Methods for estimating the volume of individual glomeruli

Methods for estimating the volume of individual glomeruli.BackgroundThe Cavalieri and maximal planar area (MPA) methods are commonly used to measure the volume of individual glomeruli. Previous studies have suggested that the MPA method, which is less laborious, yields values that are much greater than those obtained by the Cavalieri method. The current study re-examined the relationship of MPA and Cavalieri values for glomerular volume in humans and rats.MethodsBoth methods were used to measure the volume of 1201 glomeruli from 58 humans and 281 glomeruli from 15 rats. Tissue was embedded in Epon. Further mathematical analysis was performed to assess the extent to which deviation of glomeruli from spherical shape affects the relationship of values obtained by the MPA and Cavalieri methods.ResultsMPA values exceeded Cavalieri values by an average of only 14 ± 22% in humans and 6 ± 16% in rats. The relationship of MPA to Cavalieri values was similar in individual humans and rats, with widely varying values for average glomerular volume. Neither the development of sclerosis nor the loss of any connection to a tubule affected the relationship of the MPA and Cavalieri values for the volume of individual glomeruli. Mathematical analysis showed that MPA values would not exceed Cavalieri values if glomeruli had ellipsoidal rather than spherical shape.ConclusionSimilar values for glomerular volume are obtained using the Cavalieri and MPA methods in humans and rats

Application of hydroxypropylated crosslinked starch from the grains of Oryza sativa L. as potential disintegrant in compressed solid dosage form.

Context: The modification of starch from the grains of NSICRc222 rice variety may provide a new material with enhanced disintegrating functionality in compressed solid dosage forms. Aims: To synthesize hydroxypropylated crosslinked rice starch that may possess characteristics of a good disintegrant. Methods: Starch was isolated by the wet milling method. The crosslinking was performed using trisodium trimetaphosphate as crosslinking agent, and successively hydroxypropylated by using phosphorous oxychloride. The evidences of chemical modifications were determined by FTIR, TGA, and DSC. The application of modified rice starch as a disintegrant was evaluated by determining the dissolution profile, and was compared to the performance of commercially available disintegrants. The compatibility of modified rice starch was employed using FTIR, and the stability of the formulation was subjected to an accelerated stability study. Results: The changes in infrared spectra and thermogram confirmed successful chemical modification of rice starch. Allopurinol and modified starch mixture staged from 0 to 3 months showed no significant changes in the infrared spectra, which suggest compatibility of HCR with allopurinol. The dissolution rate of allopurinol tablets at pH 1.2, pH 4.5, and pH 6.8 with various superdisintegrants were comparable to the dissolution rate that utilized hydroxypropylated crosslinked rice starch as disintegrant. The accelerated stability study showed no significant changes from 0th to 3rd month, and no degradation products were detected in the HPLC analysis. Conclusions: The chemical modification of rice starch through crosslinking and hydroxypropylation yielded a novel material comparable to the commercially available superdisintegrants

Podocyte-specific expression of cre recombinase in transgenic mice

Summary: We report a transgenic mouse line that expresses Cre recombinase exclusively in podocytes. Twenty- four transgenic founders were generated in which Cre recombinase was placed under the regulation of a 2.5-kb fragment of the human NPHS2 promoter. Previously, this fragment was shown to drive beta-galactosidase (Β-gal) expression exclusively in podocytes of transgenic mice. For analysis, founder mice were bred with ROSA26 mice, a reporter line that expresses Β-gal in cells that undergo Cre recombination. Eight of 24 founder lines were found to express Β-gal exclusively in the kidney. Histological analysis of the kidneys showed that Β-gal expression was confined to podocytes. Cre recombination occurred during the capillary loop stage in glomerular development. No evidence for Cre recombination was detected in any of 14 other tissues examined. genesis 35:39–42, 2003. © 2002 Wiley-Liss, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/35267/1/10164_ftp.pd

Blockade of Oxidative Stress by Vitamin C Ameliorates Albuminuria and Renal Sclerosis in Experimental Diabetic Rats

PURPOSE: Oxidative stress has been suggested to play a role as a common mediator of apoptosis and kidney damage in diabetes. However, it is uncertain whether the apoptosis occurs in the kidney during the course of diabetes. We investigated the occurrence of apoptosis in the diabetic rat kidney, the role of oxidative stress and the effect of an antioxidant on apoptosis in the diabetic rat kidney. MATERIALS AND METHODS: Otsuka-Long-Evans-Tokushima-Fatty rats, an animal model for type 2 diabetes, were randomized into a non-treated diabetic (n=8) and a vitamin C-treated group (n=8). Long-Evans Tokushima Otsuka rats (n=8) were used as a control. RESULTS: Apoptosis was present in the epithelial cells of the proximal tubules in diabetic rats. The number of apoptotic cells, albuminuria, proteinuria, glomerular and tubulointerstitial sclerosis, and renal malondialdehyde were significantly decreased in vitamin C-treated diabetic rats when compared to the untreated diabetic rats. The decreased slit pore density (number of slit pores per underlying glomerular basement membrane length) as assessed by electron microscopy was also significantly restored by treatment with vitamin C without significantly affecting plasma glucose in diabetic rats. CONCLUSION: By blocking these pathophysiologic processes, a blockade of oxidative stress by vitamin C might become a useful adjunct to albuminuria and renal sclerosis in diabetic nephropathy.ope

Reduction in podocyte density as a pathologic feature in early diabetic nephropathy in rodents: Prevention by lipoic acid treatment

