Inherited diseases of the glomerular basement membrane

The glomerular basement membrane (GBM) is a specialized form of basement membrane that has a major role in the maintenance of the glomerular filtration barrier. Like all basement membranes, it contains four main components: type IV collagen, laminin, nidogen, and heparan sulfate proteoglycans. Different isoforms of these large molecules are produced. These isoforms have a tissue-specific distribution; in the mature GBM, the major type IV collagen molecule is the alpha 3 alpha 4 alpha 5(IV) isoform, associated with laminin-521 (alpha 5 beta2 gamma 1), nidogen and agrin heparan sulfate proteoglycans. The importance of the GBM has been demonstrated by identification of hereditary glomerular diseases linked to structural anomalies of its components; for example, type IV collagen in Alport syndrome and familial benign hematuria, and laminin in Pierson syndrome. Type III collagen, an interstitial collagen, accumulates within the GBM of patients with the nail-patella syndrome, and abnormal deposition of fibronectin, another extracellular matrix protein, is characteristic of so-called fibronectin nephropathy. Development of animal models of these diseases has facilitated precise analysis of pathogenic mechanisms, but no specific treatments are available. Therapeutic trials in Alport syndrome nephropathy are underway, following promising preliminary results obtained in rodent and canine models of the disorder.     

Glomerular proteoglycans in diabetes. Partial structural characterization and metabolism of de novo synthesized heparan-35SO4 and dermatan-35SO4 proteoglycans in streptozocin-induced diabetic rats

The metabolism of glomerular proteoglycans was studied in an effort to understand the mechanisms leading to reduction of glomerular basement membrane (GBM) heparan sulfate (heparan-SO4) proteoglycan in diabetes. Glomeruli were isolated from control and streptozocin-induced diabetic rats after exposure to [35S]sulfate. A pool of rapidly metabolized 35S-glycosaminoglycans (GAG), predominantly heparan-35SO4, was present in GBMs from controls but not diabetics, whereas intact isolated glomeruli from the two groups contained similar quantities of 35S-macromolecules after 4 and 16 h in vitro. Glomeruli from diabetics contained less 35S-proteoglycan than controls after 16 h in vivo. A more rapid disappearance of [35S]sulfate from serum and an increased inorganic sulfate concentration in diabetes may account for this difference. Glomeruli from diabetics contained more heparan-35SO4 and less dermatan-35SO4 proteoglycan than control glomeruli in vitro. Diabetic glomerular heparan-35SO4 proteoglycan and its GAG chains had hydrodynamic sizes similar to controls (Mr, 13 and 1.25 X 10(4), respectively). A heparin-releasable heparan-35SO4 proteoglycan detected in isolated control glomeruli by gel electrophoresis was present in chase medium of glomeruli from diabetics in the absence of heparin. Two dermatan-35SO4 proteoglycans were synthesized in vitro. One had size and charge properties similar to glomerular heparan-35SO4 proteoglycan. A second, larger dermatan-35SO4 proteoglycan accumulated in tissue over 16 h. It was partially excluded from Sepharose CL-6B columns and eluted from Sepharose CL-4B columns at Kav = 0.32. The hydrodynamic sizes of both tissue forms of dermatan-35SO4 proteoglycans were similar in diabetics and controls. Differences in the biochemical characteristics of the major de novo synthesized glomerular proteoglycan pools could not be invoked to explain altered metabolism of GBM heparan sulfate in diabetic animals. These changes may result from diminished affinity of heparan sulfate proteoglycan for extracellular matrix or cell surfaces and may account for altered glomerular ultrafiltration properties in diabetes mellitus.     

Altered glomerular mRNA expression of basement membrane components and type I collagen in diabetic rats treated with or without insulin therapy

Glomerular basement membrane (GBM) thickening has been reported to be a characteristic change of diabetic nephropathy. Previous studies of GBM in animal models of diabetes indicated that there are no consensus in the alteration of synthesis of GBM component. The aim of this study was to determine whether the glomerular mRNA levels encoding type I and type IV collagen. B1 laminin, and heparan sulfate proteoglycan (HSPG) are altered in streptozotocin diabetic rats with or without insulin therapy. Glomerular mRNA levels for type I and type IV collagen, laminin B1, were significantly increased, but that for HSPG were marked decreased in 4 week diabetic rats compared with age-matched control rats. Insulin therapy has normalized these abnormally regulated gene expressions. Renal morphology shows no significant changes between 4 weeks diabetic rats and age-matched control rats. These results indicate that abnormal gene expressions of BM components and type I collagen might play an important role in the initiation of glomerular changes in diabetes.     

