Development Of Glomerular Lesions Research Articles

The anti-inflammatory effects of Hedyotis diffusa Willd on SLE with STAT3 as a key target

Ethnopharmacological relevanceHedyotis diffusa Willd, also named Scleromitrion diffusum (Willd.) R.J. Wang, is one medical herb, which has been traditionally used by the She nationality in China. And H. diffusa represents a beneficial effect on Systemic lupus erythematosus (SLE) treatment in clinic. Aim of the studyThe underlying mechanisms of the protective effects of H. diffusa on SLE remain unclear. In this study, we treated MRL/lpr mice with H. diffusa water extract (HDW) to assess its therapeutic effects and verified its regulating signalling pathway through cytological experiments. Materials and methodsIn the present study, the constituents of HDW were analysed through ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) and SCIEX OS software. The protective activity and underlying mechanisms were studied in a MRL/lpr lupus mouse model. The blood cells, autoantibodies, metabolites and the cytokines in serum were identified with a hematology analyzer, specific ELISA kit, GC/MS system and cytometric assays. The histological and immunohistochemical analysis were engaged in the morphologic, and the expression and translocation of the crucial protein observation. The dual luciferase reporter assay was applied to identifying the regulative activity of HDW. The transcription and translation expression of the protein was studied by real-time PCR and Western blot assays. The network pharmacology analysis was employed to predict the IL-6/STAT3 pathway regulators and the screen the STAT3 inhibitors in HDW. ResultsThe results revealed the capability of HDW to attenuate the production of autoantibodies, secretion of inflammatory cytokines (IL-6 and IFN-γ), and suppressed the IgG and C3 deposition, the development of glomerular lesions in MRL/lpr mice. Serum metabolomics study showed the improvement in serum metabolites, especially aminoacyl-tRNA biosynthesis, by HDW. IL-6 was clarified to be highly associated with the significantly changed metabolites in network analysis. We further demonstrated the effects of HDW on the IL-6/STAT3 pathway in vivo and in vitro. ConclusionsThis study suggested that HDW exerts a therapeutic effect in SLE model mice by suppressing the IL-6/STAT3 pathway.

P0018GLOMERULAR ENDOTHELIAL CELL SENESCENCE DRIVES AGE-RELATED KIDNEY DISEASE THROUGH PAI-1

Abstract Background and Aims In age-related chronic kidney disease (CKD), the most frequent histological lesion observed is glomerulosclerosis. The molecular mechanisms involved in this deterioration process are unclear, but cellular senescence might play a role. Method By combining several murine models of physiological and accelerated aging with transgenic animals and in vitro models, we discovered the role of endothelial senescence in the development of glomerular lesions. Results These senescent glomerular endothelial cells secreted several molecules, grouped under the senescence associated secretory phenotype (SASP) including the plasminogen activator inhibitor 1 (PAI-1). Specific deletion of PAI-1 in endothelial cells prevented the development of glomerulosclerosis during physiological and accelerated aging, by decreasing podocyte loss. In addition, we showed that PAI-1 mediates a detrimental endothelial-podocyte crosstalk, as incubation of podocytes by supernatant of senescent glomerular endothelial cells led to their detachment. Consistently, preincubation of the senescent supernatant with tiplaxtinin, a PAI-1 inhibitor, preserved podocytes. More importantly, we demonstrated that these data are relevant to humans. In fact, PAI-1 staining the day of the transplantation was predictive of kidney allograft dysfunction 12 months after transplantation from elderly donors. Conclusion In conclusion, our study uncovers the critical role played by endothelial senescence in the development of glomerulosclerosis during aging and identified PAI-1 as a novel promising biomarker for predicting kidney dysfunction in patients receiving a kidney from elderly donors.

The tetraspanin CD9 controls invasive migration and proliferation of parietal epithelial cells and glomerular disease progression

