Epidemiology and referral patterns of patients living with chronic kidney disease in Johannesburg, South Africa: A single centre experience.

Chronic kidney disease (CKD) is a significant contributor to the global burden of non-communicable disease. Early intervention may facilitate slowing down of progression of CKD; recognition of at-risk patient groups may improve detection through screening. We retrospectively reviewed the clinical records of 960 patients attending a specialist nephrology outpatient clinic during the period 1 January 2011-31 December 2021. A significant proportion (47.8%) of patients were referred with established CKD stage G4 or G5. Non-national immigration status, previous diagnosis with diabetes, and advancing age were associated with late referral; antecedent diagnosis with HIV reduced the odds of late referral. Black African patients comprised most of the sample cohort and were younger at referral and more frequently female than other ethnicities; non-nationals were younger at referral than South Africans. Hypertension-associated kidney disease was the leading ascribed aetiological factor for CKD (40.7% of cases), followed by diabetic kidney disease (DKD) (19%), glomerular disease (12.5%), and HIV-associated kidney disease (11.8%). Hypertension-related (25.9%) and diabetic (10.7%) kidney diseases were not uncommon in people living with HIV. Advancing age and male sex increased the likelihood of diagnosis with hypertensive nephropathy, DKD and obstructive uropathy; males were additionally at increased risk of HIV-associated kidney disease and nephrotoxin exposure, as were patients of Black African ethnicity. In summary, this data shows that hypertension, diabetes, and HIV remain important aetiological factors in CKD in the South African context. Despite the well-described risk of CKD in these disorders, referral to nephrology services occurs late. Interventions and policy actions targeting at-risk populations are required to improve referral practices.

Thrombotic thrombocytopaenic purpura in the era of HIV: A single-centre experience.

Background: Thrombotic thrombocytopaenia purpura (TTP) is a rare disorder which carries a high mortality. HIV is an important cause of TTP. Objectives: We assessed the presentation and response to plasma exchange (PEX) by HIV status. Method: A single-centre retrospective review of all patients receiving PEX for TTP between 01 January 2010 and 31 December 2019 was undertaken. Demographics and presenting parameters were compared between HIV-associated TTP and other aetiologies using Mann-Whitney U and Kruskal Wallis analysis of variance testing, as appropriate. The effect of aetiology and presenting parameters on PEX duration was modelled using Cox proportional hazards; effect of these variables on mortality and residual renal dysfunction in survivors was analysed using stepwise multivariate regression. Results: Uncontrolled HIV infection was the commonest cause (81.9%) of TTP in the 83 patients identified. Thrombocytopaenia was more severe and neurological deficit more frequent in HIV-associated TTP; but renal dysfunction was milder in this group. Aetiology did not influence mortality risk. Aetiological category and presenting parameters did not predict PEX duration. Residual renal dysfunction was less frequent in survivors of HIV-associated TTP. Conclusion: HIV is an important cause of TTP in the local context. Haematological and neurological involvement are more severe in HIV-associated TTP. Acceptable survival rates are achievable with PEX even in advanced HIV infection; renal sequalae are less common in this group.

Survival outcomes of HIV-positive adults on peritoneal dialysis at Helen Joseph renal unit.

Background: HIV is a risk factor for the development of chronic kidney disease. People with chronic kidney disease in the state sector are likely to be prescribed continuous ambulatory peritoneal dialysis (CAPD). Previous studies have raised concern about the safety of CAPD in people living with HIV (PLWH) compared to HIV-negative patients. Objectives: To compare the risk of peritonitis, and modality and patient survival by HIV status in patients receiving CAPD at Helen Joseph Hospital. Method: A retrospective study of patients receiving CAPD between 01 January 2007 and 31 December 2017 was undertaken. Five-year patient and modality survival were modelled for PLWH and HIV-negative subgroups and analysed using the log-rank test; the effect of CD4 count, HIV viral load, and duration of antiretroviral therapy on these parameters in PLWH were additionally modelled using the Cox Proportional Hazards technique. Results: Eighty-four patients, comprising of 21 PLWH and 63 HIV-negative patients, were analysed. No difference was observed in the proportion of patients who had at least one episode of peritonitis between PLWH (61.2%) and HIV-negative patients (63.5%) (P = 0.547). A trend towards increased risk of peritonitis due to Gram-negative organisms in PLWH was noted (odds ratio: 3.20, 95% confidence interval: 0.86-11.9, P = 0.083). No difference was observed in 5-year patient or modality survival on CAPD between PLWH (log-rank P = 0.161) and HIV-negative patients (log-rank P = 0.240). Conclusion: People living with HIV should not be excluded from CAPD as a mode of kidney replacement therapy (KRT).

