Pathogenesis Of Glomerular Diseases Research Articles

A mesangium-predominant gene, megsin, is a new serpin upregulated in IgA nephropathy.

Mesangial cells play an important role in maintaining a structure and function of the glomerulus and in the pathogenesis of glomerular diseases. To identify a specific gene expressed in human mesangial cells, we used a rapid large-scale DNA sequencing and computerized data processing to compare the transcripts in cultured human mesangial cells with various different cells and organs. Using this novel approach, we discovered a new mesangium-predominant gene termed "megsin." We obtained a full-length cDNA clone of megsin, which coded for a novel 380-amino acid protein. Amino acid homology search revealed that megsin belonged to the serpin (serine protease inhibitor) superfamily. The amino acid sequences in the reactive loop site of megsin showed characteristic features of functional serpins. Northern blot and reverse-transcribed PCR analyses of various tissues and cells demonstrated that megsin was predominantly expressed in human mesangial cells. In situ hybridization studies showed the megsin expression in the mesangium of normal glomeruli, while it increased in the expanded mesangium of glomeruli from patients with IgA nephropathy with the degree of mesangial proliferation. Here we report a new human mesangium-predominant gene that may function as an inhibitory serpin in normal and abnormal biological processes of glomerulus.

Glomerular pathology: recent advances.

The last decade has seen significant advances in the fields of cellular and molecular biology and pathology. These have contributed to our understanding of the mechanisms of glomerular disease and indicate possible novel approaches to therapy. This review discusses recent insights into the pathogenesis of glomerular disease, with consideration of the roles of intrinsic glomerular cells, infiltrating inflammatory cells, circulating permeability factors, and antibodies, and recent advances in the molecular pathology of the glomerular basement membrane. Changes in the perception of some well-established glomerular entities such as focal segmental glomerulosclerosis are considered. In addition, a number of newly-recognized specific glomerulopathies including collapsing glomerulopathy, fibrillary and immunotactoid glomerulopathy, fibronectin glomerulopathy, and collagenofibrotic glomerulopathy are briefly reviewed.

In vivo transfer of engineered macrophages into the glomerulus: endogenous TGF-beta-mediated defense against macrophage-induced glomerular cell activation.

Communication between resident glomerular cells and infiltrating macrophages plays a crucial role in the pathogenesis of glomerular disease. Using matrix metalloproteinase-9 (MMP-9) as an indicator molecule, we examined the interaction between mesangial cells and macrophages. Mesangial cells cocultured with activated macrophages or exposed to macrophage-conditioned media produced abundant MMP-9. We identified the stimulator secreted by macrophages as IL-1 because mesangial cells overexpressing IL-1 receptor antagonist protein showed a blunted expression of MMP-9 in response to the macrophage-conditioned medium. In contrast, culture supernatants of mesangial cells inhibited MMP-9 production by macrophages in a dose-dependent fashion. This inhibitor was identified to be TGF-beta1, since neutralization of TGF-beta1 abrogated the inhibitory effect of the mesangial cell-conditioned medium. To investigate whether activated macrophages induce glomerular MMP-9 expression, and if so, how endogenous TGF-beta1 modulates the induction, stimulated reporter macrophages were transferred into normal rat glomeruli or glomeruli in the regeneration phase of acute anti-Thy-1 glomerulonephritis. In the normal glomeruli, MMP-9 expression was up-regulated in resident cells after the transfer of activated macrophages. This induction was substantially repressed in the regenerating glomeruli that produced active TGF-beta1. These results point to potential mechanisms involved in glomerular control of MMP-9. Based upon the in vitro evidence, TGF-beta1 was identified as an endogenous "defender" that attenuates certain actions of infiltrating macrophages in the glomerulus.

Effect of angiotensin II on plasminogen activator inhibitor-1 production by cultured human mesangial cells.

Angiotensin II is a vasoactive peptide that has been widely implicated in the pathogenesis of glomerular disease. Some of its effects are thought to be independent of changes in blood pressure. Plasmin is a key regulator of fibrinolysis and extracellular matrix turnover. The conversion of plasminogen to plasmin by plasminogen activators (PAs) is controlled by their specific inhibitor, PAI-1. In this study we report the effects of angiotensin II on the production of PA inhibitor-1 (PAI-1) and tissue-type PA (t-PA) by glomerular mesangial cells in culture. Angiotensin II significantly increased the production of PAI-1 in the supernatant of mesangial cells (p < 0.05) in a dose-dependent manner, the maximum stimulation occurring at a concentration of 10(-5) M. The effect was not mediated by transforming growth factor-beta (TGF-beta), which is known to be induced by angiotensin II; TGF-beta itself can increase PAI-1 expression. Angiotensin II did not alter t-PA production or incorporation of matrix fibronectin but did increase cellular proliferation and 3H-thymidine uptake. The increase in PAI-1 by angiotensin II may contribute to the persistence of fibrin deposits and extracellular matrix accumulation, providing another mechanism whereby angiotensin II contributes to glomerular dysfunction.

