct angiotensin-converting enzyme polymorphism

Isoproterenol treatment of Brown Norway and Lewis rats (high and low plasma angiotensin-I-converting enzyme activity, respectively) results in similar cardiac hypertrophy but higher cardiac fibrosis in Brown Norway rats. Rats were infused in vivo with isoproterenol for two or 10 days. Cardiac fibrosis and inflammation were evaluated histochemically. We measured the mRNAs of pro-fibrotic factors (transforming growth factor beta(1), endothelin-1) and pro-inflammatory factors (monocyte chemoattractant protein-1). In studies with cardiac fibroblasts incubated with isoproterenol in vitro , we measured cell proliferation, angiotensin-I-converting enzyme and matrix metalloprotease 2 activities and deposition of collagen type I and fibronectin. After treatment with isoproterenol for two days, there were large areas of myocardial injury and numerous inflammatory foci in the left ventricle, these being greater in Brown-Norway than in Lewis rats. After treatment with isoproterenol for 10 days, there were large areas of damage with extensive collagen deposition only in the left ventricle; both strains exhibited this damage which was, however, more severe in Brown-Norway than in Lewis rats. After treatment with isoproterenol for two, but not 10, days, greater amounts of monocyte chemoattractant protein-1 mRNA were found in Brown Norway than in Lewis rats. Cell proliferation, activities of angiotensin-I-converting enzyme and matrix metalloprotease 2, amounts of collagen type I and fibronectin were similar in cardiac fibroblasts from both strains; changes after isoproterenol (10 microM) were also similar in both strains. We conclude that the greater cardiac fibrosis in Brown Norway rats treated with isoproterenol correlates with the early and higher expression of proinflammatory factors.

Mesangial sclerotic change with persistent proteinuria in rats after two consecutive injections of monoclonal antibody 1-22-3.

Irreversible mesangial changes with persistent proteinuria were induced in rats given two consecutive injections 2 weeks apart of a MoAb 1-22-3 to rat mesangial cell. The characteristics of the resulting lesions were investigated and compared with those of the reversible change induced by a single injection. At 24 h after the second injection, mesangiolytic changes similar to those after a single injection were evident. The accumulation of macrophage-like cells in glomeruli observed at 1 week after the first injection was not evident during the experimental period after the second injection. Hypercellularity with the characteristics of intrinsic mesangial cell and increased mesangial matrix were already present 1 week after the second injection. And mesangial sclerotic change progressed up to 6 months. Deposition of collagen type I and type III and accumulation of collagen fibril at the ultrastructural level were evident in rats 6 months after the second injection. Proteinuria started immediately and continued for more than 6 months after the second injection. The mesangial sclerotic change with persistent proteinuria described here is considered to be a better model for investigating the mechanism of chronic progression of human mesangial proliferative glomerulonephritis.

Cartilage-Specific Matrix and Integrin Expression in Three-Dimensional Articular Chondrocyte Cultures Overexpressing Human Interleukin-10

Interleukin (IL)-10 overexpression inhibits joint inflammation, however the effect of high local concentrations of IL-10 on chondrocyte homeostasis remains unclear. The aim of this study was to determine the effects of IL-10 overexpression on cartilage matrix production in three-dimensional (3D) chondrocyte cultures.Human articular chondrocytes were transduced with adenoviral vectors alone (adv/empty) or by vectors either overexpressing enhanced green fluorescence protein (adv/EGFP) or human IL-10 (adv/hIL-10) before their transfer to a 3D culture system. Non-transduced chondrocytes were used as controls. The expression of IL-10 or EGFP was confirmed using ELISA or flow cytometry. Chondrocytes synthesis of collagen types II and I, aggrecan, fibronectin and β1-integrin was determined over a period of 14 days post transduction using flow cytometry or immunohistochemistry.adv/EGFP or adv/IL-10 transduced chondrocytes expressed EGFP or secreted IL-10 detectable over the 2 weeks culture period. No suppression of collagen type II, aggrecan or β1-integrin synthesis by IL-10 overexpression was found and the deposition of collagen type I and fibronectin remained unaffected compared to the controls.IL-10 overexpression does not impair key features of chondrocytes differentiated phenotype (e.g. collagen type II and aggrecan expression) suggesting the potential use of IL-10 for gene therapeutic approaches in the joint.

