Background and aims Resistin has been associated with atherosclerotic inflammation and cardiovascular complications. We and others have previously shown that PKC-epsilon (PKCε) is involved in resistin-induced smooth muscle cell (VSMC) dysfunction at a high pathological concentration. This study aimed to evaluate the role and potential pathways of resistin at a physiological concentration, in atherosclerosis-related inflammation. Methods Plasma from patients with atherosclerosis was analyzed for resistin concentration. Patients were divided into tertiles based on resistin levels and cytokines were compared between tertiles. Macrophages were then treated with resistin in the presence or absence of PKCε inhibitor and/or TLR4 blocking-antibody, and their inflammatory state was evaluated with ELISA, RT-PCR, immunocytochemistry, and Western blot. Results We observed significant associations between plasma resistin levels and TNF-α, IL-6, IL-12, MIP-1α, MIP-1β, and CD40L. Our in vitro analyses revealed that resistin activated PKCε via TLR4. This was followed by NF-kB activation and induction of a pro-inflammatory phenotype in macrophages, significantly upregulating CD40, downregulating CD206 and stimulating gene expression and secretion of the inflammatory cytokines, for which we found association in our plasma analysis. Resistin also induced persistent TRAM and CD40L upregulation up to 36 hours after resistin treatment. PKCε and TLR4 inhibitors suppressed gene expression to levels similar to control, especially when used in combination. Conclusions Resistin, at a physiological concentration, exacerbates the inflammatory response of macrophages. PKCε is a key upstream mediator in resistin-induced inflammation that may interact synergistically with TLR4 to promote NF-kB activation, while TRAM is an important signal. PKCε and TRAM may represent novel molecular targets for resistin-associated chronic atherosclerotic inflammation.
Backgrund and aims Resistin has been implicated in cardiovascular disease and poor interventional cardiovascular outcomes. Previous studies by our group demonstrated resistin promoted vascular smooth muscle cell (VSMC) migration through protein kinase C epsilon (PKCε) pathways, while few others showed that resistin induced reactive oxygen species (ROS) generation in various cell types. In this study, we aim to systemically examine the functional role of resistin at the cellular and tissue levels as well as the potential mechanistic relationship between resistin-induced PKCε activation and ROS production. Methods Plasma collected from patients undergoing carotid interventions was analyzed for resistin level and ROS. VSMCs were treated with resistin in the presence or absence of PKCε and NADPH oxidase (Nox)-specific inhibitors. Intracellular ROS production was analyzed using confocal microscopy and Nox activity with chemiluminescence. In vivo studies were performed in apolipoprotein E knock out (ApoE−/−) mice to determine therapeutic effects of PKCε-specific inhibitor, using the guide-wire injury model. Results We observed significant correlation between plasma resistin and circulating levels of oxidative stress in patients with severe atherosclerotic disease. We also demonstrated that resistin induced ROS production via PKCε-mediated Nox activation. Resistin-induced ROS production was time-dependent, and Nox4 was the primary isoform involved. Inhibition of Nox completely abolished resistin-exaggerated VSMC proliferation, migration and dedifferentiation, as well as pro-inflammatory cytokine release. Upstream modulation of PKCε significantly reduced resistin-mediated cytosolic ROS, Nox activity and VSMC dysfunction. Moreover, PKCε-specific inhibitor mitigated resistin-induced Nox activation and intimal hyperplasia in ApoE−/− mice. Conclusions Resistin-associated VSMC dysfunction and intimal hyperplasia are related to PKCε-dependent Nox activation and ROS generation. Targeting the PKCε-Nox pathway may represent a novel strategy in managing resistin-associated atherosclerotic complications.
Atherosclerotic cardiovascular disease has been acknowledged as a chronic inflammatory condition. Monocytes and macrophages lead the inflammatory pathology of atherosclerosis whereas changes in atheromatous plaque thickness and matrix composition are attributed to vascular smooth muscle cells. Because these cell types are key players in atherosclerosis progression, it is crucial to utilize a reliable system to investigate their interaction. In vitro co-culture systems are useful platforms to study specific molecular mechanisms between cells. This review aims to summarize the various co-culture models that have been developed to investigate vascular smooth muscle cell and monocyte/macrophage interactions, focusing on the monocyte/macrophage effects on vascular smooth muscle cell function.
Calcification of atherosclerotic plaques in elderly patients represents a potent risk marker of cardiovascular events. Plasma analyses of patients with or without calcified plaques reveal significant differences in chemokines, particularly eotaxin, which escalates with increased calcification. We therefore, hypothesize that eotaxin in circulation augments calcification of vascular smooth muscle cells (VSMCs) possibly via oxidative stress in the vasculature. We observe that eotaxin increases the rate of calcification significantly in VSMCs as evidenced by increased alkaline phosphatase activity, calcium deposition, and osteogenic marker expression. In addition, eotaxin promotes proliferation in VSMCs and triggers oxidative stress in a NADPH oxidase dependent manner. These primary novel observations support our proposition that in the vasculature eotaxin augments mineralization. Our findings suggest that eotaxin may represent a potential therapeutic target for prevention of cardiovascular complications in the elderly. J. Cell. Biochem. 118: 647-654, 2017. © 2016 Wiley Periodicals, Inc.
Objective To determine factors affecting cognition and identify predictors of long-term cognitive impairment following carotid revascularization procedures. Background Cognitive impairment is common in older patients with carotid occlusive diseases. Methods Patients undergoing carotid intervention for severe occlusive diseases were prospectively recruited. Patients received neurocognitive testing before, 1, and 6 months after carotid interventions. Plasma samples were also collected within 24 hours after carotid intervention and inflammatory cytokines were analyzed. Univariate and multivariate logistic regressions were performed to identify risk factors associated with significant cognitive deterioration (>10% decline). Results A total of 98 patients (48% symptomatic) were recruited, including 55 patients receiving carotid stenting and 43 receiving endarterectomy. Mean age was 69 (range 54–91 years). Patients had overall improvement in cognitive measures 1 month after revascularization. When compared with carotid stenting, endarterectomy patients demonstrated postoperative improvement in cognition at 1 and 6 months compared with baseline. Carotid stenting (odds ratio 6.49, P = 0.020) and age greater than 80 years (odds ratio 12.6, P = 0.023) were associated with a significant long-term cognitive impairment. Multiple inflammatory cytokines also showed significant changes after revascularization. On multivariate analysis, after controlling for procedure and age, IL-12p40 (P = 0.041) was associated with a higher risk of significant cognitive impairment at 1 month; SDF1-α (P = 0.004) and tumor necrosis factor alpha (P = 0.006) were independent predictors of cognitive impairment, whereas interleukin-6 (P = 0.019) demonstrated cognitive protective effects at 6 months after revascularization. Conclusions Carotid interventions affect cognitive function. Systemic biomarkers can be used to identify patients at risk of significant cognitive decline postprocedures that benefit from targeted cognitive training.
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