Comparison of quality of life in patients with advanced chronic kidney disease undergoing haemodialysis, peritoneal dialysis and conservative management in Johannesburg, South Africa: a cross-sectional, descriptive study.

INTRODUCTION: Mental health and quality of life are under-appreciated clinical targets which affect patient and modality survival. Lack of dialysis availability in the resource-constrained public health sector in South Africa results in assignment to treatment modalities without regard to effects on these parameters. We assessed the effect of dialysis modality, demographic and laboratory parameters on mental health and quality of life measurements. METHODS: Size-matched cohorts were recruited from patients on haemodialysis (HD), peritoneal dialysis (PD), and patients on conservative management (CM) between September 2020 and March 2021. Responses to the Hospital Anxiety and Depression Scale (HADS) and Kidney Disease Quality of Life Short Form 36 (KDQOL-SF36) questionnaires and demographic and baseline laboratory parameters were compared between modalities. Multivariate linear regression was used to evaluate independent effect of baseline characteristics on HADS and KDQOL-SF36 scores between treatment groups where significant difference was observed. RESULTS: Anxiety, depression, and reduced KDQOL measures were widespread amongst respondents. Dialyzed patients reported higher anxiety and depression scores than those on CM (p = 0.040 and p = 0.028). Physical composite (PCS), role-physical (RP), vitality (VS), and emotional well-being (EWB) KDQOL-SF36 scores were poorer in dialyzed patients (p < 0.001 for all). PCS (p = 0.005), pain (p = 0.030), vitality (p = 0.005), and social functioning KDQOL scores were poorer in PD compared to HD; HADS anxiety (p < 0.001) and KDQOL-SF36 EWB scores (p < 0.001) were better in PD. PD patients were more likely to be employed (p = 0.008). Increasing haemoglobin concentration reduced anxiety (p < 0.001) and depression scores (p = 0.004), and improved PCS (p < 0.001), and pain scores (p < 0.001). Higher serum albumin improved PCS (p < 0.001) and vitality (p < 0.001) scores. CONCLUSION: Advanced chronic kidney disease increases anxiety and depression and limits quality of life. PD improves mental health and emotional wellbeing and preserves the ability to undertake economic activity but limits social functioning and causes greater physical discomfort. Targeting haemoglobin may ameliorate modality effects on mental health and quality of life.

Clinicopathological correlation of kidney disease in HIV infection pre- and post-ART rollout.

The spectrum of HIV-associated kidney disease has expanded significantly with the introduction of antiretroviral therapy (ART). In the pre-ART era there was prominence of HIV-associated nephropathy (HIVAN). More recently, the spectrum of disease additionally reflects comorbid illness in the ageing HIV population and ART-related nephrotoxicity. We performed a clinicopathological correlation of kidney disease in HIV-positive individuals who underwent kidney biopsy between 1989 and 2014, utilizing the 2018 Kidney Disease Improving Global Outcomes pathologic classification. ART rollout began in 2004 in South Africa. Patients biopsied pre-ART rollout were compared to those biopsied post-ART rollout with respect to demographics, clinical parameters and histology. We assessed kidney survival in a cohort of these patients following biopsy. Six hundred and ninety biopsies were included, 99 (14.3%) were undertaken pre- and 591 (85.7%) post-ART rollout. Most patients were of Black African descent (97.5%). The post-ART rollout patients were older (p = 0.007), had higher eGFR at presentation (p = 0.016) and fewer presented with eGFR of less than 15ml/min/1.73m2 (p = 0.0008). There was a decrease in the prevalence of classic HIVAN (p = 0.00001); and an increase in FSGS (NOS) in the setting of HIV (p = 0.0022) and tubulointerstitial diseases (p = 0.009) post-ART rollout. Kidney function survival over 5 years was poorest in patients with classic HIVAN (p = 0.00005) and best in minimal change nephropathy (p = 0.0013). Kidney biopsy is crucial for the correct diagnosis and management of HIV-related kidney disease. ART rollout has shifted the spectrum of kidney disease away from classic HIVAN but has not eliminated it. Histological diagnosis prognosticates kidney survival.