Isoproterenol-induced impairment of heart function and remodeling are attenuated by the nonpeptide angiotensin-(1-7) analogue AVE 0991

The aim of this study was to evaluate the effects of AVE 0991 (AVE), a nonpeptide compound that mimics Ang-(1-7) actions, on cardiac remodeling. Heart hypertrophy and heart dysfunction were induced by isoproterenol (ISO) (2 mg/kg i.p./day for 7 days) in male Wistar rats. At the end of the 7-day period, the hearts were perfused according to the Langendorff method to evaluate cardiac function. The hearts, atria, and right and left ventricles wet weights were recorded, normalized for body weight and then expressed as muscle mass index (mg/g). In addition, serial sections from left ventricle were stained with hematoxylin–eosin for cell morphometry and with collagen-specific Masson's trichrome for detection of fibrosis. Immunofluorescence-labeling and confocal microscopy were used to investigate the distribution and deposition of collagen types I, III, VI, and fibronectin. AVE reduced the ISO-induced hypertrophy as quantified by myocyte diameter measurements (Control: 10.60 ± 0.08 μm; ISO: 14.60 ± 0.11 μm; ISO + AVE: 11.22 ± 0.08 μm, n = 5). In addition, AVE markedly attenuated the increase of extracellular matrix proteins induced by ISO. AVE treatment also attenuated the decrease in systolic tension and ± d T/d t and exacerbated the vasodilatation induced by ISO. These results show that AVE has a cardioprotective effect on ISO-induced cardiac remodeling.

Paeoniflorin inhibits TGF-β1-mediated collagen production bySchistosoma japonicumsoluble egg antigenin vitro

The main pathological characteristics of hepatic fibrosis in schistosomiasis are the proliferation of hepatic stellate cells (HSCs) and the deposition of collagen type I (Col I) and collagen type III (Col III). Transforming growth factor beta-1 (TGF-beta1) plays an important role in hepatic fibrosis. Paeoniflorin (PAE) has been reported to have immunoregulatory effects; however, the mechanism of its anti-hepatic fibrosis in S. japonicum has not been elucidated. In the present study, we found that mouse peritoneal macrophages (PMphis) stimulated by soluble egg antigen (SEA) of S. japonicum could secrete TGF-beta1, and the TGF-beta1 in the peritoneal macrophage-conditioned medium (PMCM) could induce proliferation of HSCs and secretion of Col I and III. We selected PMCM at 1 : 2 dilution as the optimum PMCM (OPMCM). Then we treated HSCs pre-incubated with OPMCM with PAE, and found that the inhibition of HSC proliferation or Col I and III production were closely correlated with the concentration of PAE. Further investigation found that PAE significantly decreased the Smad3 transcription and phosphorylation in HSCs stimulated by OPMCM. In conclusion, SEA plays a key role in hepatic fibrosis by inducing TGF-beta1 from PMphis. PAE can exert anti-fibrogenic effects by inhibiting HSCs proliferation and down-regulating Smad3 expression and phosphorylation through TGF-beta1 signalling.

