Biomedical subjects

Sumant S Chugh

Publications and source records attributed to Sumant S Chugh.

13 recordsLinked to original sources

PubMed2006

ZHX proteins regulate podocyte gene expression during the development of nephrotic syndrome.

Transcriptional regulation of podocyte gene expression in primary glomerular disease is poorly understood. In this study, we demonstrate a prominent role of members of the ZHX (zinc fingers and homeoboxes) family of proteins in regulating podocyte gene expression during the development of nephrotic syndrome. While studying mechanisms of glomerular disease, rat ZHX3 was cloned from a down-regulated gene fragment; its cellular localization, DNA binding, and transcriptional repressor properties were characterized; and its ability to influence podocyte gene expression directly or via ZHX1 and ZHX2 was studied. In eukaryotic promoters, ZHX3 bound to the CdxA binding motif. ZHX proteins were mostly sequestered in the non-nuclear compartment in the normal in vivo podocyte by virtue of heterodimer formation, and loss of heterodimerization was associated with entry into the nucleus. In experimental minimal change disease, ZHX3 was transiently down-regulated prior to the onset of proteinuria, and recovery of expression was associated with migration of ZHX3 protein into the nucleus and the development of proteinuria. This expression pattern mirrored the increased nuclear ZHX3 expression noted in vivo in the podocytes in human minimal change disease biopsies. In vitro, migration of ZHX3 protein into the nucleus during recovery from transient ZHX3 knockdown reproduced the gene expression profile of in vivo minimal change disease. Severe sustained knockdown of ZHX3 caused down-regulation of genes involved in focal sclerosis, including WT1, mediated mostly by increased nuclear entry of ZHX2 and ZHX1. In summary, ZHX proteins are major transcriptional mediators of podocyte disease.

Animals↗

PubMed2005

Modulation of renal-specific oxidoreductase/myo-inositol oxygenase by high-glucose ambience.

Biological properties of renal-specific oxidoreductase (RSOR), characteristics of its promoter, and underlying mechanisms regulating its expression in diabetes were analyzed. RSOR expression, normally confined to the renal cortex, was markedly increased and extended into the outer medullary tubules in db/db mice, a model of type 2 diabetes. Exposure of LLCPK cells to d-glucose resulted in a dose-dependent increase in RSOR expression and its enzymatic activity. The latter was related to one of the glycolytic enzymes, myo-inositol oxygenase. The increase in activity was in proportion to serum glucose concentration. The RSOR expression also increased in cells treated with various organic osmolytes, e.g., sorbitol, myoinositol, and glycerolphosphoryl-choline and H(2)O(2). Basal promoter activity was confined to -1,252 bp upstream of ATG, and it increased with the treatment of high glucose and osmolytes. EMSAs indicated an increased binding activity with osmotic-, carbohydrate-, and oxidant-response elements in cells treated with high glucose and was abolished by competitors. Supershifts, detected by anti-nuclear factor of activated T cells, and carbohydrate-response-element-binding protein established the binding specificity. Nuclear factor of activated T cells tonicity-enhancer-binding protein and carbohydrate-response-element-binding protein had increased nuclear expression in cells treated with high glucose. The activity of osmotic-response element exhibited a unique alternate binding pattern, as yet unreported in osmoregulatory genes. Data indicate that RSOR activity is modulated by diverse mechanisms, and it is endowed with dual properties to channel glucose intermediaries, characteristic of hepatic aldehyde reductases, and to maintain osmoregulation, a function of renal medullary genes, e.g., aldose reductase, in diabetes.

Animals↗

PubMed2005

Cell biology of diabetic kidney disease.