Living kidney donation in a developing country.

BACKGROUND: Living kidney donation has been advocated as a means to ameliorate the chronic shortage of organs for transplantation. Significant rates of comorbidity and familial risk for kidney disease may limit this approach in the local context; there is currently limited data describing living donation in Africa. METHODS: We assessed reasons for non-donation and outcomes following donation in a cohort of 1208 ethnically diverse potential living donors evaluated over a 32-year period at a single transplant centre in South Africa. RESULTS: Medical contraindications were the commonest reason for donor exclusion. Black donors were more frequently excluded (52.1% vs. 39.3%; p<0.001), particularly for medical contraindications (44% vs. 35%; p<0.001); 298 donors proceeded to donor nephrectomy (24.7%). Although no donor required kidney replacement therapy, an estimated glomerular filtration rate below 60 ml/min/1.73 m2 was recorded in 27% of donors at a median follow-up of 3.7 years, new onset albuminuria >300 mg/day was observed in 4%, and 12.8% developed new-onset hypertension. Black ethnicity was not associated with an increased risk of adverse post-donation outcomes. CONCLUSION: This study highlights the difficulties of pursuing live donation in a population with significant medical comorbidity, but provides reassurance of the safety of the procedure in carefully selected donors in the developing world.

Quality of life in patients on continuous ambulatory peritoneal dialysis in an African setting.

To determine the quality of life (QOL) of patients on continuous ambulatory peritoneal dialysis (CAPD), we studied all the CAPD patients attending their monthly follow-up care clinics at three tertiary hospitals in Johannesburg by administering the World Health Organization QOL-Bref questionnaire. The patients were grouped according to age, duration of peritoneal dialysis and gender. Data were analyzed to determine the significant differences in the QOL scores among the subgroups. There were 114 patients [64 males (56.1%), with a mean age of 42.4 ± 11.3 years) and 38 healthy control subjects (22 males (57.9%), with a mean age of 42.1 ± 12.4 years]. Twenty-one patients (18.4%) had hemoglobin <10 g/dL, while 16 patients (14%) had serum albumin <3 g/dL. The mean QOL scores in the physical, psychological, social relationships and environment domains of the CAPD patients were 55.7 ± 15.0, 56.6 ± 16.4, 55.3 ± 24.7 and 56.3 ± 16.6, respectively. The CAPD patients had significantly lower QOL scores compared with controls, and those aged <30 years had better scores in the physical and psychological domains, gender and hemoglobin concentration. Serum albumin levels did not have a significant impact on the QOL of the CAPD patients.

Hyaluronan induces the selective accumulation of matrix- and cell-associated proteoglycans by mesangial cells.