Aortic Aneurysm: Search for the Trigger

Over the past two decades, there has been a significant advancement in our understanding of the pathophysiologic processes of abdominal aortic aneurysm formation and growth. By studying specimens of aortic aneurysm wall removed at the time of surgery, numerous investigators have contributed to our understanding of the inflammatory cell infiltrate and the impact of matrix metalloproteinases, especially MMP2, MMP3, and MMP9. 1 McMillan W.D. Pearce W.H. Increased plasma levels of metalloproteinase-9 are associated with abdominal aortic aneurysms. J Vasc Surg. 1999; 29: 122-127 Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar , 2 Tamarins N.A. McMillan W.D. Shively V.P. Pearce W.H. Expression of matrix metalloproteinases and their inhibitors in aneurysms and normal aorta. Surgery. 1997; 122: 264-271 Abstract Full Text PDF PubMed Scopus (263) Google Scholar , 3 Davis V. Persidskaia R. Baca-Regen L. Itoh Y. Nagase H. Persidsky Y. Ghorpade A. Baxter B.T. Matrix metalloproteinase-2 production and its binding to the matrix are increased in abdominal aortic aneurysms. Arterioscler Thromb Vasc Biol. 1998; 18: 1625-1633 Crossref PubMed Scopus (228) Google Scholar , 4 McMillan W.D. Tamarina N.A. Cipollone M. Johnson D.A. Parker M.A. Pearce WH Size matters: the relationship between MMP-9 expression and aortic diameter. Circulation. 1997; 96: 2228-2232 Crossref PubMed Scopus (210) Google Scholar A significant increase in synthesis and deposition of collagen types I and III and the reduction of mature elastin fibers within the aneurysm wall have also been clearly demonstrated. 5 Rizzo R.J. McCarthy W.J. Dixit S.N. Lilly M.P. Shively V.P. Flinn W.R. Yao J.S. Collagen types and matrix protein content in human abdominal aortic aneurysms. J Vasc Surg. 1989; 10: 365-373 Abstract Full Text Full Text PDF PubMed Scopus (239) Google Scholar , 6 Mesh C.L. Baxter B.T. Pearce W.H. Chisholm R.L. McGee G.S. Yao J.S. Collagen and elastin gene expression in aortic aneurysms. Surgery. 1992; 112: 256-261 PubMed Google Scholar

Role of mast cells in wound healing process after glass-fiber composite implant in rats.

Glass-fiber composites are frequently used in dentistry. In order to evaluate their biocompatibility we tested, in an experimental model “in vivo”, their tissue response pointing our attention on presence of mast cells (MCs) and fibrotic process. Sprague Dawley rats were used for the experimental design. The fibers were introduced in a subcutaneous pocket along the middle dorsal line between the two scapulas for 7, 14 or 21 days. At the end of the treatments the skins were excised and then processed for Toluidine Blue, to determine the presence of MCs, and Picrosirius Red staining, to evaluate the presence of fibrotic tissue. Our preliminary results showed and increase of both MC number and deposition of collagen type I, which characterized the fibrotic tissue. So, subsequent aims of our study were to evaluate the role played by MCs in tissue fibrosis and to give a possible explanation regarding the mechanisms that were responsible of biological response observed, through the analyses of some proteins, such as metalloproteinase-2 (MMP-2), its inhibitor (TIMP-2) and transforming growth factor-β (TGF-β). Our data confirmed the involvement of TGF-β, released by MCs, in the disruption of the equilibrium between MMP-2 and TIMP-2 that were implicated in the enhancement of fibrosis. In summary, this study demonstrate that this type of materials induced an inflammatory response at the site of implant and help to clarify what type of mechanism and which proteins are involved in this biological response. Nevertheless, more extensive investigations are in progress to better evaluate the inflammatory process.

Effects of the 1,320-nm Nd:YAG laser on transepidermal water loss, histological changes, and collagen remodeling in skin

The goal of the study is to investigate the effects of the 1,320-nm Nd:YAG laser on skin barrier function and dermal remodeling. Forty-eight female mice were irradiated with the 1,320-nm laser four times at an interval of 1 week (350 micros, 10 mm, single pass 10% overlap). Three doses were used: 18, 22, and 26 J/cm2. Transepidermal water loss (TEWL) was measured at day 0, 1, 7, 21, 30, and 60 after treatments. Skin biopsies were taken to observe the changes in dermal structures and quantify the increases in the number of fibroblasts and collagen content. The dose of 22 J/cm2 induced the lowest ratio of TEWL (0.58+/-0.06, p < 0.05), highest degree of fibroblast proliferation (45.68+/-5.70, p < 0.05), and the most increase in type I collagen fibers (49.8%, p < 0.05) and hydroxyproline content (85.7%, p < 0.05). The dose of 26 J/cm2 caused marked inflammatory infiltration and the most increase in type III collagen fibers. The increase of type I collagen fibers was greater than type III collagen fibers for all three doses (p < 0.05). Our results demonstrated that 22 J/cm2 was most effective in enhancing skin barrier function and stimulating new collagen synthesis. Dermal remodeling after the 1,320-nm laser was mainly through the synthesis and deposition of collagen type I. Inflammatory reactions were in favor of the formation of type III collagen.