In large part cellular dysfunctions induced by chronic hyperglycemia are similar in type-1 and -2 diabetes. In both instances chronic hyperglycemia induces injury to a multitude of organs by affecting various target cells. The cells affected may include those derived from of epithelial or mesenchymal progenitors; and at times hyperglycemia may induce phenotypic changes with epithelial-mesenchymal transformation. In the majority of target cells the high-glucose ambience activates various intracellular pathways that are similar except for minor exceptions that are related to the selective expression of various molecules in a given cell type. Keeping in perspective a common paradigm applicable to most of the cells, a brief discussion of different hyperglycemia-induced cellular events pertaining to various pathways is described in this review. They include fluxes of glucose intermediaries in various cellular metabolic pathways, generation of advanced glycation end products (AGEs) and their extra- and intracellular effects, the role of protein kinase C, transforming growth factor-beta, guanosine triphosphate-binding proteins and reactive oxygen species (ROS) in various cellular signaling events. The latter, i.e., ROS, may be central to several intracellular pathways and modulate various events in a reciprocal manner. The information compiled under various subtitles of this synopsis is derived from an enormous amount of literature data summarized in several recent excellent reviews, and thus further reading of them is suggested to gather detailed comprehensive information on each of the subjects.

Animals↗

PubMed2005

Hyperglycemia: its imminent effects on mammalian nephrogenesis.

A sustained exposure of the mammalian embryo to very high glucose ambience is associated with a multitude of congenital birth defects, including those of the cardiovascular, CNS, skeletal and urogenital systems during the first 6-8 weeks of gestation in humans. These urogenital abnormalities may be associated with "caudal regression syndrome" or may occur alone in the form of partial or total renal agenesis. Similarly, an increase in the incidence of morphogenetic defects is observed in the offspring of streptozotocin-induced diabetic rats and mice, and also in non-obese diabetic mice. In certain cases, failure during the growth of the lower parts of embryos or newborn mice involving the genitourinary system has been observed in animals with severe diabetes. Investigators have utilized whole organ culture systems to delineate the mechanisms relevant to dysmorphogenesis of the embryonic metanephros. A marked dysmorphogenesis of the metanephros is observed upon treatment with a high concentration of D: -glucose. Associated with it are changes that include branching dysmorphogenesis of the ureteric bud iterations, reduced population of nascent nephrons, decreased expression of basement membrane proteoglycans, depletion of ATP stores, and fulminant apoptosis of the cells at the interface of mesenchyme and ureteric bud epithelium. The latter findings suggest that disruption of epithelial:mesenchymal interactions may be the major event responsible for the metanephric dysmorphogenesis induced by high glucose ambience.

Animals↗

PubMed2005

Renal-specific oxidoreductase biphasic expression under high glucose ambience during fetal versus neonatal development.

BACKGROUND: Renal-specific oxidoreductase (RSOR) has been recently identified in mice kidneys of diabetic animals, and it is developmentally regulated. Its expression during fetal, neonatal, and postnatal periods was assessed under high glucose ambience. METHODS: Whole-mount immunofluorescence and confocal microscopy were performed to assess the effect of high glucose on the morphogenesis of mice fetal kidneys. RSOR mRNA and protein expression was assessed by competitive polymerase chain reaction (PCR) and immunoprecipitation methods in embryonic kidneys (day E13 to E17) subjected to high glucose ambience and by Northern and Western blot analyses of kidneys of newborn and 1-week-old mice with hyperglycemia. The spatiotemporal changes in the RSOR expression were assessed by in situ hybridization analyses and immunofluorescence microscopy. In addition, the extent of apoptosis in the kidneys was determined by terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) nick-end labeling (TUNEL) assay. RESULTS: Whole-mount microscopy of the embryonic metanephroi revealed a dose-dependent disruption in the ureteric bud iterations with reduced population of the nascent nephrons. Both gene and protein expressions were reduced in day E13 to E17 metanephroi, while increased in kidneys of newborn and 1-week-old mice. In day E13 and day E15 kidneys, the RSOR was expressed in the ureteric bud branches and some of the immature tubules, and its expression was reduced with high glucose treatment. In day E17 kidneys the RSOR was expressed in the tubules of the deeper cortex, and its expression was marginally decreased. In newborn kidneys, this enzyme was expressed in the subcortical tubules and it spread to the entire width of the renal cortex in hyperglycemic state. In 1-week-old mice kidneys, the RSOR was localized to the entire cortex, and in animals with blood glucose above 300 mg/dL, its intensity increased with extension of expression into the outer medullary tubules. A dose-dependent fulminant apoptosis was observed in day E13 to E17 kidneys subjected to high glucose ambience. In newborn and 1-week-old mice control kidneys, the apoptosis was minimal although slightly increased during hyperglycemia. CONCLUSION: High glucose has a differential effect on the RSOR expression in kidneys during the embryonic versus neonatal/postnatal period. This may partly be related to the differential degree of apoptosis, a process reflective of oxidant stress that is seen in diabetic milieu, which as previously has been shown to adversely effect the modulators of fetal development and thereby the morphogenesis of the kidney and RSOR expression.

