Amlodipine modulates THP-1 cell adhesion to vascular endothelium via inhibition of protein kinase C signal transduction.
Inflammatory responses play an important role in atherosclerosis. To critically assess the effect of dihydropyridines in inflammatory reactions, we conducted a monocyte-endothelial adhesion assay with monocytic THP-1 cells treated with amlodipine under flow conditions in vitro. THP-1 cells were incubated in the presence of amlodipine (10 micromol/L) for 48 hours and then perfused over activated (interleukin-1beta, 10 U/mL, 4 hours) human umbilical vein endothelial cells. The adhesion of THP-1 cells was significantly reduced after amlodipine treatment (P<0.001); however, flow cytometric analysis reveled that the expression levels of integrins in THP-1 cells were not significantly altered. Furthermore, Western blotting analysis of THP-1 cell lysates revealed that translocation of RhoA from the cytosol to the membrane was significantly diminished after amlodipine treatment. In addition, activation of protein kinase C-alpha and -beta, as well as intracellular calcium influx, induced by phorbol 12-myristate 13-acetate, was diminished after amlodipine treatment. Pretreatment of THP-1 cells with calphostin C, a potent inhibitor of protein kinase C, significantly reduced THP-1 adhesion to vascular endothelium, whereas activation of beta1-integrin was reduced after amlodipine treatment in THP-1 cells, based on the immunoreactivity of an activation-specific antibody for beta1-integrin. Similar inhibitory effects were observed when we used freshly isolated peripheral blood mononuclear cells. These findings suggest a potential role for amlodipine in monocyte-endothelial interactions by modulation of protein kinase C- and RhoA-dependent mechanisms, which might account for its vascular protective effects.
Yu
Tao
T
Department of Medical Biochemistry, Graduate School of Medicine, Tokyo Medical and Dental University, Tokyo, Japan.
Morita
Ikuo
I
Shimokado
Kentaro
K
Iwai
Takehisa
T
Yoshida
Masayuki
M
D017311:Amlodipine / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002448:Cell Adhesion / Q000187:drug effects; D002460:Cell Line; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D019012:Integrin beta1 / Q000096:biosynthesis; D016023:Integrins / Q000096:biosynthesis; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009000:Monocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
D019012:Integrin beta1; D016023:Integrins; D017311:Amlodipine; D011493:Protein Kinase C; D020742:rhoA GTP-Binding Protein; D002118:Calcium