Adult stem cell homing and differentiation in vitro on composite fibrin matrix

Strategies to generate differentiated cells from haematopoetic progenitor cells will enhance potential use of adult stem cells for therapeutic transplantation or tissue engineering. Transplantation of undifferentiated stem cells into recipient tissue hinges on the hypothesis of a milieu dependent differentiation and it has been suggested that a clot-equivalent scaffold is crucial for these circulating cells to anchor and multiply. Here a natural scaffold, fibrin along with fibronectin, gelatin and growth factors has been used to induce endothelial progenitor cells and smooth muscle progenitor cells to differentiate into endothelial cells and smooth muscle cells, respectively, from peripheral blood mononuclear cells. Characteristics of endothelial cells have been verified by the detection of mRNA for and immunostaining the cells for von Willebrand factor, uptake of acetylated low-density lipoproteins and measurement of released nitric oxide in the culture medium, as nitrite. The specific molecules that characterized smooth muscle cells were alpha smooth muscle actin and calponin, besides deposition of collagen type I and elastin, onto the culture matrix. The adhesive proteins used for the fabrication of endothelial progenitor cells matrix and smooth muscle progenitor cells matrix were the same, but specific differentiation was brought about by modulating the growth factor composition in the matrix and in the culture medium. Both endothelial and smooth muscle cells were consistently developed from 20 ml of human blood.

Immunohistochemical characterization of epithelial cells implanted in the flap–stroma interface of the cornea

To investigate the expression of extracellular matrix collagens and their relationship to corneal opacities after implantation of epithelial cells in the flap-stroma corneal interface. A corneal flap was made on rabbit eyes, and epithelial cells, mechanically scraped from tissue surrounding the flap, were implanted beneath the flap. The corneas were harvested 1, 3, 7, and 30 days following surgery. Histological and immunohistochemical examinations were performed. The expression and localization of types I, III, and IV collagens and gelatinase A were determined. Slit-lamp examination showed corneal opacity in the area where the epithelial cells were implanted. Histological study revealed clusters of epithelial cells between the flap and stromal interface. One week and 1 month after the implantation, intense immunoreactivity for collagen type IV was detected at the perimeters of the intrastromal epithelial islands, but not in the interface outside the implanted epithelial cells. Weak positive staining for gelatinase A was detected in the implanted epithelial cells and surrounding keratocytes. The heavy deposition of collagen type IV surrounding the implanted epithelial cells indicated that it might be an essential component of the interface haze observed in patients following laser in situ Keratomileusis. Gelatinase A may also play a role in the regulation of stromal remodeling after epithelial ingrowth.

Effects of undernutrition on respiratory mechanics and lung parenchyma remodeling.

