α_1D-Adrenergic Receptors and Mitogen-Activated Protein Kinase Mediate Increased Protein Synthesis by Arterial Smooth Muscle

Abstract

Catecholamines may influence vascular smooth muscle cell (SMC) growth and vascular hypertrophic diseases. We previously demonstrated that stimulation of alpha1-adrenoceptors (AR) causes hypertrophy of vascular SMCs in vitro and in situ. Here, we used adult rat aorta SMCs that express alpha1D- and alpha1B-ARs (but not alpha1A-ARs) in vitro to examine the mechanisms and alpha1-AR subtypes involved. Norepinephrine (NE) increased protein synthesis and content in a time- and dose-dependent manner. To identify the responsible alpha1-AR subtype, we first documented the selectivity of two alpha1-AR subtype antagonists, BMY 7378 (alpha1D-AR antagonist) and chloroethylclonidine (CEC; alpha1B-AR antagonist), using Rat-1 fibroblasts stably transfected with the three different rodent alpha1-AR cDNAs. NE dose-dependently increased protein synthesis in each cell line. In alpha1D fibroblasts, BMY 7378 inhibited growth and protected alpha1D-ARs from CEC alkylation while having little blocking or protecting effect on the growth induced by stimulation of fibroblasts that express alpha1A- or alpha1B-ARs. In rat aorta SMCs, pretreatment with CEC in the presence of BMY 7378 to protect alpha1D-ARs had no effect on NE-induced protein synthesis. BMY 7378 inhibited the SMC growth response with a pKb of 8.4. NE caused rapid and transient p42-p44 mitogen-activated protein kinase (MAPK) activation that was alpha1D-AR dependent. Furthermore, NE caused tyrosine phosphorylation of multiple cellular proteins, phosphorylation of Raf-1, and stimulation of c-fos mRNA expression in aorta SMCs. The selective MAPK kinase inhibitor PD 98059 inhibited NE-induced protein synthesis and MAPK activation with IC50 values of 2.3 and 1.6 microM, respectively. These data demonstrate that SMC growth induced by NE is mediated by alpha1D-ARs that couple to activation of the MAPK cascade.

α1-Adrenoceptor stimulation directly induces growth of vascular wall in vivo

“…In contrast, a small hypertrophic effect was evident in organ culture studies of uninjured rat aorta, where treatment with 1 mol/l NE (a much higher concentration than endogenous wall levels, albeit only presented for 48 h) caused media protein content to increase by 10% (no increase in cell number) and caused AFB cell number and protein to increase by 10% (38). This is consistent with the action of NE in the cell culture to cause dose-dependent (0.005-1 mol/l) hypertrophy of quiescent SMCs (37) and proliferation of quiescent AFBs (13) seen after as little as 12 h of NE exposure. It is possible that the different responses reflect differences in the in vitro and in vivo models or to duration of exposure and concentration of NE.…”

“…This, together with the much greater biological half-life of KMD-3213 (12 h; Ref. 1) than NE (several minutes), suggests that the NE concentration achieved in the injured carotid wall was likely to be Ͻ1 mol/l, which would be within the physiological range for dose-dependent, NE-mediated SMC and AFB trophic and chemotactic activity as determined in previous cell and organ culture studies (13,(37)(38)(39). In the present study, blockade of ␣ 1 -ARs with benoxathian, while significantly attenuating restenosis, had no effect on the uninjured carotid.…”

Norepinephrine-Induced Stimulation of p38 Mitogen-Activated Protein Kinase Is Mediated by Arachidonic Acid Metabolites Generated by Activation of Cytosolic Phospholipase A2 in Vascular Smooth Muscle Cells

“…Activation of ␣-ARs promotes vascular smooth muscle contraction, as well as proliferation, hypertrophy, and migration of VSMC (Nebigil and Malik, 1990;Hu et al, 1996;Nishio and Watanabe, 1997;Xin et al, 1997). Stimulation of both ␣-1 and ␣-2 ARs, which are coupled to pertussis toxin-sensitive G i␣ proteins, increases Ca 2ϩ influx through voltage-gated Ca 2ϩ channels in VSMC (Nebigil and Malik, 1993).…”

Sympathetic Vascular Tone in Human Obesity

“…In animal models, norepinephrine stimulates gene and protein expression in cultured vascular smooth muscle cells [Chen et al, 1995;Xin et al, 1997]. It is possible that sympathetically mediated vascular hypertrophy could partly be caused by α 1 -adrenergic receptor activation, which induces production of reactive oxygen through activation of NADPH oxidase, a mechanism that is inhibited by anti-oxidant treatment [Bleek et al, 2004].…”