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Effect of Aging on Vascular Smooth Muscle Contractility in Skeletal Muscle Resistance Arteries
Abstract
Vasoconstrictor function, which is essential for vascular homeostasis, declines with aging. Aging induces molecular alterations, leading to the gradual loss of vascular homeostasis. Thus, aging is a significant risk factor for the development of cardiovascular disease (CVD), the leading cause of deaths globally. The mechanisms for age-associated changes in vascular function have not been fully elucidated. A number of studies have shown that Rho-kinase plays an important role in vascular smooth muscle (VSM) contraction, modulating the level of phosphorylation of the myosin light chain via inhibition of myosin phosphatase, thus yielding smooth muscle contraction. A previous study showed that age-associated reduction in vascular contractility was due to an impaired Rho-kinase pathway. However, impaired Rho-kinase pathway does not fully account for reduced contractility. The other mechanism this dissertation investigates is integrins since they play crucial roles in regulating many cellular functions including arterial contractility but the effect of aging on integrin function in VSM cells is not fully elucidated. Therefore, the purpose of this dissertation was to: 1) determine the effect of aging on vasoconstrictor responses to mechanical stimulus in rat soleus muscle feed arteries (SFA); 2) investigate the effect of aging on receptor-mediated vascular contractility in rat SFA; 3) determine the effect of aging on integrin-mediated vasoconstriction; 4) elucidate age effects on the architecture of VSM cells. To investigate vasoconstrictor function with aging in SFA were isolated from young (4 months) and old (24 months) male Fischer 344 rats. Isolated SFA were cannulated with glass micropipettes and pressurized at 90 cmH2O. To determine the role of VSM, some arteries were denuded (endothelial cells removed) by passing 5 ml of air through the lumen of artery. Myogenic responses were assessed by using step-increases in pressure up to 135 cmH2O in 15cm increments and step-decreases in pressure down to 45 cmH2O. Agonist-induced vasoconstrictor responses were assessed by adding of norepinephrine (NE; 10^-9-10^-4 M), angiotensin II (Ang II; 10^-11-10^-7 M), or phenylephrine (PE; 10^-9-10^-4 M). To determine the role of integrins, constrictor responses were assessed in the absence or presence of RGD, an integrin inhibitory peptide. RGE, a non-inhibitory peptide was used as control. To investigate changes in integrin expression and recruitment at focal adhesions with age, real-time quantitative polymerase chain reaction (RT-qPCR), cell adhesion assays, and total internal reflection fluorescence (TIRF) imaging were performed using VSM cells isolated from young and old SFA.
Old intact SFA showed impaired constrictor responses to NE, PE, and Ang II compared to young SFA. In the presence of the RGD-inhibitory peptide, constrictor responses to all three agonists were reduced in both young and old SFA. Old denuded SFA had reduced vasoconstrictor responses to NE, PE, and Ang II relative to young denuded SFA. In the presence of RGD, constrictor responses to Ang II were significantly inhibited in old and young SFA. Constrictor responses to NE and PE were not significantly altered by RGD. RGE-non-inhibitory peptide did not affect constrictor responses in any groups. Specific integrin gene expression was reduced in old VSM cells as compared to young VSM cells. Additionally, RGD-treated VSM cells demonstrated decreased adhesion to fibronectin in both young and old VSM cells compared to RGE non-inhibitory peptide. Quantitative analysis of TIRF images revealed that recruitment of integrin α5 and β3 was diminished in old VSM cells while β1 was not altered with age. Taken together, these data indicate that aging impairs vasoconstrictor function in rat SFA, and the impairment was due in part to integrin dysfunction, which leads to decreased contractile properties of VSM in SFA.
Citation
Shin, Song Yi (2022). Effect of Aging on Vascular Smooth Muscle Contractility in Skeletal Muscle Resistance Arteries. Doctoral dissertation, Texas A&M University. Available electronically from https : / /hdl .handle .net /1969 .1 /198605.