Introduction Crescentic glomerulonephritis (CGN) and focal segmental glomerulosclerosis (FSGS) are life-threatening diseases leading to irreversible renal failure. Increasing evidence supports the notion that glomerular parietal epithelial cells (PEC) are implicated in the formation of crescents in CGN and in glomerular scarring during FSGS. However, the mechanisms underlying the change in PEC phenotype remain unclear, as do those driving their oriented migration towards the glomerular tuft and their increased capacity to proliferate and form hypercellular and sclerotic lesions. Methodes Aiming to identify factors underlying PEC activation, we revealed increased expression of CD9 using comparative deep RNA sequencing of mouse and human normal and diseased glomeruli, confirmed expression by immunohistochemistry, and investigated roles of this tetraspanin in CGN and FSGS pathogeny. As CD9 de novo expression in glomeruli was observed in PEC in human CGN and FSGS and, given the recognized role of PEC in the progression of extracapillary lesions, we generated mice with a specific Cd9 deletion in PEC (iPec-Cd9lox/lox mice) that were challenged in experimental CGN and FSGS models. Cd9 gene knock-down were performed in a PEC line to evaluate its influence on migration, spreading and proliferation. Results During the CGN model, iPec-Cd9 wt/wt control mice showed a rapid increase in albuminuria, renal failure and severe glomerular crescentic lesions. Conversely, iPec-Cd9lox/lox mice displayed marked renal protection. We challenged iPec-Cd9lox/lox mice with the DOCA-salt and unilateral nephrectomy FSGS model. Glomerular ultrastructure and function was preserved in iPec-Cd9lox/lox DOCA mice, whereas iPec-Cd9 wt/wt DOCA mice exhibited flocculo-capsular synechiae and sclerosis. In vitro, we found that CD9 deficiency impairs the ability of PECs to proliferate and to migrate towards a steep gradient of chemoattractant in microfluidic channels. Conclusion These results indicate that de novo expression of CD9 in PECs participates in crescents in experimental CGN and is also critically involved in the development of glomerular lesions in FSGS.

Role of Macrophage Stimulating Protein 1 (MSP1) in the Development of Glomerulopathy in Sickle Cell Disease

BackgroundMore than 50% patients with sickle cell disease (SCD) develop renal glomerular disease. Its pathophysiology is likely to be multifactorial, being affected by hyperfiltration, glomerular hypertension, ischemia-reperfusion injury, oxidative stress, and endothelial dysfunction. Ischemia-reperfusion is associated with significant recruitment of glomerular and interstitial macrophages. Recently, Macrophage Stimulating Protein 1 (MSP1) was shown to be involved in the development of anti-Thy1 glomerulonephritis in rat model that also associated with renal macrophages infiltration. MSP1 is produced by the liver and secreted into the circulation as a non-active protein. It is cleaved and activated locally by a macrophage membrane-associated proteinase. We hypothesesize that infiltrating macrophages in SCD activate MSP1 protein that accumulates in the renal glomeruli and induces endothelial cell and podocytes injury.ObjectiveWe analyzed MSP1 expression in vivo in SCD mouse model and the effect of recombinant MSP1 on cultured human podocytes and endothelial cells.MethodsTransgenic SCD mice were obtained from Jackson Laboratory (B6;129-Hbatm1(HBA)Tow Hbbtm2(HBG1,HBB*)Tow/Hbbtm3(HBG1,HBB)Tow/J Townes strain). Townes mice produce approximately 94% human sickle (HbS), 6% human fetal hemoglobin (HbF), and no murine hemoglobin. Control animals are carrying two copies of the transgene encoding human α1-globin gene and two copies of the Hbbtm3(HBG1,HBB)Tow mutation (human hemoglobin gamma (Aγ) gene and the human wildtype hemoglobin beta (βA) gene). Townes mice spontaneously develop renal glomerular lesions. Kidneys were collected from 5 months old mice and immunohistochemistry (IHC) was carried to detect macrophages, endothelial cells (CD34), podocytes (WT-1) and MSP1. Human glomerular endothelial cell line (HGEC) and two human podocyte lines (PD1 and PD2) were treated with recombinant MCP1 and the cellular motility, permeability, growth and capillary formation were analyzed.ResultsSCD mice developed focal segmental glomerulosclerosis. It was associated with glomerular capillary aneurisms, loss of podocytes and increased macrophages infiltration. IHC demonstrated accumulation of MSP1 in the capillaries of affected glomeruli. In vitro, MSP1 treatment increased motility of both endothelial cells and podocytes. In addition, MSP1 significantly reduced podocytes growth and viability. MSP1 also inhibited capillary formation by endothelial cells on Matrigel.ConclusionActivated MSP1 is likely to be involved in the development of glomerular lesions in SCD which could be due to the modulation of endothelial cell and podocytes function. Further analysis is needed to elucidate the role of MSP1 in the development of glomerular disease in humans.AcknowledgmentsThis work was supported by NIH Research Grants (1P50HL118006, 1R01HL125005 and 5G12MD007597). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. DisclosuresNo relevant conflicts of interest to declare.