Animals↗

PubMed2004

Update of extracellular matrix, its receptors, and cell adhesion molecules in mammalian nephrogenesis.

One of the hallmarks of mammalian nephrogenesis includes a mesenchymal-epithelial transition that is accomplished by intercalation of the ureteric bud, an epithelium-lined tubelike structure, into an undifferentiated mesenchyme, and the latter then undergoes an inductive transformation and differentiates into an epithelial phenotype. At the same time, the differentiating mesenchyme reciprocates by inducing branching morphogenesis of the ureteric bud, which forms a treelike structure with dichotomous iterations. These reciprocal inductive interactions lead to the development of a functioning nephron unit made up of a glomerulus and proximal and distal tubules. The inductive interactions and differentiation events are modulated by a number of transcription factors, protooncogenes, and growth factors and their receptors, which regulate the expression of target morphogenetic modulators including the ECM, integrin receptors, and cell adhesion molecules. These target macromolecules exhibit spatiotemporal and stage-specific developmental regulation in the metanephros. The ECM molecules expressed at the epithelial-mesenchymal interface are perhaps the most relevant and conducive to the paracrine-juxtacrine interactions in a scenario where the ligand is expressed in the mesenchyme while the receptor is located in the ureteric bud epithelium or vice versa. In addition, expression of the target ECM macromolecules is regulated by matrix metalloproteinases and their inhibitors to generate a concentration gradient at the interface to further propel epithelial-mesenchymal interactions so that nephrogenesis can proceed seamlessly. In this review, we discuss and update our current understanding of the role of the ECM and related macromolecules with respect to metanephric development.

Animals↗

PubMed2003

Renal development in high-glucose ambience and diabetic embryopathy.

Maternal diabetes has an adverse influence on the intrauterine growth of the fetus, which is attributable to the exposure of the mammalian embryo to an abnormal metabolic environment. A sustained exposure of the fetus to such an environment (ie, elevated concentration of glucose), during the first 6 to 8 weeks of gestation in humans may result in diabetic embryopathy, which is characterized by a multitude of congenital birth defects, including those of the nervous, cardiovascular, skeletal, and urogenital systems. The urogenital abnormalities may be associated with caudal regression syndrome or may occur alone in the form of partial or total renal agenesis. Similarly, an increase in the incidence of morphogenetic defects is observed in offsprings of streptozotocin-induced diabetic rats and mice and also in nonobese diabetic mice. In certain instances, failure in the growth of lower part of embryos or newborn mice has been observed in animals with a severe diabetic state. For further delineation of the mechanisms involved in the pathogenesis of diabetic embryopathy, the investigators used whole-embryo culture systems, and found that glucose can induce defects mainly confined to the lower part of the body involving the genitourinary system. Similarly, dysmorphogenesis of the embryonic metanephros is observed when it is subjected to high concentrations of D-glucose and its epimer D-mannose. This article discusses certain aspects of diabetic embryopathy with an emphasis on changes that occur in the fetal metanephros in high-glucose ambience.

Animals↗

PubMed2003

Molecular structure-function relationship in the slit diaphragm.

The past 5 years have witnessed an exponential increase in our understanding of the structure and function of the glomerular slit diaphragm. The identification of nephrin as the first transmembrane slit diaphragm protein was a watershed event in slit diaphragm biology. This article correlates some of the observations of the prenephrin era with more recent studies, and elaborates on the individual characteristics of each slit diaphragm protein. Recent studies on protein-protein interactions related to slit diaphragm permeability and cell signaling are elaborated, along with observations on their expression in human disease and experimental models of proteinuria. Developmental expression of components of the slit diaphragm in normal and knockout mice also is discussed. Finally, some areas of future investigation are proposed.