Constituents of the extracellular matrices in diabetic glomerulosclerosis

This study was undertaken to elucidate the distributions of laminin, fibronectin, type I, III, IV, V and VI collagen and heparan sulfate proteoglycan (HSPG) in diabetic nephropathy, using immunohistochemical procedures. The pathological features of diabetic glomerulosclerosis were characterized as diffuse and nodular lesions, showing an expanded mesangial matrix associated with a thickened glomerular basement membrane (GBM). In the thickened GBM, laminin was present throughout the whole membrane, type IV collagen occurred along the subendothelial side, and HSPG was present with no change in its amount. On the other hand, the components detected in the slightly expanded mesangial matrix were type IV, V and VI collagen, fibronectin and HSPG, but not laminin. When the matrix was expanded markedly, collagenous components were increased over the other components. In the typical Kimmelstiel-Wilson nodules, the mesangial matrix was occupied mainly by type V and VI collagen with a relative decrease in type IV collagen. When a nodular lesion adhered to Bowman's capsule, type I and III collagen occurred not only in Bowman's space but also within the lesion itself. Furthermore, laminin and HSPG became detectable on the outside of the lesion, but not within it. These results suggested that there was a difference in the distribution and proportion of extracellular matrix components between diffuse and nodular lesions. It appeared that the nodular lesion was not simply an advanced form of the diffuse lesion.     

Release of glomerular heparan-35SO4 proteoglycan by heparin from glomeruli of streptozocin-induced diabetic rats

Abnormalities in the incorporation of heparan sulfate proteoglycan into the glomerular basement membrane have been implicated in the pathogenesis of various proteinuric states, including diabetes mellitus. To understand further the interactions between proteoglycans and glomerular extracellular matrices, glomeruli were isolated from normal and streptozocin-induced diabetic rats after in vivo exposure to 35S-labeled sulfate and were treated with heparin in vitro. Heparin treatment released a unique heparan sulfate proteoglycan from glomerular cell surface or extracellular matrix proteoglycan receptors. Another, smaller heparan sulfate proteoglycan was the most abundant proteoglycan released into medium and was released constitutively in medium with or without added heparin. While the two heparin-extracted proteoglycans copurified on anion-exchange and gel-filtration chromatographic columns, they were resolved by composite 0.6% agarose--1.8% polyacrylamide gel electrophoresis. Glomeruli from diabetic rats contained decreased proportions of the heparin-releasable heparan sulfate proteoglycan and more constitutively released heparan sulfate proteoglycan. The apparent molecular weight and intrinsic charge of the heparin-released proteoglycan mixture and the apparent molecular weight and sulfation pattern of their 35S-labeled glycosaminoglycan chains after nitrous acid deaminative cleavage were similar in the two groups. A brief trypsin digestion of heparin-treated glomeruli released proportionately less integral membrane and extracellular matrix 35S-labeled proteoglycans and 35S-labeled glycopeptides from diabetic glomeruli than form control glomeruli. Elution of these 35S-labeled macromolecules from anion-exchange columns and migration in agarose-polyacrylamide gels were similar in the two groups. Abnormalities in proteoglycan-matrix interactions or proteoglycan processing may account for changes in the proportions of heparin- and trypsin-extracted proteoglycan compartments in diabetes.     

High-resolution ultrastructural comparison of renal glomerular and tubular basement membranes