Mesangial cells (MCs) are essential for normal renal function through the synthesis of their own extracellular matrix, which forms the structural support of the renal glomerulus. In many renal diseases this matrix is reorganized in response to a variety of cytokines and growth factors. This study examines proteoglycan and hyaluronan (HA) synthesis by MCs triggered by proinflammatory agents and investigates the effect of an exogenous HA matrix on matrix synthesis by MCs. Metabolic labeling, ion exchange and size exclusion chromatography, Western blotting, and immunocytochemistry were used to identify changes in matrix accumulation. When incubated with interleukin-1, platelet-derived growth factor, or fetal calf serum, MCs initiated rapid HA synthesis associated with the up-regulation of HA synthase-2 and increased the synthesis of versican, perlecan, and decorin/biglycan. HA was both released into the medium and incorporated into extensive pericellular coats. Adding exogenous HA to unstimulated cells that had undetectable pericellular coats of HA selectively reduced perlecan and versican turnover, whereas other proteoglycans were unaffected. These results suggest that high levels of HA in the mesangium in disease is a mechanism controlling the accumulation of specific mesangial matrix components. HA may thus be an attractive target for therapeutic intervention.

Catabolism of newly synthesized decorin in vitro by human peritoneal mesothelial cells.

OBJECTIVE: Previous studies have shown that decorin and biglycan account for over 70% of the proteoglycans (PGs) synthesized by human peritoneal mesothelial cells (HPMCs). Since these PGs are involved in the control of cell growth, cell differentiation, and matrix assembly, we investigated their turnover in cultured HPMCs. METHODS: Confluent HPMCs were metabolically labeled with [35S]-sulfate and the labeled products isolated from the cell medium and the cell layer characterized by sensitivity to bacterial eliminases. Experiments were undertaken with exogenous labeled decorin, and its metabolic state was studied. RESULTS: In a 24-hour labeling period, 75% of the newly synthesized chondroitin sulfate/dermatan sulfate (CS/DS) PGs appeared in the culture medium, the majority of which (90%) was decorin. In the cell layer, protein-free glycosaminoglycan (GAG) chains accounted for 21% of the total CS/DS at 24 hours and exhibited constant specific activity at 12-16 hours. The latter material was turned over with a half-life of approximately 2.5 hours. Exogenous decorin underwent receptor-mediated endocytosis and subsequent intracellular degradation. Uptake but not degradation could be inhibited by heparin. CONCLUSIONS: HPMCs are distinguished by a rapid turnover of decorin. A characteristic metabolic feature is the existence of a large intracellular pool of protein-free DS-GAGs. Understanding the control of decorin turnover in HPMCs might lead to delineation of its potential role in both the physiology and pathophysiology of the membrane in PD patients.

Identification and analysis of the promoter region of the human hyaluronan synthase 2 gene.

Hyaluronan (HA) is a linear glycosaminoglycan of the vertebrate extracellular matrix that is synthesized at the plasma membrane by the HA synthase (HAS) enzymes HAS1, -2 and -3. The regulation of HA synthesis has been implicated in a variety of extracellular matrix-mediated and pathological processes, including renal fibrosis. We have recently described the genomic structures of each of the human HAS genes. In the present study, we analyzed the HAS2 promoter region. In 5'-rapid amplification of cDNA ends analysis of purified mRNA from human renal epithelial proximal tubular cells, we detected an extended sequence for HAS2 exon 1, relocating the transcription initiation site 130 nucleotides upstream of the reference HAS2 mRNA sequence, GenBank accession number NM_005328. A luciferase reporter gene assay of nested fragments spanning the 5' terminus of NM_005328 demonstrated the constitutive promoter activity of sequences directly upstream of the repositioned transcription initiation site but not of the newly designated exonic nucleotides. Using reverse transcription-PCR, expression of this extended HAS2 mRNA was demonstrated in a variety of human cell types, and orthologous sequences were detected in mouse and rat kidney. Alignment of human, murine, and equine genomic DNA sequences upstream of the repositioned HAS2 exon 1 provided evidence for the evolutionary conservation of specific transcription factor binding sites. The location of the HAS2 promoter will facilitate analysis of the transcriptional regulation of this gene in a variety of pathological contexts as well as in developmental models in which HAS2 null animals have an embryonic lethal phenotype.

Glucose degradation products (GDP) retard remesothelialization independently of D-glucose concentration.

