We hypothesized that SIRS/endotoxemia-associated hyporesponsiveness to vasoconstrictors is mediated by smaller increases in intracellular Ca2+ levels due to reduced signaling via the STIM/Orai. Male Wistar rats were injected either with saline or bacterial LPS (i.p.; 10 mg/kg), and experiments were performed 24 h later. LPS-injected rats exhibited decreased systolic blood pressure, increased heart rate, neutrophils’ migration into the peritoneal cavity, and elevated alanine aminotransferase levels. Additionally, second-order mesenteric arteries from endotoxemic rats displayed hyporeactivity to contractile agents such as phenylephrine and potassium chloride; decreased contractile responses to Ca2+; reduced contraction during Ca2+ loading; and smaller intracellular Ca2+ stores. Decreased Orai1, but not STIM1, expression was found in resistance mesenteric arteries from LPS-treated rats. Additionally, cultured vascular smooth muscle cell (VSMC) treated with LPS resulted in increased TLR-4 expression, but Myd-88 and STIM-1 expression were not changed. Our data suggest that in endotoxemia, Ca2+ homeostasis is disrupted in VSMC, with decreased Ca2+ influx, smaller concentrations of Ca2+ in the sarcoplasmic reticulum, and decreased activation of Orai1. Abnormal Ca2+ handling contributes to LPS-associated vascular hyporeactivity.
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This work was supported in part by Fundação de Amparo à Pesquisa do Estado de Mato Grosso (FAPEMAT) [grant number 151371/2014 (to F.R.G.], Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) [grant number 23038009165/2013-48 (to V.V.L.], Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) [grant number 2010/52214-6 (to R.C.T)], Conselho Nacional de Desenvolvimento Científico e Tecnológico [(CNPq) 471675/2013-0 and 305823/2015-9 (to F.R.G), 445777/2014-1 (V.V.L.)], and National Institutes of Health (NIH) [HL71138 and DK83685 (R.C.W)]. We would also like to thank all the technical staff, who have worked in our laboratories and contributed to the studies described here.
Fernanda Giachini and Victor Lima performed the vascular reactivity studies, in addition to being responsible for writing the paper. Vanessa Dela Justina conducted the cell culture. Arthur Nonato and Vanessa Dela Justina conducted Western blot analysis. Vania Olivon and Camila Zanotto conducted biochemistry analisys. Rita Tostes and Clinton Webb provided the animals used in the study and, along with Vania Olivon, continuously provided ideas and expertise for the project and revisions for the paper. Fernanda Giachini designed the hypothesis and supervised the entire study. All of the authors had full access to the data and take responsibility for its integrity and the accuracy of the analysis. All authors have read and agree to the paper as written.
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