BACKGROUND/AIMS: Glomerular basement membranes (GBM) and tubular basement membranes (TBM) consist of a fine meshwork composed mainly of type IV collagen. Each segment of tubules has specialized physiologic functions, and thus we investigated the ultrastructure of various basement membranes in rat kidneys. METHODS: Since purifying basement membranes from different tubule segments is technically challenging, we employed tissue negative staining rather than conventional negative staining to compare the ultrastructures of proximal and distal TBM and GBM in normal rats. We also assessed the distribution of extracellular matrix components including type IV collagen, laminin, heparan sulfate proteoglycan, and fibronectin in the basement membranes by immunohistochemistry. RESULTS: TBM and GBM of normal rats showed a fine meshwork structure consisting of fibrils forming small round to oval pores. Short- and long-pore diameters in proximal tubules were 3.3 +/- 0.5 and 3.9 +/- 0.6 nm, respectively, and in distal tubules 3.5 +/- 0.7 and 4.3 +/- 0.8 nm, respectively. For GBM the respective diameters were 2.5 +/- 0.5 and 3.0 +/- 0.5 nm. Immunohistochemical analysis showed no significant difference in distribution of extracellular matrix components between proximal and distal TBM. However, immunofluorescence scores of alpha1 chain of type IV collagen, fibronectin, and laminin were higher in the TBM than in the GBM. On the other hand, heparan sulfate proteoglycan was higher in the GBM. CONCLUSION: Ultrastructural differences in renal basement membranes may be related to differences in physiologic function in each segment. Copyright Copyright 1999 S. Karger AG, Basel     

Heparanase-1 gene expression and regulation by high glucose in renal epithelial cells: a potential role in the pathogenesis of proteinuria in diabetic patients

The molecular mechanisms of heparan sulfate proteoglycan downregulation in the glomerular basement membrane (GBM) of the kidneys with diabetic nephropathy remain controversial. In the present study, we showed that the expression of heparanase-1 (HPR1), a heparan sulfate-degrading endoglycosidase, was upregulated in the renal epithelial cells in the kidney with diabetic nephropathy. Urinary HPR1 levels were elevated in patients with diabetic nephropathy. In vitro cell culture studies revealed that HPR1 promoter-driven luciferase reporter gene expression, HPR1 mRNA, and protein were upregulated in renal epithelial cells under high glucose conditions. Induction of HPR1 expression by high glucose led to decreased cell surface heparan sulfate expression. HPR1 inhibitors were able to restore cell surface heparan sulfate expression. Functional analysis revealed that renal epithelial cells grown under high glucose conditions resulted in an increase of basement membrane permeability to albumin. Our studies suggest that loss of heparan sulfate in the GBM with diabetic nephropathy is attributable to accelerated heparan sulfate degradation by increased HPR1 expression.     

Altered steady-state mRNA levels of basement membrane proteins in diabetic mouse kidneys and thromboxane synthase inhibition

We examined steady-state levels of mRNA encoding type IV collagen, B1 chain of laminin, and the basement membrane heparan sulfate proteoglycan in the kidney cortex of a mouse model (KKAy) of non-insulin-dependent diabetes. mRNAs encoding laminin B1 and the proteoglycan were unchanged in kidneys taken from diabetic mice with demonstrable basement membrane thickening. mRNA levels for type IV collagen, in contrast, were significantly elevated (2-fold) in diabetic mice concurrent with but not preceding morphologically thickened basement membranes. There was a negative correlation between a ratio of proteoglycan/type IV collagen and levels of albuminuria in the diabetic mice. No correlation was noted with laminin. We also examined the effects of inhibiting the synthesis of thromboxane, a potent vasoconstrictor, on the steady-state levels of type IV collagen in the diabetic mice. Inhibition of thromboxane stopped the progression of albuminuria and prevented an increase in type IV collagen mRNA levels. We conclude that basement membrane thickening in diabetes, a hallmark of diabetic nephropathy, is partly a consequence of an unbalanced increase in the production of type IV collagen. The relative decrease in proteoglycan production may contribute to chronic albuminuria.     

A monoclonal antibody against GBM heparan sulfate induces an acute selective proteinuria in rats.

After immunization of mice with partially-purified heparan sulfate proteoglycan (HSPG) isolated from rat glomeruli, a monoclonal antibody (mAb JM-403) was obtained, which was directed against heparan sulfate (HS), the glycosaminoglycan side chain of HSPG. In ELISA it reacted with isolated human glomerular basement membrane (GBM) HSPG, HS and hyaluronic acid, but not with the core protein of human GBM HSPG, and not with chondroitin sulfate A and C, dermatan sulfate, keratan sulfate and heparin. Furthermore, it did not bind to laminin, collagen type IV or fibronectin. Specificity of JM-403 for HS was also suggested by results of inhibition studies, which found that intact HSPG and HS, but not the core protein, inhibited the binding of JM-403 to HS. In indirect immunofluorescence on cryostat sections of rat kidney, a fine granular to linear staining of the GBM was observed, along with a variable staining of the other renal basement membranes. Pretreatment of the sections with heparitinase completely prevented the binding of mAb JM-403, whereas pretreatment with chondroitinase ABC or hyaluronidase had no effect. The precise binding site of mAb JM-403 was investigated by indirect immunoelectron microscopy. It revealed a diffuse staining of the whole width of the GBM. One hour after intravenous injection of JM-403 into rats, the mAb was detected along the glomerular capillary wall in a fine granular pattern, which shifted towards a more mesangial localization after 24 hours. No binding was observed anymore by day 15. Intravenous injection induced a dose-dependent, transient and selective proteinuria that was maximal immediately after the injection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Revisiting the glomerular charge barrier in the molecular era