BACKGROUND: Glucose degradation products (GDP) present in heat-sterilized dialysis fluids are thought to contribute to cellular dysfunction and membrane damage during peritoneal dialysis. To examine the effects of specific GDP on the remesothelialization process, the impact of conventional and low GDP peritoneal dialysis solutions, D-glucose, and individual GDP in a scratch-wounding model was assessed. METHODS: Scratch (0.5 to 0.6 mm)-wounded human peritoneal mesothelial cells (HPMC) were treated, at pH 7.4, with either (1) control medium (M199), (2) laboratory-prepared heat or filter-sterilized solutions, (3) 10% to 80% vol/vol solution of Gambrosol or Gambrosol-trio (1.5% and 4.0% glucose), (4) D-glucose (5 to 80 mmol/L), or (5) individual or combined GDP [acetaldehyde, formaldehyde, glyoxal, methylglyoxal, 3-deoxyglucosone (3-DG), 5-hydroxy methylfufural (5-HMF), or 3,4-di-deoxyglucosone-3-ene (3,4-DGE)]. Wound closure was recorded by time-lapse photomicroscopy. RESULTS: In untreated HPMC, the rate of wound closure was linear and the process was complete by 18.4 +/- 3.6 hours (N = 16). In wounded HPMC exposed to dilutions of heat-sterilized but not filtered laboratory solutions (1.5% or 4.0% glucose, pH 7.4), remesothelialization was significantly retarded (P = 0.04 and P = 0.009 vs. M199, respectively). In Gambrosol, remesothelialization was significantly retarded in both 1.5% and 4.0% solutions. In contrast in Gambrosol-trio-treated HPMC, this rate was not significantly reduced in either 1.5% or 4.0% glucose peritoneal dialysis fluids. Remesothelialization was dose-dependently retarded in HPMC exposed to 3,4-DGE (>10 microl/L), formaldehyde (>5 micromol/L) but not by exposure to the other GDP tested even at 5 times the concentration present in low glucose solutions. The rate of remesothelialization was not significantly altered by exposure to D-glucose concentrations up to 80 mmol/L. CONCLUSION: These data identify that the formaldehyde and 3,4-DGE present in heat-sterilized peritoneal dialysis solutions are important in reducing mesothelial cell regeneration. Specifically targeting their removal may have major benefits in preserving the mesothelium during long-term peritoneal dialysis.

The human hyaluronan synthase genes: genomic structures, proximal promoters and polymorphic microsatellite markers.

The glycosaminoglycan (GAG) hyaluronan (HA) is a key component of the vertebrate extracellular matrix (ECM) and is synthesised by the HA synthase (HAS) enzymes HAS1, HAS2 and HAS3 at the plasma membrane. Accumulating evidence emphasises the relevance of HA metabolism in an increasing number of processes of clinical interest including renal fibrosis and peritoneal mesothelial wound healing. In the present study, the genomic sequences and organisation of the genes encoding the human HAS isoforms were deduced, in silico, from reference cDNA and genomic sequence data. These data were confirmed in vitro by sequencing of PCR-amplified HAS exons and flanking genomic sequences, comparison with sequence data for the corresponding murine Has orthologues, rapid amplification of 5' cDNA ends analysis and luciferase reporter assays on putative proximal promoter sequences. The HAS1 gene comprised five exons, with the translation start site situated 9bp from the 3' end of exon 1. In contrast, the genomic structures for HAS2 and both HAS3 variants spanned four exons, exon 1 forming a discrete 5'-untranslated region (5'-UTR) and the translation start site lying at nucleotide 1 of exon 2. Dinucleotide microsatellite loci were identified in intron 1 of HAS1 and HAS2, and immediately upstream of the HAS3 gene and their utility as linkage markers demonstrated in genomic DNA (gDNA) studies. We thus present a comprehensive resource for mutation detection screening of all HAS exons and/or linkage analysis of each HAS gene in a variety of disorders for which they are attractive candidates.

Structural and functional changes in heparan sulfate proteoglycan expression associated with the myofibroblastic phenotype.

