Degree Of Renal Inflammation Research Articles

MO076: GLUT2 as a Key Factor in the Pathogenesis of Diabetic Kidney Disease

Abstract BACKGROUND AND AIMS Diabetic kidney disease (DKD) affects more than 30% of all diabetic patients. It is strongly associated with increased morbidity, mortality and global healthcare burden. Furthermore, it is considered the primary cause of end-stage kidney disease (ESKD). The renal proximal tubular cells (RPTCs) are highly susceptible to diabetes-induced hyperglycaemia, and glucose handling in these cells plays a central role in the pathophysiology of DKD; yet, the molecular mechanisms underlying hyperglycaemia-induced tubulopathy are poorly understood. Although glucose transporter 2 (GLUT2) is a major glucose transporter in the RPTCs, its exclusive role and the underlying mechanism by which it contributes to the pathogenesis of DKD are poorly known. Here, we reveal its fundamental role in the pathogenesis of DKD. METHOD By using the Cre-lox technique, we developed a novel mouse strain with reduced GLUT2 expression specifically in the RPTCs and crossed it with the Akita-diabetic mouse strain to generate diabetic RPTC-GLUT2-/- mice. We evaluated the renal phenotype of the null mice and compared it with their diabetic wild-type (WT) littermate controls by assessing renal patho-morphological changes, measuring blood and urine biochemical alterations, and testing renal function by qPCR, immunohistochemistry and ELISAs. Moreover, we measured glucose uptake by the kidney in the diabetic RPTC-GLUT2-/- mice and their WT controls by utilizing µPET-MRI imaging of tail vein-injected 18[F]-deoxy-D-glucose (FDG). RESULTS Whereas no significant changes were found in the weight or the diabetic status of the null mice in comparison with the WT animals, considerable improvements of renal function in the diabetic RPTC-GLUT2-/- mice were measured. These positive changes included significant reductions in urine creatinine, albuminuria, albumin-to-creatinine ratio (ACR) and urinary kidney injury marker 1 (KIM-1) levels. Interestingly, the hyperglycemia-induced patho-morphological changes in the kidney as well as renal injury, fibrosis and inflammation were significantly attenuated in the diabetic RPTC-GLUT2-/- mice. Moreover, the renal uptake of 18[F]-FDG was significantly decreased in the diabetic RPTC-GLUT2-/- mice compared with their WT littermate controls. CONCLUSION Fundamental novel observations regarding the contribution of GLUT2 in RPTCs to the development and pathogenesis of DKD have emerged. Our findings suggest that RPTC-GLUT2 not only affects glucose reabsorption but also modulates cellular function, eventually affecting the degree of renal inflammation, tubulointerstitial fibrosis, diabetic kidney injury and renal patho-morphology. Correspondingly, our results indicate that GLUT2-linked molecular mechanisms greatly affect DKD pathophysiology. Therefore, studying these cellular pathways have a major potential for therapeutic intervention of DKD and other kidney diseases.

Mannose-binding lectin activates the nuclear factor-κB and renal inflammation in the progression of diabetic nephropathy.

Increased serum mannose-binding lectin (MBL) level has been proven to correlate with the development of diabetic nephropathy (DN). Here, we aim to find the role and mechanism of MBL involved in the progression of DN. Patients with DN were recruited and divided into two groups according to different rs1800450genotypes of the MBL2gene, and inflammatory profiles in monocytes/macrophages were compared between the two groups. MBL was given to treat macrophages, HK2, and HMC, and a co-culture transwell system was then employed. Renal inflammation and fibrosis parameters were measured after knocking down or overexpressing MBL genes in mice. Proinflammatory profile, manifesting as enhanced IL-1β production and M1 polarization, was found in monocytes/macrophages from DN with a rs1800450 GG genotype of MBL2gene who had higher MBL level, compared with those with a rs1800450 GA genotype. In mechanism, MBL directly induced inflammatory responses in macrophages, which promoted inflammatory and fibrotic markers in HK2 and HMCs during co-culture. Further experiments showed that MBL can promote macrophages transforming to the M1subset mainly by activating the nuclear factor-κB pathway. After downregulation of MBL, the blood glucose, triglyceride, urine protein, injuries of glomerulus and tubules, and the degree of renal inflammation and fibrosis were ameliorated in db/db mice treated with AAV-MBL1/2-shRNA. Overexpression of MBL promoted macrophage infiltration in the kidney. In conclusion, MBL is a crucial mediator in the progression of DN via activating the nuclear factor-κB pathway in macrophages. This will serve as a genetic base for some patients with DN who have poor outcomes and provide a direction for the screening.

