Glomerular disease and acute kidney injury in Sudan: Demographics, histological diagnosis and outcome.

BACKGROUND: Acute kidney injury (AKI) is a relatively common clinical condition, associated with high rates of mortality. Although there is extensive literature on the nature and consequence of AKI in the developed world, much less is known in the developing world and more specifically in sub-Saharan Africa (SSA). OBJECTIVES: To describe the demographics, histological diagnosis and clinical course of patients presenting with AKI to a single centre in Sudan. METHODS: Retrospective data were collected on 100 consecutive patients with AKI and an active urinary sediment, who underwent diagnostic native percutaneous renal biopsy. RESULTS: The mean (standard deviation) age of those biopsied was 33.6 (14.1) years of age, with a preponderance (58%) of females. The most common clinical indications for biopsy were AKI associated with haematuria and proteinuria (72%), AKI and proteinuria (22.5%) or AKI and haematuria (5%). The frequencies of the most common primary glomerulonephritides (GN) were focal segmental GN (15%) and mesangiocapillary GN (8%). Lupus nephritis was the most frequent secondary GN associated with AKI (31%) and the most common overall histological diagnosis. Peak creatinine, but not oliguria, at presentation predicted likelihood of remaining dialysis-dependent. Age at presentation but not baseline renal function by estimated glomerular filtration rate (eGFR), was associated with the likelihood of having residual chronic kidney disease following an episode of AKI. CONCLUSIONS: The data suggested differences in the pattern of intrinsic renal/glomerular disease leading to AKI to those published and mainly derived from the developed world and patients in SSA.

Acute kidney injury risk factor recognition in three teaching hospitals in Ethiopia.

BACKGROUND: A key objective of the Nephrology Sister Centre Programme between the renal units in Cardiff and Addis Ababa, sponsored by the International Society of Nephrology, is to facilitate development of the local clinical service in Ethiopia specifically focused on the management of acute kidney injury (AKI). OBJECTIVES: To examine the relationship between AKI risk factor recognition and monitoring of renal function in three hospitals in Ethiopia. METHODS: Cross-sectional data were gathered regarding renal function monitoring, recording the presence of AKI risk-associated comorbidities and prescription of nephrotoxic medications across the disciplines of medicine, surgery, obstetrics and gynaecology. Results. Patients were more likely to have their renal function checked at the hospital with specialist services. Across all centres, the highest proportion of patients who had renal function measurements were those admitted to a medical ward. There was a positive relationship between documented comorbidities and the measurement of renal function but not between the prescription of nephrotoxic drugs and measurement of renal function. CONCLUSION: There was great variability in the extent to which doctors recognised the presence of risk factors for the development of AKI. Failure to identify these risk factors represents a lost opportunity to identify patients at high risk of developing renal injury who would benefit from renal function monitoring.

Inhibition of TGF-ß1/Smad signal pathway is involved in the effect of Cordyceps sinensis against renal fibrosis in 5/6 nephrectomy rats.

UNLABELLED: The present study aimed to investigate the effects of Cordyceps sinensis on renal fibrosis and its possible mechanisms. Sprague-Dawley rats were randomly divided into three groups: sham operation (SHAM) group, 5/6 subtotal nephrectomy (SNx) untreated group, and 5/6 subtotal nephrectomy treated with C. sinensis (2.0 g/kg d) (CS) group. Rats were studied 12 weeks after the surgery, and the CS group presented with significantly lower proteinuria, and better renal function compared with the SNx group (p<0.05). Pathological study showed that the glomerulosclerosis tubulointerstitial injury score was significantly reduced in the CS group compared with the SNx group. Furthermore, the mRNA expression of TGF-ß1, Smad2 and Smad3 and the protein expression of TGF-ß1, TßRI, TßRII and p-Smad2/3 were attenuated by the C. sinensis treatment. In constrast, the mRNA and protein expression of Smad7 was upregulated. Furthermore, the expression of α-SMA and FSP1 was also significantly attenuated, accompanied by the increasing expression of E-cadherin, suggesting the inhibition of the epithelial-mesenchymal transition (EMT). IN CONCLUSION: C. sinensis exerted its antifibrotic effect on the SNx rats through the inhibition of the TGF-ß1/Smad pathway.

Tumour necrosis factor-stimulated gene (TSG)-6 controls epithelial-mesenchymal transition of proximal tubular epithelial cells.