PURPOSE OF REVIEW: The glomerular filtration barrier consists of fenestrated glomerular endothelium, podocyte foot processes/slit diaphragms, and intervening glomerular basement membrane. Its characterization as both a size and charge-selective barrier emerged from studies conducted decades ago. The charge selectivity phenomenon is receiving renewed attention now that the identities and mechanisms of synthesis of relevant molecules are known. Here we summarize studies employing genetic or other in-vivo strategies to investigate glomerular charge. RECENT FINDINGS: Attention has focused on glomerular basement membrane heparan sulfate proteoglycans, long considered primary charge barrier components. Agrin contributes significantly to glomerular basement membrane charge but, like perlecan and collagen XVIII, is dispensable for glomerular structure and function. Disruption of glomerular heparan sulfate through transgenic methods or administration of heparanase in vivo provides further evidence against a role for heparan sulfate in glomerular function. Disruption of glomerular sialoproteins, however, causes proteinuria and indicates a critical role for these cell-associated glycoproteins in glomerular filtration. SUMMARY: Recent in-vivo manipulations of glomerular heparan sulfate proteoglycans fail to reveal a crucial role for either them or their anionic charge in glomerular filtration. In contrast, cell-associated sialoproteins are clearly important, but whether their functions actually involve contributions to the charge barrier is unknown.     

Histochemical and immunohistochemical studies of diseased human glomeruli

In order to study the localization of Lentil lectin (LCH)-binding glycoresidues in glomeruli from patients with a variety of glomerulopathies, and to elucidate the relationship between LCH-binding sugars and the components of the extracellular matrix, laminin and type IV collagen, investigations of formalin-fixed, paraffin-embedded kidney tissues digested with trypsin were carried out by the direct and indirect immunofluorescence microscopy techniques. The glomerular basement membrane (GBM) and the mesangium reacted well with LCH, whereas areas with sclerotic lesions exhibited a decreased reactivity. The pattern of LCH binding to the GBM in various glomerulopathies was similar to that of laminin but different from that of type IV collagen. The pattern of localization of LCH-reacting sites and of laminin in the GBM included the double linear lines in diabetic nephropathy, inner linear line with outer projections (spikes) in membranous nephropathy, and reduplicated basement membrane in membranoproliferative glomerulonephritis. The results obtained by enzyme-linked immunoadsorbent assay showed that LCH had a stronger reactivity for laminin than for type IV collagen or fibronectin. These findings suggest that LCH is more reactive with laminin than with other components of the glomerular extracellular matrix.     

Glomerular extracellular matrix and growth factors in diffuse mesangial sclerosis

Diffuse mesangial sclerosis, isolated (IDMS) or observed in the context of Denys-Drash syndrome (DDS) due to WT1 mutation, is characterized by early onset nephrotic syndrome progressing to renal failure. A striking morphological feature is the rapid development of glomerulosclerosis. The aims of our study were: (1) to analyze the glomerular distribution of extracellular matrix (ECM) antigens at the early stage of DMS, (2) to determine the composition of the ECM accumulated within the mesangial areas and leading to glomerular sclerosis, and (3) to analyze the expression of growth factors, transforming growth factor-β1 (TGFβ1) and platelet- derived growth factor A (PDGFA). In glomeruli of patients with IDMS and DDS, the glomerular basement membrane (GBM) expression of the heparan sulfate chain of heparan sulfate proteoglycan (HSPG) was decreased from the early stage of DMS, at a time when ECM proteins retained a normal distribution. In fully developed lesions, mesangial and subendothelial accumulation of collagenous and noncollagenous glycoproteins normally expressed in the mesangial area (types IV [α1(IV)2 α2(IV)] and VI collagen, β1 laminin, fibronectin, tenascin, and perlecan) increased with progression of mesangial sclerosis. This was associated with mesangial expression of proteins normally restricted to the GBM (agrin, α1/α5, β2, and γ1 laminin chains) and with accumulation of chondroitin sulfate proteoglycan. The distribution of the α3−α5 chains of type IV collagen was normal. Focal accumulation of types I, III, and V collagen was seen only in severely sclerotic glomeruli. Expression of growth factors TGFβ1 and PDGFA was increased in 9 of 10 and 5 of 10 patients, respectively. Early decreased GBM expression of the heparan sulfate chain of HSPG could play a role in the proteinuria of DMS patients. Changes in the composition of the ECM accumulated within the mesangial areas are not specific. We speculate that deregulation of the expression of growth factors normally downregulated by WT1, is one of the factors responsible for the rapid and massive mesangial deposition of basement membrane material in DDS. Received: 9 July 2000 / Revised: 27 November 2000 / Accepted: 22 December 2000     