Effect of intra-amniotic lipopolysaccharide on nephron number in preterm fetal sheep

Chorioamnionitis is an antecedent of preterm birth. We aimed to determine the effect of experimental chorioamnionitis in fetal sheep during late gestation on 1) nephron number, 2) renal corpuscle volume, and 3) renal inflammation. We hypothesized that exposure to chorioamnionitis would lead to inflammation in fetal kidneys and adversely impact on the development of nephrons, leading to a reduction in nephron number. At ∼121 days of gestation (term ∼147 days), pregnant ewes bearing twin or singleton fetuses received a single intra-amniotic injection of lipopolysaccharide (n = 6; 3 singletons, 3 twins); controls were either untreated or received an intra-amniotic injection of saline (n = 8; 4 singletons, 4 twins). One twin was used from each twin-bearing ewe. At ∼128 days of gestation, fetuses were delivered via Caesarean section. Kidneys were collected and stereologically analyzed to determine nephron number and renal corpuscle volume. Renal inflammation was assessed using immunohistochemistry. Experimental chorioamnionitis did not affect body weight or relative kidney weight. There was a significant reduction in nephron number but no change in renal corpuscle volume in LPS-exposed fetuses relative to controls. On average, nephron number was significantly reduced by 23 and 18% in singleton and twin LPS-exposed fetuses, respectively. The degree of renal inflammation did not differ between groups. Importantly, this study demonstrates that exposure to experimental chorioamnionitis adversely impacts on nephron number in the developing fetus.

Renal inflammation is modulated by potassium in chronic kidney disease: possible role of Smad7

High-potassium diets have been shown to be beneficial in cardiovascular disease partly because of a blood pressure-lowering effect. The effect of potassium on inflammation has not been studied. We investigated the influence of potassium supplementation on the degree of renal inflammation and the intracellular signaling mechanisms that could mediate inflammation in chronic kidney disease (CKD). CKD was created in male Sprague-Dawley rats by subtotal nephrectomy. Two groups of CKD rats were pair fed with diets containing 2.1% potassium (potassium-supplemented diet) or 0.4% potassium (basal diet). Body weight, blood pressure, and blood and urine electrolytes were measured biweekly. The animals were euthanized at week 8, and the remnant kidneys were analyzed by histology, immunohistochemistry, Western blotting, and real-time quantitative PCR. In the CKD pair-fed groups, blood potassium concentration did not differ significantly, but blood pressure was lower in the potassium-supplemented group. Compared with the basal diet, potassium supplementation decreased renal tubulointerstitial injury and suppressed renal inflammation as evidenced by decreased macrophage infiltration, lower expression of inflammatory cytokines, and decreased NF-kappaB activation. These renoprotective effects were associated with downregulation of renal transforming growth facto-beta, upregulation of renal Smad7, and lower blood pressure. Our results show that potassium supplementation can reduce renal inflammation and hence, could modulate the progression of kidney injury in CKD.

Noninvasive Ultrasound Imaging of Inflammation Using Microbubbles Targeted to Activated Leukocytes

Lipid microbubbles used for perfusion imaging with ultrasound are retained within inflamed tissue because of complement-mediated attachment to leukocytes within venules. We hypothesized that incorporation of phosphatidylserine (PS) into the microbubble shell may enhance these interactions by amplifying complement activation and thereby allow ultrasound imaging of inflammation. In 6 mice, intravital microscopy of tissue necrosis factor-alpha-treated cremaster muscle was performed to assess the microvascular behavior of fluorescein-labeled lipid microbubbles with and without PS in the shell. Ten minutes after intravenous injection, microbubble attachment to leukocytes within inflamed venules was greater for PS-containing than for standard lipid microbubbles (20+/-4 versus 10+/-3 per 20 optical fields, P<0.05). The ultrasound signal from retained microbubbles was assessed in the kidneys of 6 mice undergoing renal ischemia-reperfusion injury and in 6 control kidneys. The signal from retained microbubbles in control kidneys was low (<2.5 video intensity units) for both agents. After ischemia-reperfusion, the signal from retained microbubbles was 2-fold higher for PS-containing than for standard lipid microbubbles (18+/-6 versus 8+/-2 video intensity units, P<0.05). An excellent relation was found between the ultrasound signal from retained microbubbles and the degree of renal inflammation, assessed by tissue myeloperoxidase activity. -We conclude that noninvasive assessment of inflammation is possible by ultrasound imaging of microbubbles targeted to activated leukocytes by the presence of PS in the lipid shell.

Role of type-1 fimbriae in the pathogenesis of chronic pyelonephritis in relation to reactive oxygen species.

The role of type-1 fimbriae in the pathogenesis of chronic pyelonephritis was studied for two Escherichia coli strains. Although both strains produced a similar total oxidative burst of chemiluminescence in macrophages from uninfected mice, the extracellular oxidative burst was greater with the non-fimbriate mutant E. coli BH-5 than its type-1 fimbriate parent E. coli 31-B. Moreover, macrophages from mice infected with the non-fimbriate mutant gave a much greater oxidative burst when stimulated with latex particles than that given by macrophages from mice infected with the type-1 fimbriate parent. These results correlated with the degree of renal inflammation and scarring as measured by malondialdehyde formation. Hence, the role of type-1 fimbriae in the pathogenesis of chronic UTI although documented does not appear to be significant.