Progressive renal disease is characterized by accumulation of extracellular matrix in the renal cortex. Proximal tubular cells (PTC) may contribute to disease through a process of epithelial-mesenchymal-transition (EMT): phenotypic change, disruption of the tubular basement membrane and migration into the interstitium. Hyaluronan (HA) synthesis and its extracellular organization by hyaladherins affect cell fate in other systems: this study investigated the role of the hyaladherin, tumour necrosis factor-stimulated gene (TSG)-6, in PTC EMT triggered in vitro by transforming growth factor (TGF)ß1. TGFß1 triggered the loss of PTC epithelial phenotype with 60% decreased expression of E-cadherin and 2-3-fold induction of alpha-smooth muscle actin (α-sma). It also increased the expression of TSG-6, HA-synthase-(HAS)2 and the HA-receptor, CD44, to a peak at 8-12h, remaining elevated thereafter. Immuno-localization of HA demonstrated that unstimulated PTC assembled HA in cables and that treatment with TGFß1 initiated cable disassembly with formation of dense HA-pericellular coats. Stable knockdown of TSG-6 with short-hairpin-RNA increased E-cadherin and HAS2 expression, produced loose HA-pericellular coats, HA cables were absent and cell migration was slowed. Treatment of transfectants with TGFß1 did not induce α-sma, alter E-cadherin, pericellular-HA or migration but did induce HAS2. This was dependent on the expression of CD44 and was inhibited by CD44-specific siRNA. In summary, TSG-6 was central to EMT through effects on HA macromolecular structure and through CD44-dependent triggering of cell responses. These findings suggest that controlling the assembly of HA by proximal tubular cells may be a novel approach towards intervention in renal disease.

Characterisation of the human ADAM15 promoter.

BACKGROUND: ADAM15 is a membrane-bound member of the adamalysin family that is up-regulated in areas of tissue remodelling. Previous studies have demonstrated the role of ADAM15 in mesangial cell migration, which is integral in tissue remodelling in pathology and repair. The current study was designed to identify and analyse the genomic regions upstream of ADAM15 that would regulate its transcription. METHODS: Using 5'-RACE and RT-PCR, the ADAM15 5'-UTR was extended and luciferase constructs assembled to examine the transcription start site and characterise the promoter region of this gene. RESULTS: A 145-bp proximal promoter construct showed full activity in unstimulated cells. Analysis of this region identified three potential Sp1-binding sites. Electromobility and supershift assays confirmed that Sp1 was constitutively present in MC nuclei. Mutations in each Sp1 site confirmed each was needed for full activity, while mutation of all three sites abrogated luciferase activity demonstrating that Sp1 was involved in the promoter activity of ADAM15. Methylation of this promoter fragment abolished the activity, while the methyltransferase inhibitor 5-aza-3'-deoxycytidine showed no increased activity in transfected cells, implying that the promoter was not methylated in our cells. CONCLUSION: These results demonstrate the intrinsic promoter activity of ADAM15 in quiescent MC and show the involvement of Sp1 in its regulation.

Regulation of proximal tubular epithelial cell CD44-mediated binding and internalisation of hyaluronan.

BACKGROUND: Increased expression of the connective tissue polysaccharide hyaluronan (HA) in the renal corticointerstitium is associated with progressive renal fibrosis. Numerous studies have demonstrated involvement proximal tubular epithelial cells in the fibrotic process and in the current study we have characterised their expression of the HA receptor, CD44, and examined changes in CD44 expression and function in response to either IL-1beta or glucose. METHODS: Characterisation of CD44 splice variant expression was carried out in primary cultures of human proximal tubular cells (PTC) and HK2 cells. Binding and internalisation HA was examined by addition of exogenous of fluorescein-HA (fl-HA), and expression of CD44 examined by immunoblot analysis and flow cytometry. Alteration in "functional" CD44 was determined by immunoprecipitation of CD44 following stimulation in the presence of fl-HA. RESULTS: PTC, both primary culture and the PTC cell line, HK2, express at least 5 CD44 splice variants, the expression of which are not altered by addition of either IL-1beta or 25mM D-glucose. Addition of either stimulus increased cell surface binding and internalisation of fl-HA and increased expression of functionally active CD44. Increased binding and internalisation of fl-HA, was blocked by anti-CD44 antibody, and by the inhibition of O-glycosylation. CONCLUSIONS: The data demonstrate that stimuli inducing PTC HA synthesis also regulate PTC-HA interactions. Furthermore increased HA binding and internalisation is the result of post-translational modification of CD44 by O-glycosylation, rather than by alteration in expression of CD44 at the cell surface, or by alternate use of CD44 splice variants.