Skimmin, a Coumarin fromHydrangea paniculata, Slows down the Progression of Membranous Glomerulonephritis by Anti-Inflammatory Effects and Inhibiting Immune Complex Deposition

Skimmin is one of the major pharmacologically active molecules present in Hydrangea paniculata, a medical herb used in the traditional Chinese medicine as an anti-inflammatory agent. In the current study, we attempted to investigate its renoprotective activity and underlying mechanisms in a rat model of membranous glomerulonephritis induced by cationic bovine serum albumin (c-BSA). Sprague-Dawley (SD) rats were divided into five groups, including normal control, model control, Mycophenolate Mofetil-treated group, and two skimming-treated groups (15 mg/kg and 30 mg/kg). Our research showed that treatment with skimmin significantly reduced the levels of blood urea nitrogen (BUN), urinary albumin excretion (UAE), and serum creatinine (Scr) as compared with model control after experimental induction of membranous glomerulonephritis (P < 0.01). Moreover, glomerular hypercellularity, tubulointerstitial injury, and glomerular deposition of IgG were less intense after skimmin treatment. By immunochemistry analysis, we demonstrated that skimmin could significantly inhibit interleukin-1β (IL1β) and IL-6 expression (P < 0.05), reduce the loss of nephrin and podocin, and suppress the infiltration of renal interstitium by CD3-positive T cell and CD20-positive B cell. These results suggest that treatment with skimmin can significantly improve renal function and suppress the IgG deposition as well as the development of glomerular lesions in a rat model of membranous glomerulonephritis.

Fractalkine expression and CD16+monocyte accumulation in glomerular lesions: association with their severity and diversity in lupus models

Fractalkine (Fkn) is expressed on injured endothelial cells and is a membrane-bound chemokine that attracts cells expressing its receptor, CX3CR1, including CD16(+) monocytes (CD16(+) Mos). To clarify the role played by Fkn in the development of glomerular lesions in lupus nephritis, we examined Fkn expression and CD16(+) Mo accumulation induced in experimental C.B-17/Inc-scid/scid (SCID) lupus model mice by injection of IgG(3)-producing hybridoma clones obtained from MRL/lpr mice. Glomerular Fkn expression and accumulation of CD16(+) Mos were semiquantitatively evaluated using laser capture microdissection and real-time PCR. Injection of the 2B11.3 and 7B6.8 clones induced formation of glomerular proliferative and wire-loop lesions, respectively. Immunohistological analysis of the localization of Fkn and CD16(+) Mos revealed that Fkn expression and CD16(+) Mo accumulation were markedly elevated in glomerular lesions induced by 2B11.3, whereas no elevation was detected in those induced by 7B6.8. In addition, to examine the contribution of glomerular Fkn to the development of proliferative lesions, L cells producing an Fkn antagonist (Fkn-AT) were transplanted into SCID mice exhibiting proliferative lupus nephritis (DPLN) induced by 2B11.3. Notably, transplantation of the Fkn-AT-producing cells was functionally and histologically protective against this DPLN. Taken together, our findings suggest that Fkn and CD16(+) Mo accumulation are partially associated with the severity and diversity of histology of lupus nephritis.

α- and β-Adducin polymorphisms affect podocyte proteins and proteinuria in rodents and decline of renal function in human IgA nephropathy

Adducins are cytoskeletal actin-binding proteins (α, β, γ) that function as heterodimers and heterotetramers and are encoded by distinct genes. Experimental and clinical evidence implicates α- and β-adducin variants in hypertension and renal dysfunction. Here, we have addressed the role of α- and β-adducin on glomerular function and disease using β-adducin null mice, congenic substrains for α- and β-adducin from the Milan hypertensive (MHS) and Milan normotensive (MNS) rats and patients with IgA nephropathy. Targeted deletion of β-adducin in mice reduced urinary protein excretion, preceded by an increase of podocyte protein expression (phospho-nephrin, synaptopodin, α-actinin, ZO-1, Fyn). The introgression of polymorphic MHS β-adducin locus into MNS (Add2, 529R) rats was associated with an early reduction of podocyte protein expression (nephrin, synaptopodin, α-actinin, ZO-1, podocin, Fyn), followed by severe glomerular and interstitial lesions and increased urinary protein excretion. These alterations were markedly attenuated when the polymorphic MHS α-adducin locus was also present (Add1, 316Y). In patients with IgA nephropathy, the rate of decline of renal function over time was associated to polymorphic β-adducin (ADD2, 1797T, rs4984) with a significant interaction with α-adducin (ADD1, 460W, rs4961). These findings suggest that adducin genetic variants participate in the development of glomerular lesions by modulating the expression of specific podocyte proteins.Electronic supplementary materialThe online version of this article (doi:10.1007/s00109-009-0549-x) contains supplementary material, which is available to authorized users.

