Myocardial glycophagy flux dysregulation and glycogen accumulation characterize diabetic cardiomyopathy.

Diabetic heart disease morbidity and mortality is escalating. No specific therapeutics exist and mechanistic understanding of diabetic cardiomyopathy etiology is lacking. While lipid accumulation is a recognized cardiomyocyte phenotype of diabetes, less is known about glycolytic fuel handling and storage. Based on in vitro studies, we postulated the operation of an autophagy pathway in the myocardium specific for glycogen homeostasis - glycophagy. Here we visualize occurrence of cardiac glycophagy and show that the diabetic myocardium is characterized by marked glycogen elevation and altered cardiomyocyte glycogen localization. We establish that cardiac glycophagy flux is disturbed in diabetes. Glycophagy may represent a potential therapeutic target for alleviating the myocardial impacts of metabolic disruption in diabetic heart disease.

Store-operated calcium entry and the localization of STIM1 and Orai1 proteins in isolated mouse sinoatrial node cells.

In many non-excitable and excitable cells, store-operated calcium entry (SOCE) represents an additional pathway for calcium entry upon Ca(2+) store depletion. In a previous study, we demonstrated SOCE activity in intact mouse cardiac pacemaker tissue, specifically from sinoatrial node (SAN) tissue. However, store content as a key determinant of SOCE activity is difficult to measure in intact SAN tissue. Therefore, to investigate the interaction between SOCE and store content and its role in cardiac pacemaking, it is necessary to investigate SOCE activity in single cardiac pacemaker cells. Furthermore, recent studies in other tissues have identified two new proteins involved in SOCE, stromal interacting molecule (STIM), which is an ER Ca(2+) sensor, and the surface membrane channel Orai, a prototypic gene encoding for SOCE. However, whether STIM and Orai are expressed in native pacemaker cells is still unknown. In this current study, we examined SOCE activity in single firing pacemaker cells isolated from mouse sinoatrial node tissue. We found a significant rise in Ca(2+) entry in response to Ca(2+) store depletion. SOCE blockers reduced the amplitude and frequency of spontaneous Ca(2+) transients and reduced Ca(2+) store content. We demonstrated for the first time that STIM and Orai are expressed in pacemaker cells. After store depletion, STIM1 redistributed to the cell periphery and showed increased co-localization with surface membrane located Orai1, indicating a possible involvement of these proteins in SOCE activity in native cardiac pacemaker cells. These results suggest the novel concept that SOCE plays a functional role in regulating intracellular Ca(2+) of cardiac pacemaker cells.

Beneficial effects of the synthetic antioxidant tert-butyl bisphenol on the hepatic microcirculation in a rat model of diabetes mellitus.

Diabetes mellitus is associated with oxidative injury to the vasculature. Here, the link between oxidative stress and ultrastructural changes in the hepatic microcirculation was investigated as well as the effects of a synthetic antioxidant, tert-butyl bisphenol (tBP). The study focused on the impact of experimental diabetes on liver sinusoidal endothelial cell (LSEC) fenestrations, which are pores in the liver endothelium that facilitate substrate transfer between blood and hepatocytes. Adult male rats were rendered diabetic using streptozotocin (60 mg/kg) and administered 1-2 IU insulin daily. After 8 weeks, animals received either 100 mg/kg tBP or vehicle alone, on 2 consecutive days. Livers were harvested 24 h later under isofluorane anaesthesia (5% v/v in O2(g) by inhalation) and fixed for scanning electron microscopy to evaluate fenestrations or for immuno-histochemical assessment of nitrotyrosine, a marker of nitrosative stress. Median fenestration diameter increased significantly following 8 weeks of diabetes (80 nm vs. 70 nm controls; P < 0.001). LSEC porosity increased by ~50% (P < 0.001). Treatment with tBP reversed these changes completely. Periportal nitrotyrosine staining was increased in diabetic livers, and this was abrogated by tBP, indicating that tBP reduced nitrosative stress in the liver. Early diabetes caused an increase in fenestration diameter and porosity. This was reversed by acute treatment with tBP, suggesting a link between nitrosative stress and regulation of liver endothelial fenestrations, and indicates that antioxidant therapy may protect the liver microvasculature against the effects of diabetes mellitus.