Simone Peyrol

Mechanical forces induce scar remodeling. Study in non-pressure-treated versus pressure-treated hypertrophic scars.

Reparative process of second and third degree burns usually results in hypertrophic scar formation that can be treated by pressure. Although this method is efficient, its mechanisms of action are not known. In this work, we have studied the histological organization of hypertrophic scars submitted to pressure. Skin biopsies were performed 2 to 7 months after the onset of treatment in two adjacent regions of the scar, non-pressure- or pressure-treated and analyzed by immunohistochemistry and transmission electron microscopy for extracellular matrix organization and cellular morphology. In non-pressure-treated regions, fibrillin deposits did not present the classical candelabra-like pattern under epidermis and were reduced in dermis; in pressure-treated regions the amount was increased compared to non-pressure-treated regions but the organization was still disturbed. In non-pressure-treated regions, elastin was present in patch deposits; in pressure-treated regions elastin formed fibers, smaller than in normal dermis. Tenascin was present in the whole dermis in non-pressure-treated regions, whereas in pressure-treated regions it was observed only under epidermis and around vessels, as in normal skin. alpha-Smooth muscle actin-expressing myofibroblasts were absent in normal skin, present in large amounts in non-pressure-treated regions, and almost absent in pressure-treated regions. The disturbed ultrastructural organization of dermal-epidermal junction observed in non-pressure-treated regions disappeared after pressure therapy; typical features of apoptosis in fibroblastic cells and morphological aspects of collagen degradation were observed in pressure-treated regions. Our results show that, in hypertrophic scars, pressure therapy restores in part the extracellular matrix organization observed in normal scar and induces the disappearance of alpha-smooth muscle actin-expressing myofibroblasts, probably by apoptosis. We suggest that the pressure acts by accelerating the remission phase of the postburn reparative process.

Coexpression of the lysyl oxidase‐like gene (LOXL) and the gene encoding type III procollagen in induced liver fibrosis

We have isolated a mouse lysyl oxidase‐like (LOXL) cDNA from a mouse embryo cDNA library and used this cDNA to measure changes in steady state levels of LOXL mRNA during the development of carbon tetrachloride‐induced liver fibrosis in adult mice. These results revealed the coincident appearance of increased steady state levels of LOXL mRNA and type III procollagen mRNA early in the development of liver fibrosis. In contrast, steady state levels of lysyl oxidase mRNA increased throughout the onset of hepatic fibrosis and appeared in parallel with the increased steady state levels of pro‐αI (I) collagen mRNA. These findings suggest that the LOXL protein (possibly an isoform of lysyl oxidase) is involved in the development of lysine‐derived cross‐links in collagenous substrates. Moreover, the substrate specificity of the LOXL protein may be different to that of lysyl oxidase and this difference may be collagen‐type specific. J. Cell. Biochem. 72:181–188, 1999.

Isolation and characterization of brain microvascular endothelial cells from Saimiri monkeys. An in vitro model for sequestration of Plasmodium falciparum-infected erythrocytes.

The adhesion of parasitized red blood cells (PRBC) to the endothelium (sequestration) may contribute to the pathogenic events in severe human malaria caused by P. falciparum. However, the factors involved in the pathophysiology, especially cerebral malaria are poorly understood. Previously, we have shown that the squirrel monkey Saimiri sciureus is a potential model for human cerebral malaria. In this paper we describe five stable clones of endothelial cell lines isolated immediately postmortem from different regions of the brain of Saimiri monkeys. The endothelial cell characteristics of these clones were confirmed by analyzing their ultrastructural aspects by transmission electron microscopy and by immunodetection of various endothelial cell markers. The Saimiri brain endothelial cell clones (SBEC) varied in their expression of different surface molecules. For example, various combinations of receptors involved in P. falciparum PRBC adherence such as CD36, ICAM-1 and E-selectin, were expressed at baseline values and could be up-regulated by human srTNF-alpha and human srIFN-gamma. One of the SBEC clones showed a strong cytoadherence for various laboratory strains of P. falciparum despite the absence of surface expression of any of the known endothelial receptors implicated in PRBC adherence. This finding suggests the existence of a new and uncharacterized PRBC binding receptor. The use of target organ specific endothelial cell lines expressing a number of different potential P. falciparum PRBC cytoadherence receptors, will be a useful in vitro system for the evaluation of strategies for the development of vaccine and antimalarial drugs to prevent human cerebral malaria.

Lobular - but not periovular - inhibition of collagen deposition in the liver of S. Mansoni infected mice using interferon-γ