A murine model of Denys–Drash syndrome reveals novel transcriptional targets of WT1 in podocytes

The Wilms tumor-suppressor gene WT1, a key player in renal development, also has a crucial role in maintenance of the glomerulus in the mature kidney. However, molecular pathways orchestrated by WT1 in podocytes, where it is highly expressed, remain unknown. Their defects are thought to modify the cross-talk between podocytes and other glomerular cells and ultimately lead to glomerular sclerosis, as observed in diffuse mesangial sclerosis (DMS) a nephropathy associated with WT1 mutations. To identify podocyte WT1 targets, we generated a novel DMS mouse line, performed gene expression profiling in isolated glomeruli and identified excellent candidates that may modify podocyte differentiation and growth factor signaling in glomeruli. Scel, encoding sciellin, a protein of the cornified envelope in the skin, and Sulf1, encoding a 6-O endosulfatase, are shown to be expressed in wild-type podocytes and to be strongly down-regulated in mutants. Co-expression of Wt1, Scel and Sulf1 was also found in a mesonephric cell line, and siRNA-mediated knockdown of WT1 decreased Scel and Sulf1 mRNAs and proteins. By ChIP we show that Scel and Sulf1 are direct WT1 targets. Cyp26a1, encoding an enzyme involved in the degradation of retinoic acid, is shown to be up-regulated in mutant podocytes. Cyp26a1 may play a role in the development of glomerular lesions but does not seem to be regulated by WT1. These results provide novel clues in our understanding of normal glomerular function and early events involved in glomerulosclerosis.

Possible role of autoantibodies against nephrin in an experimental model of chronic graft-versus-host disease

Nephrin, a product of the NPHS1 gene, is a component of the slit diaphragms that are found between glomerular foot processes and is a crucial element for glomerular filtration barrier. Recently, nephrin has been focused in a number of studies of proteinuria development including various types of acquired glomerular diseases including minimal change nephrotic syndrome and membranous nephropathy. However, the precise role of nephrin in such acquired glomerular diseases is still unknown. To analyse the role of nephrin further, two kinds of anti-nephrin antibodies were raised in the rabbits and applied to an experimental mouse model of chronic graft-versus-host disease, in which (C57BL/10 x DBA/2) F1 mice developed clinically apparent severe proteinuria with significant glomerular lesions 7 weeks after parental DBA/2 cell transfer. Antibody-sandwich ELISA detected anti-nephrin antibodies during week 2 to week 6, with the peak at week 2 or week 4. Colocalization of nephrin and IgG on week 4, week 6, and week 8 was revealed by confocal microscopic analysis, suggesting that in situ immune complex formation with nephrin in glomerular lesion. Taken together, it seems to be suggested nephrin and its autoantibody have a certain role in the development of glomerular lesion in our model mice.

Treatment of experimental immune complex glomerulonephritis by sodium alginate

We have studied the therapeutic efficacy of the sodium alginate in experimental immune complex glomerulonephritis. Bovine serum albumin (BSA) nephritis was induced in rats by a subcutaneous immunization and daily intravenous administration of BSA. Sodium alginate at two different doses (25 and 50 mg/kg) was administered intraperitoneally at regular 72-h intervals for 6 weeks. Onset of treatment was day 42. Urinary protein was measured weekly and serum anti-BSA antibody was assessed by ELISA method at different intervals. Animals were euthanized at the 12th experimental week and blood samples and kidney specimens were obtained. BUN, serum creatinine and serum cholesterol and triglyceride were measured at the time of sacrifice. Kidney specimens were processed for light and immunofluorescent microscopic examination. The tolerability and inhibitory effect of LVA on matrix metalloproteinase 2 (MMP-2) were tested using WEHI-164 cell line and zymography method. Results of this experiment showed that treatment with sodium alginate could significantly reduce the urinary protein excretion and serum creatinine in treated rats vs. nontreated controls. Anti-BSA antibody titers were lower in treated rats than in controls at the 12th week post-immunization. There was no significant difference in the level of BUN and serum lipids between two groups. Whereas, glomerular hypercellularity, PMN infiltration and glomerular deposition of BSA were less intense in treated rats vs. controls. Moreover, in vitro examinations revealed that treatment with LVA, as a very safe agent could diminish MMP-2 activity. These results suggest that treatment with sodium alginate as a new immunosuppressive agent can reduce proteinuria, inhibit MMP-2 activity and suppress the antibody production as well as the development of glomerular lesions in a rat model of immune complex glomerulonephritis.

