Comparative proteomic analysis of cardiac mitochondria-associated reticulum membranes in type 2 diabetes
Authors: Dia M , Givre L , Leon C , Chouabe C , Chanon S , Rieusset J , Thibault H , Paillard M 
Published: June 2022

Abstract

Funding Acknowledgements
Type of funding sources: Public grant(s) – National budget only. Main funding source(s): INSERM U1060

Introduction
The prevalence of type 2 diabetes (T2D) is increasing worldwide with cardiovascular complications being a leading cause of T2D-related morbi-mortality, notably diabetic cardiomyopathy (DCM). The involvement of the mitochondria-associated reticular membranes (MAM) in diabetic cardiomyopathy starts to be demonstrated, notably for Ca²+ regulation of cellular processes. However, while the MAM protein composition was studied in several organs, none was conducted on cardiac MAM.

Objective
We aim to characterize the cardiac MAM proteome by an exhaustive mass spectrometry-based proteomics together with the effect of early T2D using two well-established obesogenic diabetic mouse models (high-fat high-sucrose diet, HFHSD, and leptin-deficient ob/ob).

Methods
12 weeks old ob/ob mice versus 16 weeks HFHSD-fed mice were characterized at the subcellular level. MAM were enriched by subcellular fractionation, then protein compostion was assessed by mass spectrometry. Uniprot and Panther softwares were used to study the MAM proteomes. At the cellular level, mitochondrial calcium transfer was assessed using a mitochondrial Ca²+ sensor, while contractile function was evaluated by studying cardiomyocyte Ca²+ transients and echocardiography.

Results
Compartmental analysis of MAMs showed an enrichment of proteins from mitochondria, reticulum, cytoskeleton and plasma membrane. On the functional level, molecular and biological annotations revealed that cardiac MAM presents mainly enzymes and transporters, associated to 4 main biological processes: cellular and metabolic processes, localization and cellular component organization. Both the cellular response to stress and the lipid metabolism processes were found upregulated in both diabetic cardiac MAM. While Ca²+ transport was downregulated in the HFHSD MAM, this process was upregulated in the ob/ob MAM. However, no change in the histamine-induced reticulum-mitochondria Ca²+ transfer was observed in the ob/ob cardiomyocytes. Contrary to the HFHSD mice, the ob/ob mice did not exhibit any cardiac hypertrophy, by echocardiography or electrophysiology. Echocardiography further revealed no diastolic dysfunction in the ob/ob mice but a mild strain rate reduction with preserved ejection fraction.

Conclusion
Our data investigate the protein composition of the cardiac MAM and unravel the main alterations of the MAM proteome in ob/ob model versus our HFHSD results. Importantly, while being two models of early diabetic cardiomyopathy, the MAM proteome is altered differently, emphasizing on the complexity of diabetic animal models. Indeed, these ob/ob mice, recognized as a T2D and obese mouse model, do not recapitulate the main hallmarks of DCM, i.e. diastolic dysfunction and cardiac hypertrophy, contrary to the heart failure with preserved ejection fraction (HFpEF) observed in the HFHSD mice, yet the lipotoxicity in the ob/ob cardiomyocytes may contribute to the early systolic dysfunction.