Undersulfation of glomerular basement membrane heparan sulfate in experimental diabetes and lack of correction with aldose reductase inhibition

In this study we examined the effect of experimental diabetes and of treatment with an aldose reductase inhibitor on the level of sulfation of glomerular basement membrane (GBM) heparan sulfate, the principal glycosaminoglycan in this extracellular matrix. Glycosaminoglycans were isolated from GBM purified from control, streptozocin-induced diabetic, and sorbinil-treated diabetic rats and analyzed for sulfate and uronate content. Glomerular yields from diabetic kidneys were greater than those from control animals, but the amount of sulfate per glomerulus in diabetic samples, both untreated and sorbinil treated, did not differ significantly from that in control samples. However, the sulfate-to-uronate ratio in heparan sulfate isolated from diabetic GBM (0.34 +/- 0.08) was significantly less than in control samples (0.69 +/- 0.11), and treatment of diabetic rats with an aldose reductase inhibitor did not correct this reduced ratio (0.36 +/- 0.06). The results indicate that there is an undersulfation of heparan sulfate of GBM in experimental diabetes, an abnormality that may contribute to loss of anionic sites and decreased charge selectivity of the glomerular filtration barrier. The findings further suggest that this abnormality results from disturbances in glycosaminoglycan synthesis and/or metabolism in diabetes that are independent of polyol-pathway activation in the renal glomerulus.     

Occurrence of type VI collagen in extracellular matrix of renal glomeruli and its increase in diabetes

Human and bovine glomerular basement membrane (GBM) preparations, representing the extracellular matrix of the renal filtration units, were found to contain type VI collagen. This protein was solubilized by guanidine and guanidine-dithiothreitol extractions and characterized after polyacrylamide gel electrophoretic resolution by immunoblotting with an antiserum directed against the alpha 1(VI) and alpha 2(VI) polypeptide chains and by its insensitivity to collagenase digestion in the nonreduced state. In contrast to GBM, which is the product of three distinct cells, type VI collagen could not be detected in extracts from calf lens capsule, an epithelial cell-derived basement membrane. Quantitation by radioimmunoassay of the type VI collagen content of GBM from 17 diabetic and 15 nondiabetic human subjects indicated a 2.8-fold higher level (P less than 0.001) in the diabetic preparations. Because in the glomerulus type VI collagen is considered on the basis of immunohistochemistry to be localized to the mesangium, we believe that measurement of this protein in GBM preparations can provide a valuable index of mesangial expansion in diabetic and other glomerulopathies.     

SLE nephritis: an ultrastructural immunogold study to evaluate the relationship between immune complexes and the basement membrane components type IV collagen, fibronectin and heparan sulphate proteoglycans

The nature of immune complexes and their relationship to the normal glomerular basement membrane (GBM) components type IV collagen, fibronectin, and heparan sulphate proteoglycans (HSPG) have been examined in the glomeruli of 7 cases of systemic lupus erythematosus (SLE) glomerulonephritis using an ultrastructural immunogold technique. In paraformaldehyde-fixed, Lowicryl resin-embedded tissue, the electron-dense deposits contained IgG, IgM, IgA, and C3 whether they were subepithelial, intramembranous, subendothelial, or mesangial and there was no particular relationship between the class of immunoglobulin and site of immune complex localization within the glomerulus. The normal GBM components type IV collagen, fibronectin, and HSPG were found within all the glomeruli, but did not have the same distribution. Type IV collagen and fibronectin were found predominantly on the inner aspect of the GBM and diffusely throughout the more central regions of the mesangial matrix. By contrast the HSPG was seen mainly on the outer aspect of the GBM and at the periphery of the mesangial matrix. In none of the cases were GBM antigens localized within the electron-dense deposits, results which suggest that autoantibodies to these GBM components may not play a role in the development of the glomerulonephritis.     