Aggregated IgG Bind to Glomerular Epithelial Cells to Stimulate Urokinase Release through an Endocytosis-Independent Process

Background: In membranous nephropathy, the development of glomerular lesions is related to the formation of immune complexes at subepithelial sites. These deposits are associated with modifications in the fibrinolytic activity of glomerular cells leading to the appearance of fibrin degradation products in the deposits and the urine. A previous study has shown that immune complexes interact with glomerular epithelial cells (GEC) through the neonatal Fc receptor (FcRn). We therefore determined whether this binding could be responsible for a modification in the fibrinolytic activity of GEC. Methods: Endocytosis of heat-aggregated immunoglobulins (AgIgG) in cultured human GEC was studied by immunofluorescence and confocal microscopy. The release of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor 1 (PAI) by GEC or whole glomeruli was assessed by ELISA, fibrin zymography and Northern blot. Results: Human GEC in culture bound AgIgG that possess characteristics similar to soluble immune complexes and internalized them by 10 min. This process was mediated by FcRn since chicken aggregated IgG (AgIgY), that do not bind FcRn, did not colocalize with AgIgG in GEC. AgIgG but not AgIgY induced a decrease of FcRn expression at the membrane and within the cells. The binding of AgIgG to GEC elicited a dose- and time-dependent increase in the release of uPA activity, as in the uPA protein and mRNA expression without modification in the release of PAI. This process was not abrogated by agents inhibiting endocytosis and/or transcytosis such as cytochalasin B, suggesting an endocytosis-independent uPA regulation. Conclusion: GEC response to AgIgG overload comprises at least two sequential steps: (1) a FcRn-mediated endocytosis; (2) an endocytosis-independent fibrinolytic imbalance leading to plasmin generation which could favor in vivo AgIgG clearance and matrix remodeling.

Endothelial adhesion molecules in glomerular lesions: Association with their severity and diversity in lupus models

To clarify whether vascular endothelial adhesion molecules in glomeruli might contribute to the severity and diversity of glomerular lesions in lupus nephritis, their expression in lupus models was analyzed. The expression levels of E- and P-selectin and vascular cell adhesion molecule-1 (VCAM-1) in glomeruli of different histopathologic grades of MRL/MpJ-lpr/lpr (MRL/lpr) lupus mice was studied using laser-capture microdissection of the glomeruli, followed by reverse transcription-polymerase chain reaction (RT-PCR) analysis. The glomerular lesions in SCID mice injected with the 2B11.3 and 7B6.8 clones, which are derived from an MRL/lpr mouse and induce endocapillary proliferative and wire loop type of glomerular lesions, respectively, were analyzed. To investigate the effect of a soluble form of E-selectin (sE-selectin) on the development of glomerular lesions, sE-selectin-producing L cells were prepared by transfection of the cDNA encoding sE-selectin and injected into SCID mice. The glomeruli in MRL/lpr mice showed increased expression of these adhesion molecules, corresponding to the severity of the glomerular lesions. The endocapillary proliferative type lesions in SCID mice induced by the 2B11.3 clone showed significantly increased expression of the adhesion molecules, especially E-selectin and P-selectin, but the wire loop type lesion induced by the 7B6.8 clone expressed only VCAM-1. Formation of the endocapillary proliferative type lesions induced by the 2B11.3 clone was markedly prevented in association with elevation of the serum level of sE-selectin produced by the tansfected L cells. The severity and diversity of the histopathology of lupus nephritis are partially associated with the expression of vascular endothelial adhesion molecules in glomeruli.

Downregulation of Bcl-2 by podocytes is associated with progressive glomerular injury and clinical indices of poor renal prognosis in human IgA nephropathy.