TGF-beta1-mediated fibroblast-myofibroblast terminal differentiation-the role of Smad proteins.

It is now clear that resident myofibroblasts play a central role in the mediation of tissue fibrosis. The aim of the work outlined in this study is to increase our understanding of the mechanisms which drive the phenotypic and functional changes associated with the differentiation process. We have used an in vitro model of transforming growth factor-beta1 (TGF-beta1)-induced pulmonary fibroblast-myofibroblast differentiation to examine the role of the TGF-beta1 Smad protein signaling intermediates, in alterations of fibroblast phenotype and function associated with terminal differentiation. TGF-beta1 induced marked alteration in cell phenotype, such that cells resembled "epithelioid-postmitotic fibroblasts." This was associated with marked reorganization of the actin cytoskeleton and upregulation of alphaSMA gene expression. TGF-beta1 stimulation also induced alphaSMA protein expression with increased incorporation of alphaSMA into stress fibers. Following stimulation with TGF-beta1, subsequent addition of serum-free medium did not reverse TGF-beta1-induced morphological change, suggesting that TGF-beta1 induced a relatively stable alteration in fibroblast cell phenotype. Functionally, these phenotypic changes were associated with induction of type I, type III, and type IV collagen gene expression and an increase in the concentrations of the respective collagens in the cell culture supernatant. The role of Smad proteins in terminal differentiation of fibroblasts was examined by transfection of cells, with expression vectors for the TGFbeta1 receptor-regulated Smads (R-Smads) or the co-Smad, Smad 4. Transfection with Smad2 but not Smad3 resulted in TGF-beta1 independent alteration in fibroblast cell phenotype, up-regulation of alphaSMA mRNA and reorganization of the actin cytoskeleton. Transfection with Smad4 also induced alteration in cell phenotype, although this was not as pronounced as the effect of overexpression of Smad2. Overexpression of the Smad2, Smad3, or Smad4 proteins was associated with increased production of all collagen types. The study suggests that the phenotypic and functional changes associated with TGF-beta1-induced fibroblast terminal differentiation are differentially regulated by Smad proteins.

Interaction between the transforming growth factor-beta type II receptor/Smad pathway and beta-catenin during transforming growth factor-beta1-mediated adherens junction disassembly.

The aim of the current study was to examine the influence of transforming growth factor (TGF)-beta 1 on proximal tubular epithelial cell-cell interaction, with particular emphasis on the regulation of adherens junction complex formation. Stimulation of the proximal tubular cell line HK-2 cells by TGF-beta 1 led to loss of cell-cell contact and disassembly of both adherens and tight junctional complexes. Adherens junction disassembly was associated with reduction of both Triton-soluble and Triton-insoluble E-cadherin, and an increase in detergent-soluble beta-catenin. Under these conditions, immunoprecipitation and Western analysis demonstrated decreased association of beta-catenin, both with E-cadherin, alpha-catenin, and the cell cytoskeleton. Confocal microscopy after immunostaining, showed decreased intensity of peripheral E-cadherin staining, and redistribution of beta-catenin expression to a perinuclear location. Tight junction disassembly was manifest by a reduction in the expression of Triton-soluble occludin and ZO-1 by Western analysis and their disassociation manifested by immunostaining and confocal microscopy. Loss of cell-cell contact and disassembly of adherens junctions were seen after addition of TGF-beta 1 to the basolateral aspect of the cells. Immunoprecipitation experiments demonstrated co-localization of E-cadherin, beta-catenin, and TGF-beta 1 RII in unstimulated cells. After TGF-beta 1 stimulation, the TGF-beta 1 RII no longer associated with either E-cadherin or beta-catenin. Dissociation of the adherens junction protein from the TGF-beta 1 receptor was associated with increased beta-catenin tyrosine phosphorylation and decreased threonine phosphorylation. Furthermore after receptor ligand binding, beta-catenin became associated with the TGF-beta 1-signaling molecules Smad3 and Smad4.

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.