Animals↗

PubMed2003

Neph1 and nephrin interaction in the slit diaphragm is an important determinant of glomerular permeability.

Neph1-deficient mice develop nephrotic syndrome at birth, indicating the importance of this protein in the development of a normal glomerular filtration barrier. While the precise subcellular localization of Neph1 remains unknown, its relationship with other components of the glomerular filtration barrier is of great interest in this field. In this paper, we localize the expression of Neph1 to the glomerular slit diaphragm by immunogold electron microscopy in rodents and describe its direct interaction with two other components of the slit diaphragm, nephrin and ZO-1. Both native and recombinant Neph1 associate with each other as dimers and multimers and interact with nephrin via their extracellular segments. Disruption of the Neph1-nephrin interaction in vivo by injecting combinations of individual subnephritogenic doses of anti-Neph1 and anti-nephrin results in complement- and leukocyte-independent proteinuria with preserved foot processes. This disruption modestly reduces Neph1 and nephrin protein expression in podocytes and dramatically reduces ZO-1 protein expression via the interaction of ZO-1 PDZ domains with the cytoplasmic tail of Neph1, independent of changes in mRNA expression of all three genes. The interaction between nephrin and Neph1 is specific and not shared by either protein with P-cadherin, another integral slit diaphragm protein. The interaction between nephrin and Neph1 therefore appears to be an important determinant of glomerular permeability.

Amino Acid Sequence↗

PubMed2002

High glucose stimulates synthesis of fibronectin via a novel protein kinase C, Rap1b, and B-Raf signaling pathway.

The molecular mechanism(s) by which high glucose induces fibronectin expression via G-protein activation in the kidney are largely unknown. This investigation describes the effect of high glucose (HG) on a small GTP-binding protein, Rap1b, expression and activation, and the relevance of protein kinase C (PKC) and Raf pathways in fibronectin synthesis in cultured renal glomerular mesangial cells (MCs). In vivo experiments revealed a dose-dependent increase in Rap1b expression in glomeruli of diabetic rat kidneys. Similarly, in vitro exposure of MCs to HG led to an up-regulation of Rap1b with concomitant increase in fibronectin (FN) mRNA and protein expression. The up-regulation of Rap1b mRNA was mitigated by the PKC inhibitors, calphostin C, and bisindolymaleimide, while also reducing HG- induced FN expression in non-transfected MCs. Overexpression of Rap1b by transfection with pcDNA 3.1/Rap1b in MCs resulted in the stimulation of FN synthesis; however, the PKC inhibitors had no significant effect in reducing FN expression in Rap1b-transfected MCs. Transfection of Rap1b mutants S17N (Ser --> Asn) or T61R (Thr --> Arg) in MCs inhibited the HG-induced increased FN synthesis. B-Raf and Raf-1 expression was investigated to assess whether Rap1b effects are mediated via the Raf pathway. B-Raf, and not Raf-1, expression was increased in MCs transfected with Rap1b. HG also caused activation of Rap1b, which was largely unaffected by anti-platelet-derived growth factor (PDGF) antibodies. HG-induced activation of Rap1b was specific, since Rap2b activation and expression of Rap2a and Rap2b were unaffected by HG. These findings indicate that hyperglycemia and HG cause an activation and up-regulation of Rap1b in renal glomeruli and in cultured MCs, which then stimulates FN synthesis. This effect appears to be PKC-dependent and PDGF-independent, but involves B-Raf, suggesting a novel PKC-Rap1b-B-Raf pathway responsible for HG-induced increased mesangial matrix synthesis, a hallmark of diabetic nephropathy.

Animals↗

PubMed2002

Isolation and functional analysis of mouse UbA52 gene and its relevance to diabetic nephropathy.

