Endocrine Abstracts

Background: Sodium glucose co-transporter 2 inhibitors reduce the incidence of cardiovascular events in patients with type 2 diabetes mellitus based on the results of recent cardiovascular outcome studies. Herein, we investigated the effects of long-term treatment with empagliflozin on biochemical and immunohistochemical markers related to atherosclerosis development in apolipoprotein E knockout [Apo-E^(−/−)] mice.

Methods: At the age of 5 weeks, mice were switched from normal to high-fat diet. After 5 weeks, Apo-E^(−/−) mice were divided into control-group (n=10) treated with 0.5% hydroxypropyl methylcellulose and Empa-group (n=10) treated with Empagliflozin (10 mg/kg/day) per os. After 10 weeks of intervention, animals were culled, and heart and aorta were removed. Sections stained with hematoxylin–eosin (H&E) were used for histomorphometry. Immunohistochemistry was performed to evaluate protein expression of MCP-1, CD68, α-SMA, MMP-2, MMP-9, TIMP-1, ICAM-1 and VCAM-1. mRNA expression of matrix metalloproteinases and their inhibitors, ICAM-1, VCAM-1, IL-6, MCP-1 was measured by q-PCR. Blood pressure, heart rate and biochemical indices were measured at the onset and by the end of the intervention in both groups.

Results: Empa-group mice had lower glucose levels and HDL-cholesterol (P<0.01), while totalcholesterol, LDL and triglycerides did not change significantly compared to control group. Diastolic blood pressure and heart rate were significantly lower (P≤0.01 and P< 0.05 respectively) in Empa-group compared to control-group. Histomorphometry of the aorta root revealed that 6 in 10 Empa-group mice versus 8 in 10 control mice developed atheromatosis. The lumen area was wider (approximately 50%) in Empa-group compared with control-group reaching statistical significance (P=0.06). Empagliflozin administration significantly reduced VCAM-1 and MCP-1 mRNA levels (P<0.05 and P≤0.01 respectively) while marginally induced TIMP-1 and TIMP-2 mRNA expression (P≤0.08 and P=0.1 respectively). There were no significant alterations in IL-6, ICAM-1, MMP-2 and MMP-9 mRNA levels compared to controls; however, TIMP-1/MMP-2 mRNA ratio was significantly higher in Empa-group (P<0.05) compared to control-group. Immunohistochemistry staining of aortic root sections revealed that treatment with Empagliflozin led to border line increase in TIMP-1 (P=0.1) and marginal reduction in VCAM-1 and MMP-9 (P=0.1) without affecting the expression of ICAM-1, MMP-2, TIMP-2, MCP-1, CD68 and α-SMA in atherosclerotic lesions.

Conclusions: Empagliflozin attenuates the progression of atherosclerosis, reducing (1) hyperglycemia, and (2) inflammatory process by lowering the expression of inflammatory molecule VCAM-1. Moreover, it increased TIMP-1/MMP-2 mRNA ratio expression possibly leading to increased plaque stability.