CD45RO and CD45RA positive cell populations in idiopathic membranous and IgA glomerulopathy.

To immunophenotype and quantitate glomerular and interstitial inflammatory cells in cases of idiopathic membranous and IgA glomerulopathy; to correlate cell numbers with aspects of clinical data and renal function. Routine indirect immunoperoxidase staining was performed on frozen section renal biopsy specimens for T and B lymphocyte related antigens, macrophages and MHC class II antigens. Double immunohistochemical staining was performed to identify CD45RO+ and CD45RA+ cells. In IgA glomerulopathy correlations were found relating interstitial cell numbers to creatinine concentration at biopsy (CD45RO+ and CD45RA+ cells) and follow up creatinine concentration (CD3+, CD4+, CD8+, CD45RO+, and CD45RA+ cells). Also in IgA glomerulopathy mean arterial pressure at biopsy correlated with interstitial cell numbers and most recent follow up creatinine concentration. There were no correlations between glomerular inflammatory cells and renal function in either disease. Double staining showed that although most glomerular CD45RO+ and CD45RA+ cells were macrophages, positive cells in the interstitium were lymphocytes. The interstitial CD45RO+:RA+ ratio in normal renal biopsy specimens was approximately 5:1; for IgA glomerulopathy it was 1.5 and was 1.0 in idiopathic membranous glomerulopathy. This study demonstrates that interstitial, and not glomerular, inflammatory cell numbers correlate with renal function in primary glomerular disease and that double staining is necessary to interpret positive immunostaining for antigens located on more than one type of inflammatory cell. Detailed investigation of the interstitial CD45RO+ and CD45RA+ cells may give an insight into the pathogenesis of glomerular disease.

Transforming growth factor-beta and the pathogenesis of glomerular diseases.

Transforming growth factor-beta (TGF-beta) is a cytokine that is important in embryogenesis, development, carcinogenesis, and tissue repair. TGF-beta is unique among cytokines in its widespread actions on the regulation of extracellular matrix. In a model of acute mesangial proliferative glomerulonephritis, it was shown that overproduction of TGF-beta is the cause of pathologic matrix accumulation in the nephritic glomeruli. TGF-beta acted to increase matrix production, inhibit matrix degradation, and modulate matrix receptors in the glomerulonephritic rats. Elevated expression of TGF-beta was also found in other experimental glomerular diseases, including diabetic nephropathy. Studies of humans with glomerulonephritis and diabetic nephropathy also strongly implicated TGF-beta in the pathogenesis of glomerular matrix build-up. Recently, the proteoglycan decorin was shown to neutralize TGF-beta. When injected into glomerulonephritic rats, decorin markedly suppressed pathologic matrix deposition in the glomeruli. Thus, decorin offers hope as a treatment for progressive kidney diseases caused by the overproduction of TGF-beta.

The pathobiology of the mesangium.

Although Marcello Malpighi in 1666 was the first to call attention to the glomerulus as an important part of the mammalian kidney, it was not until the detailed microscopic observations of von Mollendorf and Zimmermann around 1930 that the central or axial, connective tissue-like space of the glomerular capillary tuft was distinguished from the glomerular endothelial and epithelial layers. This inter- or pericapillary region, harboring a third cell type, came to be known as the mesangium and became a focus of investigative interest for students of glomerular morphology, function and pathobiology [reviewed in 1]. To pathologists and nephrologists, it has been a most important finding that the expansion of the mesangium, due to increases in cellularity and extracellular matrix, is a prominent histologic abnormality in all types of chronic, progressive glomerular disease. The proliferative response in the human mesangium is most pronounced in diseases such as mesangiocapillary glomerulonephritis and IgA nephropathy. The sclerotic response as characterized by the accumulation of mesangial matrix, is a common feature of all advanced glomerular lesions and is especially prominent in diabetic nephropathy and chronic diseases such as focal and segmental glomerulosclerosis. In view of the obvious involvement of the mesangium in the pathogenesis of glomerular disease, it is important to understand the mechanisms which control the behavior of mesangial cells under physiological and abnormal conditions.

