Influence of non-vascular cells in accelerated coronary aging in diabetes

2021 Fiscal Year Annual Research Report

• Project/Area Number: 19H03405
• Research Institution: National Cardiovascular Center Research Institute
• Principal Investigator: Pearson James 国立研究開発法人国立循環器病研究センター, 研究所, 部長 (30261390)
• Co-Investigator(Kenkyū-buntansha): 土持 裕胤 国立研究開発法人国立循環器病研究センター, 研究所, 室長 (60379948) ; 曽野部 崇 国立研究開発法人国立循環器病研究センター, 研究所, 室長 (70548289) ; 岩田 裕子 国立研究開発法人国立循環器病研究センター, 研究所, 室長 (80171908)
• Project Period (FY): 2019-04-01 – 2023-03-31
• Keywords: coronary / senescence / diabetes / inflammation / oxidative stress

Outline of Annual Research Achievements

Diabetes accelerates vascular ageing when excess ROS exceed antioxidant capacity, leading to SASP, vascular dysfunction and vessel rarefaction. Therefore, we tested whether Nox2 contributes to coronary dysfunction in diet induced early-stage diabetes (B6D2 hybrid mice) induced by chronic high fat diet (HFD) and increased salt intake (1% NaCl water) in mice treated with and without apocynin (Nox2 inhibitor). Synchrotron microangiography revealed that the coronary capacity to produce NO was diminished with insulin resistance onset, but was potentiated by apocynin treatment, suggesting that Nox2 overactivation in insulin resistance reduces NO bioavailability. To investigate SASP activation we used CRISPR/Cas9 to generate Nfkbib gene deletion mice that are unable to activate NFkB signaling.

Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

In SAMP8 mice glycolysis was inhibited and abnormal purine metabolism increased xanthine oxidase activation and ROS generation relative to SAMR1 on HFD, leading to SASP activation and microvascular dysfunction. In the absence of insulin resistance, Nox2 activation was essential for NO mediated vasodilation and limiting senescence progression. We characterised diet induced insulin resistance with transcriptome (RNAseq) and proteome profiling. B6D2 mice was found to develop mild coronary dysfunction due to elevated ROS production, metabolic inflexibility and mitochondrial dysfunction. We found that insulin resistance greatly increases ROS through Nox2 to reduce NO bioavailability. Experiments in May will establish if Nox1 activation contributes also to exacerbation of ROS in this model.

Strategy for Future Research Activity

Having now confirmed our novel diabetic mouse model has metabolic dysregulation and coronary dysfunction we will investigate the molecular mechanisms underlying abnormal purine metabolism, mitochondrial dysfunction and DNA damage. We will also utilize our Nfkbib deletion mouse exposed to HFD to examine the role of NF-kB/p53 activation in non-vascular cells accelerating vascular ageing associated with diabetes. We predict Nfkbib deletion will prevent SASP activation that ages coronary vessels. We established cell co-culture to investigate the roles of specific cardiac cell types in accelerated ageing with diet-induced insulin resistance and Nfkbib deletion mice. Quantitative PCR will be used to determine gene expression changes in endothelial and non-vascular cells in these mouse models.

Research Products

• [Journal Article] Using synchrotron radiation imaging techniques to elucidate the actions of hexarelin in the heart of small animal models (2021)
  • Author(s): Mark T Waddingham, Hirotsugu Tsuchimochi, Takashi Sonobe, Ryotaro Asano, Huiling Jin, Connie PC Ow, Daryl O Schwenke, Rajesh Katare, Kohki Aoyama, Keiji Umetani, Masato Hoshino, Kentaro Uesugi, Mikiyasu Shirai, Takeshi Ogo, James T Pearson
  • Journal Title: Frontiers in Physiology Volume: 12 Pages: 766818
  • DOI: 10.3389/fphys.2021.766818
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Presentation] Exercise reverses the sarcomeric and cytoskeletal contributions to diastolic dysfunction in mice with diet-induced insulin resistance and obesity (2022)
  • Author(s): James Pearson
  • Organizer: 第９9回日本生理学会大会