Worsening Cardiac Autonomic Neuropathy on Progression to Type 2 Diabetes: Localized vs. Systemic Inflammation

Abstract

Cardiac autonomic neuropathy (CAN) occurs early in the course of diabetes with significant cardiovascular risks. While linked to hyperglycemia, current understanding extends CAN development to pre‐diabetes. The instigating cause of CAN, its temporal progression, and responsiveness to treatment remain ill‐defined. Here, we used a rat model of mild metabolic challenge with delayed development of hyperglycemia to trace these parameters in different disease stages.Male Sprague Dawley rats (5–6 weeks old) were fed a mild hypercaloric diet (4.035 vs. 3 KCal/g) leading to hyperinsulinemia after 12 weeks, representing the prediabetic stage. Type 2 diabetes was induced by two consecutive injections of low‐dose streptozotocin (40 mg/kg) at weeks 8 and 9. Drug treatment (metformin 100 mg/Kg/day, pioglitazone 2.5 mg/Kg/day, insulin titrated to control glucose levels, or their combination) was started at week 10. CAN was assessed by estimating baroreceptor sensitivity using the vasoactive method. ELISA, immunohistochemistry, and western blotting were used to examine markers of structural deterioration, inflammation, and autophagy in serum, heart, and brainstem.CAN was detected in the prediabetic state as blunted bradycardic reflex to phenylephrine stimulation reflected by a decrease in the slope of the DMAP vs. DHR line. It progressed with the transition to diabetes to further parasympathetic deterioration and de novo sympathetic insult. Interestingly, diabetic sympathetic dysfunction was readily reversed by insulin treatment. While prediabetic parasympathetic dysfunction responded to treatment with non‐hypoglycemic, anti‐inflammatory doses of metformin or pioglitazone, this was not the case in diabetic rats. Worsened parasympathetic function in diabetic rats was only restored by combination treatments, with insulin and Met/Pio. Notably, whereas prediabetic dysfunction was associated with localized perivascular adipose inflammation, the exaggerated CAN observed in diabetes was accompanied by significant systemic inflammation manifesting as elevated serum IL‐1b levels. This was concomitant with centralization of inflammation presenting in the brainstem along with microglial activation and autophagy suppression. Significantly, only combination treatment produced a considerable decrease in serum IL‐1b, consistent with their corrective effect on CAN in diabetes. Moreover, hyperglycemic rats showed signs of focal cardiac ischemia and left ventricular dysfunction which were ameliorated with combination treatments. In a cellular framework, we showed that systemic inflammation, rather than metabolic stress, induced neuronal damage. Differentiated PC12 cells simulating sympathetic neurons only replicated the central manifestations observed in diabetic rats when exposed to diabetic rat serum as opposed to the altered serum insulin, glucose, and lipid levels.Our results highlight a temporal framework whereby deteriorating metabolic state leads to exaggerated CAN possibly related to the progression of localized adipose inflammation to a systemic inflammatory state affecting brainstem and cardiac function.Support or Funding InformationFunding by AUB‐FM grant #320148