Diabetic eNOS knockout mice develop distinct macro- and microvascular complications

Abstract

Functional consequences of impaired endothelial nitric oxide synthase (eNOS) activity causing organ-specific abnormalities on a diabetic setting are not completely understood. In this study, we extensively characterized a diabetic mouse model (lepr db/db ) in which eNOS expression is genetically disrupted (eNOS À/À ). The eNOS À/À / lepr db/db double-knockout (DKO) mice developed obesity, hyperglycemia, hyperinsulinemia and hypertension. Analysis of tissues from DKO mice showed large islets in the pancreas and fat droplets in hepatocytes. Interestingly, the aorta was normal and atherogenic lesions were not observed. Abnormalities in the aorta including poor re-endothelialization and increased medial wall thickness were evident only in response to deliberate injury. In contrast, significant glomerular capillary damage in the kidney was identified, with DKO mice demonstrating a robust diabetic nephropathy similar to human disease. The vascular and renal impairments in DKO mice were pronounced despite lower fasting plasma glucose levels compared to lepr db/db mice, indicating that eNOS is a critical determinant of hyperglycemia-induced organ-specific complications and their severity in diabetes. Results provide the first evidence that absence of eNOS in diabetes has a greater deleterious effect on the renal microvasculature than on the larger aortic vessel. The DKO model may suggest novel therapeutic strategies to prevent both vascular and renal complications of diabetes. KEYWORDS: diabetes; endothelial dysfunction; eNOS; micro-and macrovasculature; nephropathy and vasculopathy Macro-and microangiopathy including accelerated atherosclerosis and nephropathy are the most common complications of diabetes. Reduced bioavailabilty of nitric oxide (NO), due to impaired nitric oxide synthase (NOS) activity, is implicated in the generation of these abnormalities. 1-3 NO is formed from L-Arginine by three isoforms of NOS with stoichiometric production of L-citrulline. Among the three isoforms, endothelial NOS (eNOS) is constitutively expressed predominantly in the macrovascular endothelium and along the renal microvascular tree. Basal release of eNOS-driven NO by endothelial cells contributes to the maintenance of normal vasodilatory tone and thromboresistance. Under conditions of hyperglycemia and associated increase in advanced glycation end products, there is excessive generation of superoxide, which interacts with NO resulting in the formation of peroxynitrites. The peroxynitrites oxidize tetrahydrobiopterin, an important cofactor involved in normal eNOS activity. The deficiency of tetrahydrobiopterin causes eNOS uncoupling, which results in increased generation of superoxide rather than NO. Overall, superoxide generation is amplified in macrovessels in diabetes with concomitant reduction in the level of bioavailable NO.Nitric oxide also acts as a potent modulator of renal function and controls both afferent and efferent vascular tone, glomerular ultrafiltration coefficient 4,5 and medullary blood flow. 6 However, the pr...

Chronic Hyperphosphatemia and Vascular Calcification Are Reduced by Stable Delivery of Soluble Klotho

“…[41][42][43] The db/db-eNOS 2/2 model serves as a useful tool in studying therapeutic interventions for CKD-MBD phenotypes evident in DN. 26,27 On histologic characterization of these mice, it was confirmed that the db/db-eNOS 2 /2 animals exhibited a prominent presence of tubular protein (arrowheads and inset in Figure 1A, iv) and dilated tubules (arrows in Figure 1A, iv) with undulation of the subcapsular cortex due to underlying fibrosis. Noticeable mesangial matrix deposition obliterating normal capillary loops and cellularity in the glomeruli (arrows and inset in Figure 1A, v) was observed in db/db-eNOS 2/2 mice compared with control lean mice ( Figure 1A, i-iii).…”

“…With the loss of activity of the leptin receptor and disruption of eNOS, these animals develop highly elevated blood glucose and progressive renal damage, including glomerular and interstitial fibrosis. 26,27 Because DN is now the leading cause of CKD-MBD, 28,29 understanding the nature of this syndrome is important for identifying potential novel management approaches. The aKL-and Fgf23-knockout mice share key phenotypes associated with CKD-MBD, including uncontrollably elevated serum phosphate as well as ectopic calcification and VC.…”

Structural profiling of endogenous S-nitrosocysteine residues reveals unique features that accommodate diverse mechanisms for protein S-nitrosylation

“…Leptin is an adipocytesecreted hormone that primarily acts on the central nervous system to regulate energy homeostasis. Leptin also regulates liver metabolism, evident by the significant accumulation of lipids (fatty liver) in mice deficient in leptin (ob/ob) (25,26) or leptin long-form receptor (db/db) (27). The regulation of liver metabolism is attributed to the leptin-dependent repression of liver stearoyl-CoA desaturase-1, the rate limiting step in monosaturated fat biosynthesis (28).…”

Gasotransmitters in Vascular Complications of Diabetes

“…The important role of NO in the macrovasculature is also shown in animal models of diabetes. Lepr db/db eNOS 2/2 double knockout mice showed an aggravated vascular phenotype compared with diabetic Lepr db/db or eNOS 2/2 single knockouts, as evidenced by an increased aortic wall thickness and reduced re-endothelialization after arterial injury (117). In ApoE 2/2 mice and mice with STZinduced type 1 diabetes, treatment with bone marrowderived mononuclear cells overexpressing eNOS resulted in reduced plaque progression and improved postischemic neovascularization, an effect that was completely inhibited by NOS inhibitor L-N G -nitro-L-arginine methyl ester (L-NAME) (118).…”