Bcl-2 defines a new class of proto-oncogenes that block cell death without promoting cell proliferation. To elucidate the role of Bcl-2 in the development of glomerular lesions in human IgA nephropathy (IgAN), we applied immunohistochemistry coupled with in situ hybridization to detect the expression of Bcl-2 products and their association with Bax, p27(kip1), and p57(kip2) in modulating the apoptotic, proliferative, and sclerotic events in progressive glomerular injury. Glomerular cell apoptosis was examined by TdT-mediated dUTP-biotin nick-end labeling (TUNEL) staining. A total of 51 IgAN cases were categorized into four subgroups (A to D) according to the severity of their histopathological lesions. Creatinine levels, creatinine clearance, and magnitude of proteinuria based on 24-h urine collections at the time of diagnostic renal biopsy were available for the majority of subjects. Bcl-2 expression was observed predominantly in podocytes in IgAN. Podocyte expression of Bcl-2 was found to be upregulated in early-stage disease and downregulated in late-stage disease. Bcl-2 downregulation in progressive IgAN was associated with an increased Bax/Bcl-2 ratio in glomerular epithelial cells and correlated with the downregulation of high endogenous podocyte p27(kip1) and p57(kip2) expression. Bax/Bcl-2 ratios positively correlated with glomerular cell apoptosis and the degree of glomerulosclerosis, whereas p27(kip1) and p57(kip2) expression levels were inversely correlated with mesangial hypercellularity and glomerulosclerosis. Clinicopathologic correlations demonstrated that downregulation of Bcl-2 protein expression was associated with indices of poor renal prognosis in human IgAN. The results suggest that Bcl-2 expression by podocytes may exert modulatory effects on cellular processes that contribute to progressive glomerular injury and play an important role in determining renal outcome in human IgA nephropathy.

Up-regulation of macrophage colony-stimulating factor (M-CSF) and migration inhibitory factor (MIF) expression and monocyte recruitment during lipid-induced glomerular injury in the exogenous hypercholesterolaemic (ExHC) rat.

Although macrophages play an important role in lipid-induced glomerular injury, we know little of the mechanisms by which hyperlipidaemia induces monocyte recruitment. This study investigated the role of M-CSF and macrophage MIF in monocyte recruitment during the development of lipid-induced glomerular injury in the susceptible ExHC rat strain. Groups of five ExHC rats were fed a high cholesterol diet (HCD) containing 3% cholesterol, 0.6% sodium cholate and 15% olive oil, and killed after 3 days, 1, 2 or 6 weeks. Control animals were killed on day 0 or after 6 weeks on a normal diet. Animals were hypercholesterolaemic 3 days after the induction of the HCD, but showed no change in plasma triglycerides over the 6-week period. Glomerular macrophage accumulation was first evident at 1-2 weeks and increased up to week 6, when macrophage-derived foam cells were seen in almost all glomeruli, and segmental lesions and mild proteinuria were also evident. Combined in situ hybridization and immunohistochemistry staining demonstrated that, coincident with the induction of hypercholesterolaemia on day 3, there was marked up-regulation of M-CSF and MIF mRNA expression by intrinsic glomerular cells (mostly mesangial cells and podocytes) which preceded monocyte recruitment. There was a highly significant correlation between the number of M-CSF and MIF-positive cells and glomerular macrophage accumulation over the 6-week period. Although some glomerular macrophages and foam cells exhibited M-CSF and MIF expression, the major source of these molecules was intrinsic glomerular cells. No local macrophage proliferation was observed during the development of glomerular lesions. In conclusion, hypercholesterolaemia caused marked up-regulation of M-CSF and MIF expression by intrinsic glomerular cells, which correlated with monocyte recruitment and the development of lipid-induced glomerular injury. This is the first study to implicate local synthesis of MIF in the pathogenesis of lipid-induced lesions.

Suppression of Experimental Membranous Glomerulonephritis in Rats by an Anti-MHC Class II Antibody

Background: We previously reported that idiopathic membranous nephropathy (IMN) strongly correlated with HLA-DRB1*1501-DRB5*0101-DQAI*0102-DQB1* 0602, a specific haplotype of human major histocompatibility complex (MHC), in Japanese patients. To investigate the role of MHC in the development of rat Heymann nephritis (HN), an animal model of membranous nephropathy, a monoclonal antibody (mAb) specific to rat MHC class II antigen (RT1B) was administered, and its effectiveness in inhibiting HN was assessed. Methods: Active HN was induced in HN-sensitive Lewis rats by administering brush border proteins of rat proximal uriniferous tubules (FX1A). Rats were divided into four groups: rats treated with 1,000 µg anti-rat MHC class II mAb, rats treated with 100 µg anti-rat MHC class II mAb, rats treated with murine myeloma IgG, and rats that did not receive either FX1A or any other mAb. We examined the differences in 24-hour urinary protein excretion and serum alloantibody titers against FX1A between groups at different time intervals, and the histologic features of kidneys at the end of the study. Results: HN was induced in Lewis rats by inoculation with FX1A antigen. Administration of anti-MHC class II mAb successfully lowered urinary proteins, production of anti-FX1A alloantibodies, and the development of glomerular lesions in a dose-dependent manner. Conclusion: The present results demonstrated that the MHC class II molecule itself is directly involved in the pathogenesis of HN, and suggest that this therapy would be any better (or less toxic) than nonselective immunosuppressants in the treatment of IMN.