Undernutrition thwarts lung structure and function, but there are disagreements about the behavior of lung mechanics in malnourished animals. To clarify this issue, lung and chest wall mechanical properties were subdivided into their resistive, elastic, and viscoelastic properties in nutritionally deprived (ND) rats and correlated with the data gathered from histology (light and electron microscopy and elastic fiber content), and bronchoalveolar lavage fluid analysis (lipid and protein content). Twenty-four Wistar rats were assigned into two groups. In the control (Ctrl) group the animals received food ad libitum. In the ND group, rats received one-third of their usual daily food consumption until they lost 40% of their initial body weight. Lung static elastance, viscoelastic and resistive pressures (normalized by functional residual capacity), and chest wall pressures were higher in the ND group than in the Ctrl group. The ND group exhibited patchy atelectasis, areas of emphysema, interstitial edema, and reduced elastic fiber content. The amount of lipid and protein in bronchoalveolar lavage fluid was significantly reduced in the ND group. Electron microscopy showed 1) type II pneumocytes with a reduction in lamellar body content, multilamellated structures, membrane vesicles, granular debris, and structurally aberrant mitochondria; and 2) diaphragm and intercostals with atrophy, disarrangement of the myofibrils, and deposition of collagen type I fibers. In conclusion, undernutrition led to lung and chest wall mechanical changes that were the result from a balance among the following modifications: 1) distorted structure of diaphragm and intercostals, 2) surfactant content reduction, and 3) decrease in elastic fiber content.

Alterations in myocardial ultrastructure and protein expression after a single injection of isoproterenol.

Immunochemical and electron microscopic characterization of rat myocardium was conducted 2 h and 3 weeks after a single injection of isoproterenol in rats. The relative content of several myospecific proteins (KRP--kinase-related protein, desmin), cytoskeletal proteins (tubulin, vinculin, myosin light chain kinase--MLCK) and extracellular matrix protein fibronectin was determined by immunoblotting. Two hours after injection of 50 mg/kg isoproterenol a destruction of some cardiomyocytes, contracture of myofibrils and mild edema of intercellular space was observed. The content of all the studied proteins except KRP decreased below control levels. This situation sustained 3 weeks after injection and paralleled alterations in cardiomyocyte ultrastructure. Areas of myofibrillar contracture and lysis were noted, glycogen granules were sparse; mitochondria contained arrow-like inclusions that are characteristic for calcium overload, also huge mitochondria contacting each other by specialized intermitochondrial contacts were detected. Clumps of unripe elastic fibers in enlarged intercellular space were combined with increased deposition of collagens type I and III forming areas of fibrosis. The smaller dosage of isoproterenol (10 mg/kg) rendered no significant damage in the acute postinjection period but 3 weeks later it induced the thickening of extracellular matrix around cardiac cells and the increase in KRP and tubulin content by 26 and 32%, correspondingly. MLCK levels remained depressed throughout the experiment. The rise in KRP expression was also observed after the addition of isoproterenol to cultured chicken embryo cardiomyocytes. Obtained results indicate that even a single injection of isoproterenol creates long lasting structural alterations in cardiac muscle accompanied by the increased expression of extracellular matrix proteins and several sarcoplasmic proteins apparently involved in hypertrophic response of cardiomyocytes.

Repeated allergen exposure changes collagen composition in airways of sensitised Brown Norway rats.

Increased or altered collagen deposition in the airway wall is one of the characteristics of airway remodelling in asthma. The mechanisms underlying this increase, and its functional consequences remain to be established further. Representative in vivo animal models might be useful in this respect. In the present study, collagen deposition after prolonged allergen exposure was characterised in the airway wall of Brown Norway rats. Sensitised rats were repeatedly exposed to ovalbumin (OA) or phosphate-buffered saline during 2 and 12 weeks. The deposition of collagen type I, III, IV, V and VI was not altered in animals exposed to OA for 2 weeks. After 12 weeks of OA exposure, more collagen type I was deposited in the inner and outer airway wall and more type V and VI collagen was observed in the outer airway wall. At 12 weeks the number of vessels, identified via type IV collagen staining was not increased, but the total vessel area was. In conclusion, prolonged allergen exposure in sensitised rats is associated with enhanced deposition of type I, V and VI collagens and increased vascularity. This suggests that some aspects of airway remodelling in asthma could be driven by long-term allergen exposure.