BACKGROUND: A reduction in the number of podocytes and podocyte density has been documented in the kidneys of patients with diabetes mellitus. Additional studies have shown that podocyte injury and loss occurs in both diabetic animals and humans. However, most studies in animals have examined relatively long-term changes in podocyte number and density and have not examined effects early after initiation of diabetes. We hypothesized that streptozotocin diabetes in rats and mice would result in an early reduction in podocyte density and that this reduction would be prevented by antioxidants. METHODS: The number of podocytes per glomerular section and the podocyte density in glomeruli from rats and mice with streptozotocin (STZ)-diabetes mellitus was determined at several time points based on detection of the glomerular podocyte specific antigens, WT-1 and GLEPP1. The effect of insulin administration or treatment with the antioxidant, α-lipoic acid, on podocyte number was assessed. RESULTS: Experimental diabetes resulted in a rapid decline in apparent podocyte number and podocyte density. A significant reduction in podocytes/glomerular cross-section was found in STZ diabetes in rats at 2 weeks (14%), 6 weeks (18%) and 8 weeks (34%) following STZ injection. Similar declines in apparent podocyte number were found in STZ diabetes in C57BL/6 mice at 2 weeks, but not at 3 days after injection. Treatment with α-lipoic acid substantially prevented podocyte loss in diabetic rats but treatment with insulin had only a modest effect. CONCLUSION: STZ diabetes results in reduction in apparent podocyte number and in podocyte density within 2 weeks after onset of hyperglycemia. Prevention of these effects with antioxidant therapy suggests that this early reduction in podocyte density is due in part to increased levels of reactive oxygen species as well as hyperglycemia

The Protective Role of Smad7 in Diabetic Kidney Disease: Mechanism and Therapeutic Potential

OBJECTIVE - Although Smad3 has been considered as a downstream mediator of transforming growth factor-β (TGF-β) signaling in diabetes complications, the role of Smad7 in diabetes remains largely unclear. The current study tests the hypothesis that Smad7 may play a protective role and has therapeutic potential for diabetic kidney disease. RESEARCH DESIGN AND METHODS - Protective role of Smad7 in diabetic kidney disease was examined in streptozotocin-induced diabetic mice that have Smad7 gene knockout (KO) and in diabetic rats given Smad7 gene transfer using an ultrasound-microbubble-mediated technique. RESULTS - We found that mice deficient for Smad7 developed more severe diabetic kidney injury than wild-type mice as evidenced by a significant increase in microalbuminuria, renal fibrosis (collagen I, IV, and fibronectin), and renal inflammation (interleukin-1β [IL-1β], tumor necrosis factor-α [TNF-α], monocyte chemoattractant protein-1 [MCP-1], intracellular adhesion molecule-1 [ICAM-1], and macrophages). Further studies revealed that enhanced renal fibrosis and inflammation in Smad7 KO mice with diabetes were associated with increased activation of both TGF-β/Smad2/3 and nuclear factor-κB (NF-κB) signaling pathways. To develop a therapeutic potential for diabetic kidney disease, Smad7 gene was transferred into the kidney in diabetic rats by an ultrasound-microbubble-mediated technique. Although overexpression of renal Smad7 had no effect on levels of blood glucose, it significantly attenuated the development of microalbuminuria, TGF-β/Smad3-mediated renal fibrosis such as collagen I and IV and fibronectin accumulation and NF-κB/p65-driven renal inflammation including IL-1β, TNF-α, MCP-1, and ICAM-1 expression and macrophage infiltration in diabetic rats. CONCLUSIONS-Smad7 plays a protective role in diabetic renal injury. Overexpression of Smad7 may represent a novel therapy for the diabetic kidney complication. © 2011 by the American Diabetes Association.link_to_subscribed_fulltex

Podocyte specific knock out of selenoproteins does not enhance nephropathy in streptozotocin diabetic C57BL/6 mice

<p>Abstract</p> <p>Background</p> <p>Selenoproteins contain selenocysteine (Sec), commonly considered the 21^stgenetically encoded amino acid. Many selenoproteins, such as the glutathione peroxidases and thioredoxin reductases, protect cells against oxidative stress by functioning as antioxidants and/or through their roles in the maintenance of intracellular redox balance. Since oxidative stress has been implicated in the pathogenesis of diabetic nephropathy, we hypothesized that selenoproteins protect against this complication of diabetes.</p> <p>Methods</p> <p>C57BL/6 mice that have a podocyte-specific inability to incorporate Sec into proteins (denoted in this paper as PodoTrsp^-/-) and control mice were made diabetic by intraperitoneal injection of streptozotocin, or were injected with vehicle. Blood glucose, body weight, microalbuminuria, glomerular mesangial matrix expansion, and immunohistochemical markers of oxidative stress were assessed.</p> <p>Results</p> <p>After 3 and 6 months of diabetes, control and PodoTrsp^-/-mice had similar levels of blood glucose. There were no differences in urinary albumin/creatinine ratios. Periodic acid-Schiff staining to examine mesangial matrix expansion also demonstrated no difference between control and PodoTrsp^-/-mice after 6 months of diabetes, and there were no differences in immunohistochemical stainings for nitrotyrosine or NAD(P)H dehydrogenase, quinone 1.</p> <p>Conclusion</p> <p>Loss of podocyte selenoproteins in streptozotocin diabetic C57BL/6 mice does not lead to increased oxidative stress as assessed by nitrotyrosine and NAD(P)H dehydrogenase, quinone 1 immunostaining, nor does it lead to worsening nephropathy.</p