Receptor mediated endocytosis by mesangial cells modulates transmigration of macrophages.

Accumulation of immune complexes in the mesangium is a common finding. Since migration of macrophages (Mphi) in the mesangium has been demonstrated to be an important event in the development of glomerular lesions, we studied the role of immune complexes and mesangial cell (MC) interaction in the transmigration (Tm) of Mphi. To determine the effect of MC and immune complexes (aggregated IgG, IgGAg) on transmigration of Mphi. MC were incubated with or without IgGAg in the lower compartment of a modified Boyden Chamber. To determine the effects of the secretory products (as a result of endocytosis of IgGAg by mesangial cells), MC-IgAg conditioned media was prepared and placed in the lower compartment of the Boyden chamber. We evaluated the effects of MC alone, MC + IgGAg, or MC-IgGAg conditioned media on the transmigration of macrophages across a filter. To determine the effect of free radicals on MC-IgAg endocytosis-induced Mphi migration we evaluated the effect of free radical scavengers such as dimethyl thiourea (DMTU) and tetramethylthiourea (TMTU) in MC-IgAg endocytosis-induced Mphi migration. To determine the role of chemokines in MC-IgAs endocytosis-induced Mphi migration we evaluated the effect of ani-MCP-1 antibodies on MC-IgAg endocytosis-induced Mphi migration, and also studied the effects of IgAg on MC mRNA expression of MCP-1 and RANTES. In addition, we evaluated the role of Fc receptors and actin cytoskeleton of MC in transmigration of Mphi. Mesangial cell endocytosis of IgG aggregates (IgGAg) is associated with enhanced (P < 0.001) transmigration of Mphi (control, 11.2 +/- 0.2 vs. MC + IgGAg, 22.1 +/- 0.9 migrated Mphi/field). IgGAg also induced MC mRNA expression for RANTES and MCP-1 on MC. DMTU and TMTU attenuated (P < 0.001) the MC + IgGAg-induced migration of Mphi as well as IgGAg-induced mRNA expression for RANTES and MCP-1. MC and IgGAg interaction products (MC-IgGAg conditioned media) also increased (P < 0.01) transmigration of Mphi (control, 18.3 +/- 1.7 vs. MC-IgGAg conditioned media, 30.7 +/- 0.6 Mphi/field). This effect of MC-IgGAg conditioned media on the migration of macrophages was dose dependent. Anti-MCP-1 antibody partially inhibited MC-IgGAg-induced migration of macrophages. MC and monomeric IgG (MIgG) interaction (MC-MIgG conditioned media) showed a lower (P < 0.05) migration of Mphi, when compared to the MC-IgGAg conditioned media. MC-IgGAg conditioned media prepared from cytochalasin B pretreated MCs also showed a lower (P < 0.001) migration of Mphi when compared with MC-IgGAg conditioned media-induced migration. These results indicate that MC-IgGAg conditioned media-induced transmigration of macrophages may be mediated through the generation of RANTES and MCP-1 by MC.

Diabetes--renal function--what are the special problems?

Diabetic nephropathy has become the single most important cause of endstage renal failure in most countries of the Western world. Against this background, the role of the renin-angiotensin system (RAS) and its blockade command considerable interest. In diabetic patients and in diabetic animals, the circulating components of the RAS are suppressed. Although the evidence is not completely uniform, there are indirect arguments (renal hemodynamic response to RAS blockade, AT1 receptor expression), however, which would be consistent with increased intrarenal action of angiotensin (ANG) II. There is solid evidence that ACE inhibitors effectively interfere with progression of micro-albuminuria both in IDDM and NIDDM. They also prevent progression of advanced renal failure in IDDM, while there is only preliminary evidence in this respect for NIDDM. ACE inhibitors are superior to conventional antihypertensive agents (with the possible exception of some calcium channel blockers), but such superiority is seen only when the levels of blood pressure are relatively high. In diabetic animals, treatment with ANG II receptor blockers interferes with the development of glomerular lesions. In acute and subacute studies on diabetic patients, ANG II receptor blockers reduced albuminuria (or proteinuria) more than beta-blockers. Head-on comparison of equipotent doses ACE inhibitors and ANG II receptor blockers in non-diabetic patients produced equal reductions in proteinuria. The long-term effects of ANG II receptor blockers on progression of advanced diabetic nephropathy is the object of two large international studies. The results will not be available before the year 2000.

