Involvement of renal cytochrome P450 and arachidonic acid metabolites in diabetic nephropathy

Abstract

Background and Objectives: Diabetic nephropathy (DN), a serious complication of diabetes, is characterized by hyperfiltration, hypertrophy, extracellular matrix accumulation, fibrosis and proteinuria leading to loss of renal function. In renal hypertrophy, tubules increase in size and cause accumulation of the extracellular matrix, and are also associated with alterations in renal sodium handling as well as hypertension; processes linked by involvement of the arachidonic acid (AA) metabolites 20-HETE and EETs. This study aims to determine the specific AA-metabolizing CYP450 isoforms present in proximal tubules (PT) that are altered by high glucose (HG) in cultured PTs, and in an animal model of diabetes. It intends to investigate the effects of alterations in CYP isoforms and/or AA-metabolite levels in DN. This work will investigate the mechanism of PT injury and the effect of inhibition of AA-metabolites in vitro and will also provide insight into the cross-talk between CYP450 isoforms and other sources of reactive oxygen species (ROS). Methods: Immunohistochemistry, hypertrophy, apoptosis, fibrosis, ROS generation, 20-HETE and EET formation, CYP4A and Nox protein expression, and mRNA levels were measured in vitro and in vivo. Results: Exposure of PT cells to HG resulted in apoptosis and hypertrophy. HG treatment increased ROS production and was associated with CYP4A and CYP2C upregulation, 20-HETE and EETs formation, and Nox oxidases upregulation. The effects of HG on Nox proteins and mRNA expression, matrix protein accumulation and apoptosis were blocked by HET0016, an inhibitor of CYP4A, and were mimicked by 20-HETE. Inhibition of EETs in vitro promoted the effects of HG on cultured proximal tubular cells. In parallel, the levels of CYPs 2B, 2C, and 4A were assessed in a rat model of streptozotocin-induced diabetes. There was significant induction of expression and activity over control of these CYPs associated with an increase in ROS production, Noxs expression, PTs injury, and this was prevented by insulin therapy. Conclusion: Our results indicate that hyperglycemia in diabetes has a significant effect on the expression of AA-metabolizing CYPs, manifested by increased AA metabolism, and might thus alter kidney function through alteration of type and amount of AA metabolites; this pathway is through an oxidative stress-dependant mechanism.

mTORC2 Signaling Regulates Nox4-Induced Podocyte Depletion in Diabetes

“…Oxidative stress, particularly ROS derived from the NAD(P)H oxidases Noxs, is crucial for the pathogenesis of diabetic kidney disease (4,(16)(17)(18)(19)(20)(21)(22)29). The most abundant Nox isoform in the kidneys is Nox4 (3,24,25,28).…”

“…Twenty-two-week-old control FVB mice and OVE26 mice (FVB background; The Jackson Laboratory) were used. At 17 weeks of age, OVE26 mice were treated by either phosphorothioated sense or AS oligonucleotides for Rictor (90 ng$g body wt 21 …”

Vascular actions of 20-HETE

“…Several studies have established a role of 20-HETE in diabetic nephropathy [95–97]; however, the specific mechanisms involving 20-HETE and its effect on the vascular system leading to progression of this disease are not completely understood. In a streptozotocin-induced diabetic rat model, renal hypertrophy was associated with increased CYP4A expression and 20-HETE production, increased fibronectin and TGF-β1 expression, as well as increased ROS and NADPH oxidase activity [96]. Gangadhariah et al treated male mice with genetic disruption of Cyp4a14 with streptozotocin to induce diabetes [97].…”

The Metabolites of Arachidonic Acid in Microvascular Function

“…Similar to findings in hypertension and high salt diet, endothelial dysfunction in diabetes could also be related to the release of vasoconstrictor mediators, for example, increased production of 20-HETE leading to activation of ROS through an NAD(P)H-dependent pathway. This may have an important therapeutic potential in the treatment of diabetic vascular complications, for example, nephropathy [159].…”