Increased Migration of Monocytes in Essential Hypertension Is Associated with Increased Transient Receptor Potential Channel Canonical Type 3 Channels

et al. (2012) Increased Migration of Monocytes in Essential Hypertension Is Associated with Increased Transient Receptor Potential Channel Canonical Type 3 Channels. PLoS ONE 7(3): e32628. doi:10.1371/journal.pone.0032628 Increased Migration of Monocytes in Essential Hypertension Is Associated with Increased Transient Receptor Potential Channel Canonical Type 3 Channels Zhigang Zhao 0 Yinxing Ni 0 Jing Chen 0 Jian Zhong 0 Hao Yu 0 Xingsen Xu 0 Hongbo He 0 Zhencheng Yan 0 Alexandra Scholze 0 Daoyan Liu 0 Zhiming Zhu 0 Martin Tepel 0 Harry Mellor, University of Bristol, United Kingdom 0 1 Department of Hypertension and Endocrinology, Center for Hypertension and Metabolic Diseases, Daping Hospital, Third Military Medical University, Chongqing Institute of Hypertension , Chongqing , China , 2 Department of Nephrology , Charite , Berlin , Germany; and University of Southern Denmark, Institute for Molecular Medicine , Odense , Denmark Increased transient receptor potential canonical type 3 (TRPC3) channels have been observed in patients with essential hypertension. In the present study we tested the hypothesis that increased monocyte migration is associated with increased TRPC3 expression. Monocyte migration assay was performed in a microchemotaxis chamber using chemoattractants formylated peptide Met-Leu-Phe (fMLP) and tumor necrosis factor-a (TNF-a). Proteins were identified by immunoblotting and quantitative in-cell Western assay. The effects of TRP channel-inhibitor 2-aminoethoxydiphenylborane (2-APB) and small interfering RNA knockdown of TRPC3 were investigated. We observed an increased fMLP-induced migration of monocytes from hypertensive patients compared with normotensive control subjects (246614% vs 151610%). The TNF-ainduced migration of monocytes in patients with essential hypertension was also significantly increased compared to normotensive control subjects (221620% vs 138618%). In the presence of 2-APB or after siRNA knockdown of TRPC3 the fMLP-induced monocyte migration was significantly blocked. The fMLP-induced changes of cytosolic calcium were significantly increased in monocytes from hypertensive patients compared to normotensive control subjects. The fMLPinduced monocyte migration was significantly reduced in the presence of inhibitors of tyrosine kinase and phosphoinositide 3-kinase. We conclude that increased monocyte migration in patients with essential hypertension is associated with increased TRPC3 channels. - Funding: This study was funded by 973 Program (grant no. 2011CB503902 and 2012CB517806); Natural Science Foundation of China (grant no. 30871058 and 30900619); Natural Science Foundation of Chongqing (CSTC, 2008BB5016) and Chinese Medical Association Clinical Research Grant (no. 07060700078). This study was supported in part by Else-Kro ner-Fresenius-Stiftung. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. . These authors contributed equally to this work. An increased transient receptor potential canonical type 3 (TRPC3) protein expression has been observed both in patients with essential hypertension and in animal models of hypertension [15]. In patients with hypertension an increased TRPC3 expression has been reported in several tissues including vascular smooth muscle cells and peripheral blood monocytes [1,2,5]. It is well established that monocytes play a crucial role in atherogenesis by recruitment to the vessel wall [6]. Monocyte activation, adhesion to the endothelium, and transmigration into the subendothelial space are key events in early pathogenesis of atherosclerosis [7]. Earlier studies from Doerffel et al. indicated that monocyte activation is increased in hypertension [8]. Monocytes from patients with essential hypertension show elevated secretion patterns of pro-inflammatory cytokines, an increased expression of adhesion molecules, and an increased adhesion to vascular endothelial cells [9]. Increased activation of monocytes in hypertension may be due to increased change of cytosolic calcium. TRPC3 channels are non-selective cation channels mediating transplasmamembrane calcium influx [10]. TRPC3 channels are likely candidates to produce increased activation of monocytes. An increased calcium influx through TRPC3 channels may cause increased migration of monocytes. However, the role of TRPC3 for regulating the migration of monocytes has not been investigated to date. In the present study we tested the hypothesis that increased TRPC3 channel expression causes increased migration of monocytes from patients with essential hypertension. Increased migration of monocytes from patients with essential hypertension First we evaluated the migration of monocytes using the chemoattractants fMLP and TNF-a. Figure 1A shows representative images of the fMLP-induced migration of monocytes from normotensive contr (...truncated)