Fosinopril ameliorates exogenous cholesterol-induced incipient glomerular lesions in obese Zucker rats. Effects on eicosanoid secretion

To date, the role of dietary cholesterol as a risk factor for some diabetic nephrophathy, such as mesangial expansion and glomerular lesions, is unknown. Controversy also exists regarding the effects of prostaglandin-induced changes in glomerular haemodynamics on the appearance of glomerulosclerosis. We have used obese Zucker rats (OZRs) as a model of early nephrophathy to evaluate the effect of hypercholesterolaemic diet on glomerular prostaglandin secretion and on the development of glomerular lesions. Due to the role of angiotensin II (Ang II) in glomerular haemodynamics, we have also evaluated its effects on glomerular eicosanoid secretion. Furthermore, as it has been suggested recently by clinical studies that angiotensin-converting enzyme inhibitors (ACEIs) reduce serum lipids associated with proteinuria, we have also evaluated the effect of the ACEI, fosinopril, both in vivo and in vitro, using 24 h glomeruli cultures. Results showed that a cholesterol-rich diet significantly increased serum cholesterol, proteinuria and glomerular eicosanoid secretion, and caused macrophage-ED1 cell deposits in the glomerular mesangium. Segmentary lesions only appeared in those rats with the highest percentage of glomerular xanthomatous (macrophage-ED1) cells. Ang II, per se, caused a marked rise in glomerular prosaglandin E2 and thromboxane B2. The inhibition of Ang II synthesis with fosinopril reduced all the parameters listed above, whereas Ang II (10(-6)M) increased the secretion of TxB2 and tended to increase PGE2 secretion in glomerular culture. In conclusion, exogenous cholesterol per se may contribute to nephropathy by increasing eicosanoid secretion, serum lipid profile, urinary protein excretion and the development of glomerular lesions. Fosinopril reduced all these parameters probably by its effects on Ang II.

Adaptation and distortion of podocytes in rat remnant kidney.

A sequence of alterations of podocytes as well as the development of glomerulosclerosis was analyzed in a five-sixths nephrectomy rat model. Morphometric measurements showed a gradual enlargement in glomerular size without any increase in the number of podocytes in the glomerulus. Structural changes of podocytes appeared as early as 4 weeks after operation, and increased their degree and extent with time. In electron microscopic studies, changes of podocytes in the early stage consisted of foot process stretching and thinning, and cell body attenuation accompanied by pseudocyst formation. As weeks passed, these changes became prominent and were occasionally associated with podocyte detachment from the glomerular basement membrane followed by tuft adhesion and the development of segmental sclerosis. In parallel to the structural changes, immunohistochemical staining demonstrated a marked increase of desmin expression in the glomerulus with intense staining in podocytes. Podocyte alterations were identified as an adaptation to the enlargement of the glomerulus and, consequently, the podocytes seemed to fall to adapt to the structural changes of the glomerulus. The present study supports the hypothesis that podocyte defects are responsible for both the initiation and progression of glomerular damage. Furthermore, the findings may provide the hypothesis that the native disposition of podocytes in the glomerulus participates in the development of glomerular lesions in a focal and segmental fashion.

Hepatitis C and glomerulonephritis

Summary: Several systemic viral infections have been associated with the development of glomerular lesions. of the viruses that cause liver disease hepatitis B was the first to be recognized. the recent availability of serologic tests for the hepatitis C virus (HCV) has uncovered an association between HCV infection and renal disease. the principal glomerular lesion that develops is that of a membranoproliferative glomerulonephritis (MPGN), usually in association with antigenaemia and circulating immune complexes that have the characteristics of mixed cryoglobulinaemia represented by polyclonal IgG and monoclonal IgM with rheumatoid factor activity. the presence of a very high percentage of anti‐HCV seropositivity in cryoglobulinaemic forms of MPGN suggests that the virus plays an important role in the pathogenesis of the associated immune complex glomerulonephritis. Precipitates containing HCV‐RNA and circulating anti‐HCV IgG and IgM have been found in the majority of such cases. the course of the renal disease that develops is progressive. Treatment with interferon alpha appears to attenuate the progression of the renal lesions, and the response to treatment appears to be closely related to the clearance of hepatitis C viraemia. Renal lesions also occur in the absence of clinical evidence of liver disease or mixed cryoglobulinaemia. In addition to MPGN, membranous glomerulonephropathy, IgA nephropathy and focal segmental glomerulosclerosis have been reported in these cases of HCV infection. the prevalence of glomerular lesions in patients with HCV infection remains to be determined. the available serologic tests for HCV are still in evolution. In the meantime, all patients presenting with glomerular disease should be screened for HCV.