Persistent Methylmalonate Overload Despite Glycemic Control Drives Mitochondrial Dyshomeostasis and Subclinical Heart Damage in Diabetes

Background Heart failure burden is still considerable in diabetes patients despite optimal glycemic control. Metabolic remodeling is involved in this process, but current understanding is still in its infancy. Methylmalonic acid (MMA) is a diagnostic marker of cobalamin (Cbl) deficiency in clinical practice. Paradoxically, MMA elevation-related cardiovascular mortality is more pronounced in diabetic patients with higher Cbl levels. We investigated the mechanisms underlying these contradictory features in diabetic heart and translational relevance. Methods We assessed serum Cbl, MMA, and cardiac biomarkers in 12,751 participants, and the Mmut protein (allosteric enzyme of MMA catabolism) in failing human hearts with diabetes and healthy donors. Cardiomyocyte-specific Mmut knockout and Mmut-overexpressing (delivered by AAV-Mmut with cTnT promoter) mice were subjected to high-fat diet/streptozotocin-induced diabetes. High glucose and/or palmitate stimulation of primary neonatal mouse ventricular cardiomyocytes (NMVMs) imitates diabetic conditions in vitro. Molecular mechanism was explored through RNA sequencing, immunoprecipitation, isotope tracing mass spectrometry and biolayer interferometry assays. Results Higher serum MMA was significantly associated with subclinical heart damage and poor outcomes among adults with diabetes in the absence of Cbl deficiency. Cardiac MMA overload and decreased protein expression of Mmut were observed in humans and mice with diabetes. Notably, MMA dysmetabolism in diabetic mice occurred before detectable cardiac damage and persisted even after glycemic control. Mechanistically, hyperglycemic memory-related molecule miR-499 binds to the mRNA of Mmut and inhibits its expression, leading to MMA overload under diabetes. Mmut knockout amplified cardiac MMA overload and exacerbated disturbances in glycolipid metabolism and mitochondrial quality control in diabetic hearts. Conversely, AAV-Mmut injection significantly reduces MMA load and alleviates the progression of cardiac remodeling in diabetic mice. Isotope tracing identified isoleucine and valine as the main sources of cardiac MMA under diabetes. A restricted-branched chain amino acid diet alleviated diabetes-induced MMA accumulation and heart damage. However, Cbl supplementation did not correct MMA overload in diabetic mice, even at high doses or in activated forms. Strikingly, both clinical and preclinical data showed metformin, a risk factor for Cbl deficiency, may mitigate MMA overload and related heart damage via dual mechanisms—activating AMPK-dependent mitochondrial quality control, enhancing tolerance to MMA insults; and direct Mmut-cobalamin cooperation, promoting MMA clearance independent of AMPK. Conclusions: This study provides a foundation for understanding diabetes-related MMA dysmetabolism as a trigger for subclinical heart damage resistant to glycemic control and Cbl supplementation. Our findings challenge the prevailing clinical consensus regarding the impacts of Cbl and metformin use on MMA elevation.