BACKGROUND/AIMS Interferon-gamma (IFNgamma) elicits antiproliferative and antifibrogenic activity in a variety of mesenchymal cells, including hepatic stellate cells (Ito cells), and therefore represents a possible drug for liver fibrosis. However, IFNgamma binds to heparan sulfate, and is localized by these molecules in a restricted area within the tissue. For example, in rat liver, it has been shown that following injection, IFNgamma was concentrated in a restricted area by heparan sulfate. The aim of this study was to analyze, at the tissular level in the liver, the antifibrogenic activity of IFNgamma. METHODS Chronic inflammation due to Schistosoma infection induces hepatic fibrogenesis around the parasite eggs (portal fibrosis) and in the parenchyma (lobular fibrosis). Infected mice were treated with recombinant IFNgamma, and the collagen content of the liver was evaluated by means of biochemical dosages, histologic and morphometric examination of liver tissue, and electron microscopic analysis. RESULTS IFNgamma reduced the whole liver collagen content by 28% compared to control mice. In control mice, collagen was found around eggs and infiltrating the parenchyma, associated with a diffuse array of inflammatory cells, while in treated mice the collagen was present only around eggs and surrounded by a dense layer of inflammatory cells. Therefore, collagen was measured in isolated granulomas and in the remaining parenchyma. We found that IFNgamma strongly reduced the parenchymal collagen (74%), but had no effect on the granuloma collagen content. CONCLUSIONS Together these data demonstrate that IFNgamma did not act in a homogeneous manner in the liver. Since granulomas are almost completely devoid of heparan sulfate, these data could suggest, among others hypotheses, that heparan sulfate which binds IFNgamma either localizes or mediates the cytokine activity outside the granulomas.

Connective matrix organization in human pulmonary fibrosis. Collagen polymorphism analysis in fibrotic deposits by immunohistological methods.

SummaryIn the interstitium of the alveolar septa in the peripheral parts of the lung, four molecular types of collagen (I, III, IV and V) each with different morphological appearances, can be identified. The structural integrity of collagens accounts for the physiological efficiency of the lung. Fibrous thickening of alveolar septa is an invariable result of various diseases affecting the interstitium of the lung.The light and electron microscopic findings, and the immunological typing of collagens in six cases of fibrotic alveolar disease, are described. In the alveolar septa, two different compartments (the alveolo-capillary junction and the supportive axis) were affected by fibrosis: the alveolo-capillary junction was widened by the addition of interstitial collagens to basement membranes. In the axis, the increase of interstitial (types I and III) collagen gave rise to different patterns of connective matrix organization, graded as Loose or Dense depending on quantitative alterations of the type I/III ratio.The mode of organization of the fibrotic lung connective matrix, which depends on the quality of deposits in the matrix, may be correlated with the evolution of interstitial pulmonary fibrosis, in terms of its stability, remodelling ability and reversibility.

Fibrosing vasculitis in Wegener's granulomatosis : ultrastructural and immunohistochemical analysis of the vascular lesions

This study of two cases of pulmonary Wegeners granulomatosis (WG) focuses on the ultrastructural aspects of the vascular wall injury and on the immunohistochemical characterization of the perivascular connective matrix. The iterative waves of endothelial cell necrosis and regeneration are demonstrated by the multiplamellar appearance of the basal lamina. Neutrophils infiltrate the vessel wall and myofibroblasts are recruited to injured vessels. The perivascular connective matrix associates basement-membrane like and fibrillar material with fibrin deposits. The initiation of the fibrosing process was assessed by the visualization of matrix molecules involved in targeting (p-fibronectin), organizing (cellular fibronectin and tenascin) and stabilizing (lysyl-oxidase) the fibrogenic activity. These elementary lesions affect different levels of the vascular tree, and capillaritis is involved in the extension of the pathological process. Lysyl-oxidase labelling reveals the fibrosing front which is located on the border of dense fibrosis. The markers of fibrosing activity disappear in the areas of fibrosis following vasculitis and/or ischaemic necrosis and/or granulomatosis. Vasculitis plays a major role in both the genesis and progression of the fibrosis observed in the late stage of WG.

Ultrastructural aspects of corneal fibrous tissue in the scheie syndrome

SummaryAn ultrastructural study of the corneal fibrous tissue was performed in a case of Scheie’s syndrome. Mucopolysaccharidosis deposits in keratocytes were observed as electron-clear and electron-dense inclusions.Modifications of the extracellular space included modifications of lamellar collagen organization and local hypertrophy of collagen bundles; presence of microfibrillar dense material isolating large irregular collagen fibers; and presence of fibrous long spacing type collagen fibers.The significance of these changes is discussed. This special form of collagen organization is supposed to appear in a modified microenvironment, that is the presence of an abnormal concentration of proteoglycans.

Constrictive bronchiolitis obliterans. Characterisation of fibrogenesis and lysyl oxidase expression patterns.

Abstract. The process leading to irreversible fibrotic constriction of the bronchioles was studied in two cases of bronchiolitis obliterans (BO) after bone marrow transplantation. Because lysyl oxidase (LOX) is the main collagen cross-linking enzyme that might account for irreversible fibrosis, its expression was studied together with expression of extracellular matrix (ECM) proteins. Characteristic types of lesions could be distinguished on the basis of histological and immunohistological criteria. An inflammatory stage was characterised by infiltration restricted to the bronchioles by lymphocytes and dendritic cells. A fibro-inflammatory stage was characterised by the coexistence of a persistent immune cellular lesion pattern with further focal modelling of a sub-epithelial neo-synthesised connective matrix. LOX expression was observed at the tips of intra-luminal fibrotic protrusions, together with tenascin and cellular fibronectin. A fibrotic stage was characterised by dense ECM deposits spreading throughout the peri-bronchiolar connective tissue, resulting in bronchiole obliteration and final disappearance. In contrast to reversible cases of fibrosis, persistence of long-term LOX expression reflecting continuing fibrosing activity might account for the irreversible status of BO. Our two cases illustrated that, at inflammatory and fibro-inflammatory stages, BO may be stabilised by immunosuppressive treatment, while the persistence of LOX expression in the fibrotic stage might correspond to a disease that becomes irreversible and fatal.