Renal hypertrophy and extracellular matrix accumulation are early features of diabetic nephropathy. We investigated the role of the NAD(P)H oxidase Nox4 in generation of reactive oxygen species (ROS), hypertrophy, and fibronectin expression in a rat model of type 1 diabetes induced by streptozotocin. Phosphorothioated antisense (AS) or sense oligonucleotides for Nox4 were administered for 2 weeks with an osmotic minipump 72 h after streptozotocin treatment. Nox4 protein expression was increased in diabetic kidney cortex compared with non-diabetic controls and was downregulated in AS-treated animals. AS oligonucleotides inhibited NADPH-dependent ROS generation in renal cortical and glomerular homogenates. ROS generation by intact isolated glomeruli from diabetic animals was increased compared with glomeruli isolated from AS-treated animals. AS treatment reduced whole kidney and glomerular hypertrophy. Moreover, the increased expression of fibronectin protein was markedly reduced in renal cortex including glomeruli of AS-treated diabetic rats. Akt/protein kinase B and ERK1/2, two protein kinases critical for cell growth and hypertrophy, were activated in diabetes, and AS treatment almost abolished their activation. In cultured mesangial cells, high glucose increased NADPH oxidase activity and fibronectin expression, effects that were prevented in cells transfected with AS oligonucleotides. These data establish a role for Nox4 as the major source of ROS in the kidneys during early stages of diabetes and establish that Nox4-derived ROS mediate renal hypertrophy and increased fibronectin expression.

BMP-7, a member of the bone morphogenic protein subfamily (BMPs) of the transforming growth factor-β superfamily of secreted growth factors, is abundantly expressed in the fetal kidney. The precise role of this protein in renal physiology or pathology is unknown. A cDNA that encodes rat BMP-7 was cloned and used as a probe to localize BMP-7 mRNA expression by in situ hybridization in the adult rat kidney. The highest expression of BMP-7 mRNA could be seen in tubules of the outer medulla. In glomeruli, a few cells, mainly located at the periphery of the glomerular tuft, showed specific and strong signals. Also, high BMP-7 mRNA expression could be localized to the adventitia of renal arteries, as well as to the epithelial cell layer of the renal pelvis and the ureter. Preliminary evidence suggests that BMP-7 enhances recovery when infused into rats with ischemia-induced acute renal failure. We examined BMP-7 mRNA expression in kidneys with acute renal failure induced by unilateral renal artery clamping. BMP-7 mRNA abundance as analyzed by solution hybridization was reduced in ischemic kidneys after 6 and 16 h of reperfusion compared with the contralateral kidney. In situ hybridization in ischemic kidneys showed a marked decrease of BMP-7 mRNA in the outer medulla and in glomeruli. Utilizing rat metanephric mesenchymal cells in culture, we also demonstrate that BMP-7 induces epithelial cell differentiation. Taken together, these data suggest that BMP-7 is important in both stimulating and maintaining a healthy differentiated epithelial cell phenotype.

The changes in glucose and potassium are consistent with the magnitude and duration of the temperature difference between the flown and stationary samples. Long drone flights of biological samples are feasible but require stringent environmental controls to ensure consistent results.

These data indicate that mesangial cells are susceptible to the effects of Stx1 in vitro. Stx1 exerts a spectrum of biologic effects on mesangial cells ranging from activation of chemokine genes to a lethal toxic injury. Immunoinflammatory cytokines potentiate the effects of Stx1. Thus, glomerular pathology in HUS may also result from a direct effect of Stx1 on mesangial cells.

Eosinophils have been widely investigated in asthma and allergic diseases. More recently, new insights into the biology of these cells has illustrated eosinophils contribute to homeostatic functions in health such as regulation of adipose tissue glucose metabolism. Human translational studies are limited by the difficulty of obtaining cells taken directly from their tissue environment, relying instead on eosinophils isolated from peripheral blood. Isolation techniques for tissue-derived eosinophils can result in unwanted cell or ribonuclease activation, leading to poor cell viability or RNA quality, which may impair analysis of effector activities of these cells. Here we demonstrate a technique to obtain eosinophils from human adipose tissue samples for the purpose of downstream molecular analysis. From as little as 2 g of intact human adipose tissue, greater than 10 4 eosinophils were purified by fluorescence-activated cell sorting (FACS) protocol resulting in ≥ 99% purity and ≥ 95% viable eosinophils. We demonstrated that the isolated eosinophils could undergo epigenetic analysis to determine differences in DNA methylation in various settings. Here we focused on comparing eosinophils isolated from human peripheral blood vs human adipose tissue. Our results open the door to future mechanistic investigations to better understand the role of tissue resident eosinophils in different context.