The principal cells implicated as the source of the extracellular matrix in areas of progressive fibrosis are fibroblasts with the phenotypic appearance of myofibroblasts. This report describes differences in heparan sulfate proteoglycan expression between myofibroblasts and normal fibroblasts, associated with impaired responses to fibroblast growth factor-2 (FGF-2). Although both cell types responded to platelet-derived growth factor, myofibroblasts, unlike fibroblasts, did not proliferate to FGF-2. A response was acquired, however, when myofibroblasts were incubated with FGF-2 in the presence of heparan sulfate (HS) and heparin. Selective digestion with pronase, NaOH/NaBH(4), heparinase I, or low pH nitrous acid showed that each HS-glycosaminoglycan region comprised a pronase-resistant peptide separating two HS chains. The HS-glycosaminoglycan chains from myofibroblasts were larger (K(av), 0.32; molecular weight, 50 kd) than those from fibroblasts (K(av), 0.4; molecular weight, 33 kd), although their disaccharide composition was identical. The chains from myofibroblasts, however, contained three, compared to two, heparinase 1-resistant sequences separated by larger contiguous areas of low sulfation. Furthermore, although there was no difference in FGF-2-binding affinity between the two cell types, the chains secreted by myofibroblasts had twice the binding capacity of those from fibroblasts. Thus, it is likely that the difference in response to FGF-2 is because of a difference in FGF-2 sequestration and receptor interaction with FGF-2-HS complexes. A comparative investigation into HS fine structure is being undertaken to examine these findings in more detail.

Involvement of human heparanase in the pathogenesis of diabetic nephropathy.

BACKGROUND: Decreased heparan sulfate proteoglycan content of the glomerular basement membrane has been described in proteinuric patients with diabetic nephropathy. Heparanase is an endo-beta-D-glucuronidase that cleaves negatively charged heparan sulfate side chains in the basement membrane and extracellular matrix. OBJECTIVES: To investigate whether urine from type I diabetic patients differs in heparanase activity from control subjects and whether resident glomerular cells could be the source of urinary heparanase. METHODS: Using soluble 35S-HSPG and sulfate-labeled extracellular matrix we assessed heparanase activity in human glomerular epithelial cells, rat mesangial cells, and urine from 73 type I diabetic patients. Heparanase activity resulted in the conversion of a high molecular weight sulfate-labeled HSPG into heparan sulfate degradation fragments as determined by gel filtration analysis. RESULTS: High heparanase activity was found in lysates of both epithelial and mesangial cells. Immunohistochemical staining localized the heparanase protein to both glomeruli capillaries and tubular epithelium. Heparanase activity was detected in the urine of 16% and 25% of the normoalbuminuric and microalbuminuric diabetic patients, respectively. Urine from 40 healthy individuals did not possess detectable heparanase. Urinary heparanase activity was associated with worse glycemic control. CONCLUSION: We suggest that heparanase enzyme participates in the tunover of glomerular HSPG. Hyperglycemia enhances heparanase activity and/or secretion in some diabetic patients, resulting in the loss of albumin permselective properties of the GBM.

The role of ADAM 15 in glomerular mesangial cell migration.

Mesangial cells (MC) occupy the core of the renal glomerulus and are surrounded by a mesangial matrix. In certain diseases, MC migrate through this matrix into the pericapillary space. The mechanisms involved, however, are poorly understood. Members of the ADAM (A Disintegrin And Metalloproteinase) family of membrane proteins have the potential to be key modulators of cell-matrix interactions through the activities of their constituent domains. We have studied the possible role of ADAM 15 in human (H) MC migration in vitro. HMC ADAM 15 was expressed at low levels in serum-free medium but was increased during migration. Antibodies to the individual domains of ADAM 15 and the incorporation of antisense ADAM 15, (but not control oligonucleotide) inhibited this migration. Furthermore, inhibition of migration by the broad spectrum metalloproteinase inhibitor BB3103, demonstrated that metalloproteinase activity was essential for migration. ADAM 15, extracted from HMC membranes, was an active metalloproteinase, which degraded both type IV collagen and gelatin prepared from fibrillar collagen. Activity was inhibited by EDTA but not by phenylmethylsulfonyl fluoride. This is the first report of the potential of ADAM 15 for involvement in the restructuring of the mesangial matrix and in the migration of MC in disease.