Degradation of renal basement membrane by neutral proteinases from rabbit polymorphonuclear leucocytes [proceedings

Studies of the pathogenesis of glomerular disease have suggested that polymorphonuclear leucocytes promote glomerular damage. The pattern of events observed in the autologous phase of nephrotoxic serum nephritis in rabbits indicates that the fixation of antibodies to glomerular basement membrane initiates the activation and binding of complement. This results in the chemotaxis of polymorphonuclear leucocytes t o the glomerulus where they become closely associated with the basement membrane and basement-membrane antigens are liberated into the urine; there is a concomitant alteration in the permeability characteristics of the basement membrane (Hawkins & Cochrane, 1968). These changes are prevented if the leucocytes are depleted prior to antibody administration. The cytoplasmic granules of human polymorphonuclear leucocytes are known to contain a number of neutral proteinases (Barrett, 1977), two of which (elastase and cathepsin G) can degrade renal basement membrane (Davies er al., 1978), and it has been proposed that release of these enzymes results in damage to the basement membrane in vivo. In the case of rabbits however, the granules contain mainly acidic proteinases and, although neutral proteinases with low activity against histones have been described (Davies et al., 1971), there have been no direct studies of enzymes that might hydrolyse basement membrane. Here we report the presence of neutral proteinase activity in the granules of rabbit polymorphonuclear leucocyges that act on renal basement membrane. Rabbit polymorphonuclear leucocytes were prepared from peritoneal exudates (Cohn & Hirsch, 1960) and the leucocytes were washed in 0 .34~-sucrose and disrupted by homogenization. Intact cells and nuclei were removed by centrifugation at 600g for lOrnin and the granules were harvested by centrifugation at 12000g for 20min. These were suspended in 0.01 M-sodium phosphate buffer, pH7.4, and the suspension was subjected to freezing in acetone/solid CO, and thawing at room temperature three times before centrifuging at 30000g for 30min to pellet the granular membranes. This pellet was then extracted at 0°C with 0.01 M-sodium phosphate buffer, pH7.4, containing 1 M-NaCI and 0.1 % (v/v) Triton X-100, for 20min with shaking. The suspension was centrifuged at 30000g for 30min and the pellet was extracted twice more before combining the supernatants as the membrane extract. Renal basement membrane was prepared by the method of Ligler & Robinson (1977). Hydroxyproline was measured by the method described by Blumenkrantz & Asboe-Hansen (1975), protein was determined by using the Fohn procedure described by Lowry et al. (1951). Each fraction was examined for basement-membrane-degrading activity by using a protein ratio (fraction/basement membrane) of 1 :20 incubated in 0.01 M-Tris/HCI buffer, pH7.4, containing 0.15M-NaC1, for 18 h a t 37°C with constant shaking. After incubation the undigested basement membrane was removed by centrifugation a t 2000g for lOmin and the supernatant was examined for basement-membrane breakdown products. Since collagen-like proteins are major structural components of basement membrane the release of hydrocyproline was taken as a measure of basement-membrane breakdown. Only the granular-membrane extract showed any basement-membrane-degrading activity and this could be stimulated 3-fold by increasing the NaCl concentration to 0 . 5 ~ . Under these conditions up to 15% of the basementmembrane hydroxyproline was released. The neutral proteinase activity against basement membrane was optimally active slightly above neutral pH (Fig. 1). Polyacrylamide-gel electrophoresis with 7.5 % acrylamide gels containing 1 % (w/v) sodium dodecyl sulphate revealed that high-molecular-weight (>90000) proteins had been degraded. The effects of a number of inhibitors on the enzymes were examined

Influence of cyclophosphamide treatment and neonatal thymectomy on glomerular changes in rats with long-term alloxan diabetes.

The effect of cyclophosphamide treatment and neonatal thymectomy on glomerular lesions in rats with long‐term alloxan diabetes was studied. There were no significant differences between treated and untreated diabetic rats in the occurrence of glomerular IgG, graded and measured basement membrane thickness, and mesangial area. In cyclophosphamide treated animals, however, there was a tendency toward reduced thickening of the glomerular basement membrane without decrease of glomerular IgG and of the degree of diabetes, as measured by diuresis. The results do not support the assumption that immunological mechanisms are important in the pathogenesis of glomerular disease in diabetes.