In delineating the mechanism(s) of diabetic nephropathy various novel genes have been isolated, whereas others remain to be discovered. We identified several up-regulated genes in the kidneys of diabetic newborn mice. Among them was UbA52, a ubiquitin ribosomal fusion protein. Its mRNA expression in the kidney was proportional to blood glucose levels. By in situ hybridization and immunohistochemistry, UbA52 was exclusively localized to renal tubules, and its expression was markedly increased in diabetic mice. The up-regulated UbA52 mRNA and protein expression were also observed in Madin-Darby canine kidney cells, a tubular cell line, treated with 30 mm glucose in both cell lysates and ribosomal fractions. To explore the mechanism(s) of its increased expression, UbA52 genomic DNA was isolated. A transcription start site at -22 bp from the initiation codon was identified and confirmed by primer extension analysis. The UbA52 promoter region included glucose response-related E-box sequences and stress response elements (STRE). Unlike in humans, mouse UbA52 gene had no introns in the coding or 5'-ATG-flanking regions. To identify the DNA segment with maximal promoter activity, deletion constructs were prepared using a pSEAP vector system and transfected into COS7 kidney cells. Maximal activity was confined to -198 to +68 bp, which included E-boxes and STRE motifs. A dose-dependent increase in the promoter activity was observed in cells exposed to high glucose. Mutations in the first E-box (CAGCTG --> TGGCTG) or STRE (CCCCT --> CATCT) resulted in a decrease in the SEAP activity under high glucose ambience. Given the presence of glucose-responsive motifs in the promoter region and decrease in the SEAP activity in E-box mutants in the presence of glucose, these data suggest that UbA52, a ribosomal fusion protein, may be relevant in the pathogenesis of diabetic nephropathy.

Animals↗

PubMed2002

Renal gene expression in embryonic and newborn diabetic mice.

Several novel genes that are upregulated in diabetic kidneys have been identified. Recently, transforming growth factor beta driven secreted proteins, i.e., connective tissue growth factor and gremlin (bone morphogenetic protein 2), have been identified, and their expression has been correlated with the tissue changes seen in diabetic nephropathy in the adult population. However, there are very few studies reported in the literature that describe the gene expression in the diabetic state during embryonic and neonatal life. It is well known that exposure to glucose or its epimer, i.e., mannose, induces marked dysmorphogenesis of the embryonic metanephros in an organ culture system. These changes are associated with ATP depletion and marked apoptosis, suggesting an oxidant stress in the induction of dysmorphogenesis of the embryonic metanephros. In view of the glucose-induced changes in the fetal metanephros, a diabetic state was induced by the administration of streptozotocin during pregnancy, and newborn mouse kidneys were processed for suppression subtractive hybridization-PCR. In addition, a diabetic state was induced in newborn diabetic mice, and after 1 week their kidneys were harvested and subjected to representational difference analysis of cDNA. Four novel genes with upregulated mRNA expression were identified. They included: (1) a translocase inner mitochondrial membrane 44 that is involved in the ATP-dependent import of preproteins from the cytosol into the mitochondrial matrix; (2) a kidney-specific aldo-keto reductase that utilizes NADPH and NADH as cofactors in the reduction of aromatic aldehydes and aldohexoses; (3) Rap1b, a Ras-related small GTP-binding protein that behaves as a GTPase and cycles between GTP-bound (active) and GDP-bound (inactive) states associated with conformational change, and (4) a fusion protein of ubiquitin polypeptide and ribosomal protein L40 (UbA(52) or ubiquitin/60) that is intimately involved in the ubiquitin-dependent proteasome pathway related to the accelerated degradation of proteins under various stress conditions, such as those seen in patients with cancer and diabetes mellitus.

Alcohol Oxidoreductases↗

PubMedDate not supplied

Co-existent crescentic glomerulonephritis and renal amyloidosis: a case report and literature review.

Co-existent crescentic glomerulonephritis and renal amyloidosis have only rarely been reported thus far. To our knowledge, only about eighteen convincing cases (with >50% crescents) have been described, mostly in the context of rheumatoid arthritis. In this report, we add a case from our institution and review the existing literature, including a proposed mechanism of crescent formation in this setting. Although the relationship between crescentic glomerulonephritis, renal amyloidosis, and rheumatoid arthritis needs further investigation, we suggest that one should consider the development of crescentic glomerulonephritis with amyloidosis in patients with long-standing rheumatoid arthritis and worsening renal function.

Amyloidosis↗