Goto-Kakizaki rat is protected from proteinuria after induction of anti-Thy1 nephritis.

Hyperglycemia, although necessary, alone is insufficient for the development of progressive diabetic nephropathy. Two factors implicated in its pathogenesis are mesangial cell activation and/or proliferation and monocyte/macrophage influx. We have shown that prolonged hyperglycemia in the Goto-Kakizaki (GK) rat is associated with renal structural changes similar to those in patients with diabetes before the onset of progressive nephropathy. The aim of the current study is to examine the role of mesangial cell injury and macrophage influx on renal structure and function. After induction of nephritis in either hyperglycemic GK rats or normoglycemic Wistar rats by the administration of Ox-7 antibody, the degree of mesangiolysis and subsequent mesangial proliferation was no different between GK and Wistar rats. Similarly, macrophage influx and mesangial cell activation (assessed by alpha-smooth actin expression) was no different between the two groups. Wistar rats developed marked albuminuria; conversely, no significant proteinuria or albuminuria was seen in GK rats. Analysis of glomerular proteoglycans (PGs) showed an increase in (35)S incorporation into heparan sulfate PGs of GK compared with Wistar rats, with no alteration in glycosaminoglycan chain size or charge density. These changes were kidney specific and not seen in spleen, lung, or heart tissue. Western blot analysis showed increased agrin core protein expression in whole-kidney homogenates of untreated GK rats. Induction of Thy1.1 nephritis was associated with reduced expression of agrin in both GK and Wistar rats. However, agrin expression was greater in GK rats at all times. In summary, acute mesangial cell injury associated with a macrophage influx did not initiate progressive diabetic nephropathy in GK rats. Despite a similar magnitude of glomerular/mesangial injury, GK rats, in contrast to normoglycemic Wistar rats, did not develop proteinuria after the administration of anti-Thy1 antibody. We postulate that altered expression of agrin in this model accounts for the lack of proteinuria and thus may protect against progressive nephropathy.

Induction of metalloproteinases by glomerular mesangial cells stimulated by proteins of the extracellular matrix.

Human glomerular mesangial cells (HMC) are embedded in the mesangial matrix (MM) and control its turnover through a dynamic equilibrium between synthesis and degradation. Degradation is controlled by matrix metalloproteinases (MMP), whose activity has been causally implicated in the progression of glomerular disease. In other systems, MMP secretion may be directly affected by exposure to specific matrix proteins. The present study, therefore, investigated the effect of different matrix components on the adherence of HMC and on their secretion and activation of the gelatinases MMP2 and MMP9. HMC adhered strongly (quantified using crystal violet staining) to collagen IV and collagen I (P < 0.01, relative to binding to control, bovine serum albumin (BSA)-coated wells) and to a lesser extent to gelatin IV and fibronectin (P < 0.05). Binding to vitronectin and laminin was not statistically different to control wells. After the addition of these matrix proteins (0.1 microg/ml to 100 microg/ml) to growth-arrested HMC for 72 h, zymography of the conditioned medium established that only fibronectin and collagens I and IV dose-dependently increased latent (72 kD) MMP2 secretion and activation. Fibronectin, however, also induced the secretion of MMP9. Membranes from HMC that had been co-cultured with fibronectin for 72 h were prepared to investigate whether the activation of MMP2 in this system was due to the action of membrane-type (MT)-MMP. When incubated with latent MMP2 for times up to 24 h, these membranes activated the enzyme in a time- and dose-dependent manner. The results demonstrate that specific matrix components increased the secretion of MMP2 and MMP9 from HMC. In addition, MT-MMP activity, selectively induced by fibronectin, was implicated in the activation of the secreted proteinases.