Loss of heparan sulfate glycosaminoglycan assembly in podocytes does not lead to proteinuria

Podocytes synthesize the majority of the glomerular basement membrane components with some contribution from the glomerular capillary endothelial cells. The anionic charge of heparan sulfate proteoglycans is conferred by covalently attached heparan sulfate glycosaminoglycans and these are thought to provide critical charge selectivity to the glomerular basement membrane for ultrafiltration. One key component in herparan sulfate glycosaminoglycan assembly is the Ext1 gene product encoding a subunit of heparan sulfate co-polymerase. Here we knocked out Ext1 gene expression in podocytes halting polymerization of heparin sulfate glycosaminoglycans on the proteoglycan core proteins secreted by podocytes. Glomerular development occurred normally in these knockout animals but changes in podocyte morphology, such as foot process effacement, were seen as early as 1 month after birth. Immunohistochemical analysis showed a significant decrease in heparan sulfate glycosaminoglycans confirmed by ultrastructural studies using polyethyleneimine staining. Despite podocyte abnormalities and loss of heparan sulfate glycosaminoglycans, severe albuminuria did not develop in the knockout mice. We show that the presence of podocyte-secreted heparan sulfate glycosaminoglycans is not absolutely necessary to limit albuminuria suggesting the existence of other mechanisms that limit albuminuria. Heparan sulfate glycosaminoglycans appear to have functions that control podocyte behavior rather than be primarily an ultrafiltration barrier.     

Recent insights into the structure and functions of heparan sulfate proteoglycans in the human glomerular basement membrane.

As the first barrier to be crossed on the way to urinary space, the glomerular basement membrane (GBM) plays a key role in renal function. The permeability of the GBM for a given molecule is highly dependent on its size, shape and charge. As early as 1980, the charge-selective permeability was demonstrated to relate to the electrostatic properties of covalently bound heparan sulfates (HS) within the GBM. Since the identification of perlecan as a heparan sulfate proteoglycan (HSPG) of basement membranes, the hypothesis that perlecan could be a crucial determinant of GBM permselectivity received considerable attention. In addition to perlecan, the GBM also contains other HSPG species, one of which was identified as agrin. The high local expression of agrin in the GBM, together with the presence of agrin receptors at the cell matrix interface, suggests that this HSPG contributes to glomerular function in multiple ways. Here, we review the current knowledge regarding the structure and functions of HSPGs in the GBM, and discuss how these molecules could be involved in various glomerular diseases. Possible directions for future investigation are suggested.     

Effects of methylprednisolone on glomerular and medullary mRNA levels for extracellular matrices in puromycin aminonucleoside nephrosis.

We examined the effects of methylprednisolone (MPSL) on type IV collagen, laminin and heparan sulfate proteoglycan (HSPG) mRNA levels in the renal glomeruli and medulla of puromycin aminonucleoside (PAN) nephrosis. mRNA levels encoding for type IV collagen and laminin increased markedly, whereas those for HSPG decreased significantly in glomeruli of PAN nephrosis. Administration of MPSL partially ameliorated the abnormal gene expression for basement membrane components. Furthermore, we showed that medullary mRNA levels for all these basement membrane components decreased with age in PAN nephrosis with or without MPSL treatment, suggesting that neither PAN nor MPSL has any effect on basement membrane component mRNA levels in the renal medulla. In contrast, mRNA levels for the interstitial collagens including alpha 1 (I) and alpha 1 (III) chains in glomeruli showed little change with or without MPSL treatment, whereas those in medulla increased significantly in PAN nephrosis when compared with the control. MPSL ameliorated the abnormal gene expression of alpha 1 (I) and alpha 1 (III) collagen in renal medulla. These results indicate that PAN affects both glomerular mRNA encoding for basement membrane components and medullary mRNA encoding for interstitial collagens, and that MPSL has marked effects on the amelioration of abnormal gene expression in both glomeruli and medulla of PAN nephrosis.