Sitagliptin attenuates arterial calcification by downregulating oxidative stress-induced receptor for advanced glycation end products in LDLR knockout mice

Chih-Pei Lin,Jia-Shiong Chen,Po-Hsun Huang,Shing-Jong Lin,Chi-Yu Chen,Jaw-Wen Chen,Meng-Yu Wu

doi:10.1038/s41598-021-97361-w

Abstract

Diabetes is a complex disease characterized by hyperglycemia, dyslipidemia, and insulin resistance. Plasma advanced glycation end products (AGEs) activated the receptor for advanced glycation end products (RAGE) and the activation of RAGE is implicated to be the pathogenesis of type 2 diabetic mellitus (T2DM) patient vascular complications. Sitagliptin, a dipeptidyl peptidase-4 (DPP4) inhibitor, is a new oral hypoglycemic agent for the treatment of T2DM. However, the beneficial effects on vascular calcification remain unclear. In this study, we used a high-fat diet (HFD)-fed low-density lipoprotein receptor deficiency (LDLR−/−) mice model to investigate the potential effects of sitagliptin on HFD-induced arterial calcification. Mice were randomly divided into 3 groups: (1) normal diet group, (2) HFD group and (3) HFD + sitagliptin group. After 24 weeks treatment, we collected the blood for chemistry parameters and DPP4 activity measurement, and harvested the aorta to evaluate calcification using immunohistochemistry and calcium content. To determine the effects of sitagliptin, tumor necrosis factor (TNF)-α combined with S100A12 was used to induce oxidative stress, activation of nicotinamide adenine dinucleotide phosphate (NADPH), up-regulation of bone markers and RAGE expression, and cell calcium deposition on human aortic smooth muscle cells (HASMCs). We found that sitagliptin effectively blunted the HFD-induced artery calcification and significantly lowered the levels of fasting serum glucose, triglyceride (TG), nitrotyrosine and TNF-α, decreased the calcium deposits, and reduced arterial calcification. In an in-vitro study, both S100A12 and TNF-α stimulated RAGE expression and cellular calcium deposits in HASMCs. The potency of S100A12 on HASMCs was amplified by the presence of TNF-α. Sitagliptin and Apocynin (APO), an NADPH oxidase inhibitor, inhibited the TNF-α + S100A12-induced NADPH oxidase and nuclear factor (NF)-κB activation, cellular oxidative stress, RAGE expression, osteo transcription factors expression and calcium deposition. In addition, treatment with sitagliptin, knockdown of RAGE or TNF-α receptor blunted the TNF-α + S100A12-induced RAGE expression. Our findings suggest that sitagliptin may suppress the initiation and progression of arterial calcification by inhibiting the activation of NADPH oxidase and NF-κB, followed by decreasing the expression of RAGE.

Highlights

Diabetes is a complex disease characterized by hyperglycemia, dyslipidemia, and insulin resistance
It is suggested that the activation of nuclear factor (NF)-κB led to activation of receptor for advanced glycation end products (RAGE), which led to a wide spectrum of pathological inflammation conditions and various diseases such as atherosclerosis[20]
We found that the high-fat diet (HFD)-fed LDLR−/− mice had a significantly higher body weight than the regular chow-fed mice (Fig. 1a; 33.8 ± 1.9 vs. 41.8 ± 2.3 g)

Summary

Introduction

Diabetes is a complex disease characterized by hyperglycemia, dyslipidemia, and insulin resistance. Abbreviations AGEs Advanced glycation end products APO Apocynin ApoE Apolipoprotein-E BMP-2 Bone matrix protein-2 T2DM Type 2 diabetes mellitus DPP4 Dipeptidyl peptidase-4 LDLR Low-density lipoprotein receptor MSX2 Msh homeobox 2 NADPH Nicotinamide adenine dinucleotide phosphate Nox Nicotinamide adenine dinucleotide phosphate oxidase NF-κB Nuclear factor (NF)-κB ROS Reactive oxygen species RAGE Receptor for advanced glycation end products RUNX2 Runt-related transcription factor 2 VSMCs Vascular smooth muscle cells TNF Tumor necrosis factor GLP-1 Type I glucagon-like peptide. The activation of NF-κB was reported to upregulate RAGE expression and this positive feedback loop between RAGE and NF-κB signaling pathway results in a perpetuation of inflammation stats[21] This RAGE/NF-κB interaction theory explained why oxidative stress caused by ureteral obstruction only caused severe arterial calcification in transgenic S100A12 C57BL/6J mice but not the wild-type animals[19]. RAGE-S100A12 signaling is reported to cause coronary artery diseases and vascular calcification[23,24]

Methods

Results

Conclusion