Effect of intracellular angiotensin II on diabetic rat vascular smooth muscle cell proliferation

Background: The mechanism underlying vascular smooth muscle cell proliferation caused by diabetes and intracellular angiotensin II (Ang II) is not fully understood. It is also known that components of the renin angiotensin system in diabetes and hypertension increase intracellularly and systemically. Many studies have shown the role of systemic Ang II, angiotensin II type 1 receptor (ATR1) and angiotensin II type 2 receptor (ATR2) in smooth muscle cell proliferation. However, there has not been enough studies about the effect of angiotensin II, which is synthesized intracellularly by vascular smooth muscle cell on proliferation. Therefore, the effect of intracellular angiotensin II, ATR1 and ATR2 on diabetic rat vascular smooth muscle cell proliferation was investigated. Materials and Methods: We used Wistar albino male rats weighing 180-200 g as experimental animals. An experimental diabetic group was formed by administering streptozotocin 45 mg/kg intravenously via a tail vein of the rats. All experimental applications were made in accordance with the decisions of the Ethical Committee of the University of Çukurova Experimental Medicine Research and Application Center. Primary smooth muscle cell culture was performed from the aorta smooth muscle tissue of the diabetic and control group of rats. Some of the obtained cells were treated with Ang II (0,001 μM, 0,01 μM, 0,1 μM, 1 μM, 10 μM, 100 μM and 1000 μM), ATR1 antagonist Olmesartan (1 μM) and ATR2 antagonist PD123,319 (1 μM) for 24 hours to evaluation of cell proliferation and viability spectrophotometrically with 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT). Ang II level was measured in cell medium and cell lysate by ELISA kit. Findings: In normal, diabetic and high glucose treated vascular smooth muscle cell (VDKH), 0,1 μM and 0,01 μM Ang II application significantly increased cell proliferation. In our study, Ang II, Ang II+PD123,319, Olmesartan+PD123,319 and Ang II+Olmesartan+PD123,319 resulted in a significant increase in cell proliferation whereas Ang II+Olmesartan administration significantly decreased cell proliferation in normal VDKH. However, there was no decrease in Ang II+Olmesartan administration in VDKH with diabetic and high glucose. Intracellular Ang II was detected with ELISA in VDKH and medium with normal, diabetic and high glucose. In our study, Ang II levels measured in normal, diabetic and hyperglucose intracellular and cell media were calculated as mg/protein, but no significant difference was found between normal group and diabetic group. Conclusion: Although there was an increased tendency in diabetic VDKH proliferation in our study, the difference between intracellular and extracellular Ang II levels compared to the control group suggests that there is no relationship between diabetes proliferation, Ang II levels and ATR. Key Words: Angiotensin II, diabetes, smooth muscle cell, intracellular, proliferation.