ISG15 signalling promotes coronary microvascular endothelial dysfunction in experimental diabetes

Objective
Diabetic cardiomyopathy (DCM) is a common complication of type 2 diabetes, which is characterised by dysfunction of the coronary microvascular endothelium as an important determinant of adverse cardiac remodeling. Whilst multiple signaling pathways are linked with DCM, specific mediators remain poorly understood. This study focused on interferon-stimulated gene 15 (ISG15) as a novel driver of endothelial dysfunction in experimental DCM and potential therapeutic target.

Methods and Results
Ingenuity Pathway Analysis (IPA) was performed on data obtained from both bulk and single nuclei RNAseq analyses of mouse DCM tissue using R and Partek. Data sets met the criteria of an adjusted p-value of 0.05 and a log2 fold change ranging from -1.5 to 1.5, focusing on differentially expressed genes with endothelial cell enrichment. Interferon signalling and activation of interferon-related factors were identified as the most significantly altered canonical pathways linked with endothelial dysfunction, with expression of several component genes found to be significantly upregulated within the dataset, including ISG15 which was validated by real time RT-PCR analysis of DCM mouse tissue. Parallel in vitro studies established treatment of human coronary microvascular endothelial cells (HCMECs) with 25 mmol/L D-glucose for 14 days (versus L-glucose osmotic control) as a reliable model of experimental diabetes, characterised by barrier dysfunction (increased FITC-Dextran transfer, reduced expression of e.g. ZO-1, claudin-5, β-catenin) and significantly increased ISG15 expression. ISG15 siRNA knockdown restored high glucose-induced HCMEC barrier dysfunction and expression of the key adherens junction protein, β-catenin, but had no impact on expression of tight junction proteins, ZO-1 and claudin-5.

Conclusion
Taken together, these data highlight ISG15 is an important driver of coronary microvascular endothelial dysfunction in experimental diabetes which may represent a potential therapeutic target to reduce progression of adverse cardiovascular remodelling.