Myofibroblasts are responsible for the desmoplastic reaction surrounding human pancreatic carcinomas

Background. The cell type responsible for the desmoplastic reaction surrounding human pancreatic carcinoma is unknown. Hepatic stellate cells, which activate to a myofibroblast-like form, are responsible for collagen deposition in cirrhosis and around hepatocellular carcinomas. Recently, pancreatic stellate cells have been described and implicated in the fibrosis of chronic pancreatitis. We sought to determine whether these cells are responsible for the scirrhous reaction surrounding pancreatic adenocarcinomas. Methods. Archival formalin-fixed, paraffin-embedded pancreatic tissues from 10 patients undergoing pancreaticoduodenectomy for ductal adenocarcinoma and from 2 patients with pancreatic islet cell tumors were examined immunohistochemically for alpha-smooth muscle actin (α-SMA), smooth muscle myosin heavy chain (SMMHC), procollagen I, collagen IV, and endothelial cell markers, von Willebrand factor and cluster of differentiation 31. Results. In non-neoplastic areas, staining for α-SMA and SMMHC was confined to interlobular septal regions. In contrast, the desmoplastic reaction surrounding all 10 pancreatic adenocarcinoma specimens displayed intense interstitial staining for α-SMA, SMMHC, and collagen IV but no staining for von Willebrand factor and cluster of differentiation 31. Procollagen I staining localized intracellularly to fibroblast-shaped cells within this α-SMA/SMMHC-positive scirrhous region. Islet cell tumors demonstrated an increase in α-SMA staining, although this was not as marked as in ductal adenocarcinomas. Conclusions. A massive increase in myofibroblast activity, compatible with the activation of stellate cells, is associated with the deposition of collagen types I and IV in the desmoplastic reaction around pancreatic adenocarcinomas. (Surgery 2002;131:129-34.)

Tubular phenotypic change in progressive tubulointerstitial fibrosis in human glomerulonephritis

There is much debate over the origins of fibroblast-type cells that accumulate in interstitial fibrosis. A controversial hypothesis, supported by data from animal and cell-culture studies, is that fibroblast-type cells can derive from tubular epithelial cells by a process of epithelial-mesenchymal transdifferentiation. However, to date, no evidence supports this postulate in human glomerulonephritis. This study sought to provide evidence that tubular epithelial cells can undergo phenotypic change toward a fibroblast-like cell in human glomerulonephritis. One hundred twenty-seven open renal biopsy specimens from patients with minimal change disease (MCD), immunoglobulin A (IgA) nephropathy, and rapidly progressive glomerulonephritis (RPGN) were examined for tubular phenotypic change by two-color immunohistochemistry using the criteria of de novo expression of α-smooth muscle actin (α-SMA), a myofibroblast marker; loss of the epithelial marker cytokeratin; and collagen production. In normal human kidney and MCD, tubular epithelial cells expressed cytokeratin with no evidence of α-SMA staining. However, in 36 of 90 cases of IgA nephropathy and 9 of 18 cases of RPGN, small numbers of tubular epithelial cells in areas of fibrosis showed de novo α-SMA expression, accounting for 0.4% ± 0.2% (IgA nephropathy) and 3.8% ± 1.5% (RPGN) of cortical tubules. An intermediate stage of phenotypic change was observed in some cuboidal epithelial cells that expressed both cytokeratin and α-SMA. Tubules containing α-SMA-positive (α-SMA+) cells also stained for collagen types I and III, suggesting that tubular cells undergoing phenotypic change have an active role in the fibrotic process. There also was a marked increase in transforming growth factor-β1 (TGF-β1) tubular expression in areas with interstitial fibrosis, including tubules with phenotypic change. There was a highly significant correlation between tubular α-SMA expression and interstitial fibrosis, interstitial α-SMA+ myofibroblast accumulation, deposition of collagen types I and III, tubular TGF-β1 expression, and renal dysfunction. In conclusion, this study provides evidence that tubular epithelial cells can undergo phenotypic change toward a myofibroblast-like phenotype on the basis of de novo α-SMA expression, loss of cytokeratin, and de novo collagen staining. These data, although not conclusive, provide the first support for the hypothesis that transdifferentiation of tubular epithelial cells has a role in progressive renal fibrosis in human glomerulonephritis. © 2001 by the National Kidney Foundation, Inc.