title: High glucose activates Raw264.7 macrophages through RhoA kinase-mediated signaling pathway.
abstract: Hyperglycemia has been shown to accelerate atherogenesis, an inflammation process resulting from macrophage activation. Although high glucose (HG) was previously demonstrated to accentuate ROCK activity in macrophages and enhance their activation in vitro, the role of ROCK signaling in HG-mediated macrophage activation remains unclear. This study aimed to elucidate potential signal transduction pathways of HG-mediated ROCK upregulation and macrophage activation, including c-Jun or NF-κB pathways. A macrophage cell line, RAW264.7, was used to investigate the atherogenic effects of HG on RhoA/ROCK activity and macrophage functions. Exposure to HG significantly induced RhoA membrane translocation, RhoA-kinase activity, and phosphorylation of myosin-binding subunit, a RhoA-kinase substrate. Macrophage behaviors, including cell proliferation, adhesion, migration, and TNF-α de novo synthesis, were also increased by HG exposure. However, pharmacological ROCK inhibition by hydroxyfasudil attenuated the HG-enhanced adhesion and TNF-α production. Nuclear translocation of c-Jun and transcription factor NF-κB was simultaneously noted after HG stimulation. Pharmacological ROCK inhibition by hydroxyfasudil and siRNA-mediated ROCK1 or ROCK2 gene silencing confirmed the ROCK-dependent JNK and ERK phosphorylation, but not NF-κB activation in macrophages. In addition, both interventions effectively ameliorated the HG-mediated macrophage activation under the conditions mimicking diabetes. These findings suggest that hyperglycemia activates macrophages mainly through ROCK/JNK and ROCK/ERK pathways, which results in a more pro-inflammatory phenotype and eventually contributes to atherogenesis. In conclusion, ROCK inhibition might become a novel therapeutic strategy in atherosclerosis treatment and prevention in diabetic patients.
authors.affiliation: Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang-Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung 83301, Taiwan.
authors.forename: Cheng-I
authors.initials: CI
authors.lastname: Cheng
authors.affiliation: Department of Medical Research, E-DA Hospital, Kaohsiung 82445, Taiwan.
authors.forename: Po-Han
authors.initials: PH
authors.lastname: Chen
authors.affiliation: Department of Medical Research, E-DA Hospital, Kaohsiung 82445, Taiwan.
authors.forename: Yu-Chun
authors.initials: YC
authors.lastname: Lin
authors.affiliation: Department of Medical Research, E-DA Hospital, Kaohsiung 82445, Taiwan. Electronic address: danyhkao@gmail.com.
authors.forename: Ying-Hsien
authors.initials: YH
authors.lastname: Kao
mesh_terms: D000818:Animals; D002460:Cell Line; D005947:Glucose / Q000494:pharmacology*; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D015533:Transcriptional Activation / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
chemical_list: D016328:NF-kappa B; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; C515854:Rock1 protein, mouse; C515855:Rock2 protein, mouse; D054460:rho-Associated Kinases; D020928:Mitogen-Activated Protein Kinases; D020742:rhoA GTP-Binding Protein; D005947:Glucose
keywords: Atherosclerosis; Hyperglycemia; Macrophages; Rho-associated coiled coil forming kinase; Signal transduction