CLP is currently the most widely used animal model of sepsis and generally recognized as a reliable and clinically relevant animal model of the human septic condition. CLP surgery is straightforward: ligation distal to the ileocecal valve and needle puncture of ligated cecum cause leakage of fecal contents into the peritoneum, with subsequent polymicrobial bacteremia and sepsis. Multiple species of bacteria are found in the bloodstream, and progressive systemic inflammatory response syndrome followed by septic shock and multiorgan injury ensues.CLP-induced sepsis models show a cytokine profile similar to that in human sepsis [15]. Sepsis initiates a complex immune response that varies over time, with the concomitant occurrence of both pro-inflammatory and anti-inflammatory mechanisms. As a result, most patients with sepsis rapidly display signs of profound immunosuppression, which is associated with deleterious consequences [17, 18]. Previous studies have suggested that following the onset of sepsis, the production of both Th1 and Th2 cell-associated cytokines is decreased. Moreover, a shift from a Th1 to a Th2 response occurs, contributing to immunosuppression during sepsis [19, 20]. Our findings are consistent with those reported in previous studies, and indicate that anti-CD3/CD28 antibody-induced IL-2, IFN-γ, and IL-4 secretion from septic CD4+ T cells was markedly decreased on 12 h and 24 h. The proliferation activities of splenic CD4+ T cells in CLP 12h and CLP 24h groups were also lower than those in the sham group.
Th17 cells, characterized by the production of proinflammatory cytokines IL-17A, are a major contributor in multiple diseases [21, 22]. In contrast, Treg cells have been identified as dedicated suppressors of diverse immune responses and inflammations through the secretion of the anti-inflammatory cytokine IL-10 [23, 24]. These two cells play opposite roles in the inflammatory responses to jointly maintain immune homeostasis. The potential role of the Th17/Treg balance in sepsis has not been fully elucidated. In this study, we observed that septic mice exhibited a significant increase in the frequency of Th17 cells and CD4+CD25+Treg, compared to the sham group. Furthermore, we confirmed that the Treg/Th17 ratio in septic mice were higher as compared to the sham group. These results are consistent with previous studies. A study also showed an imbalance in cell mediated immune response and disturbance in Th1/Th2/Th17 and Treg population of T helper cells in patients with post traumatic sepsis [25]. Therefore, the skewing of the Treg/Th17 ratio along with Th2/Th1 may contribute to the pathogenesis of sepsis.
Calcium (Ca2+) is a ubiquitous intracellular signaling entity responsible for controlling numerous cellular processes. Calcium SOCs activated by the depletion of Ca2+ from the ER are a primary Ca2+ entry pathway in nonexcitable cells and are essential for T cell activation and adaptive immunity [26–28]. SOCE is required not only for Treg development but also for their suppressive function [29]. Our study confirmed that Ca2+-calcineurin-NFAT signaling pathways as well as SOCE channels were inhibited in septic mice. Research also shows that CaN-NFAT pathwayof T cells were inhibited in Cryptococcus neoformans infected rats [30]. However, it has been reported that inhibition of Ca2+ channels, rather than activation of SOCE process, is beneficial for the survival of septic patients [31]. In sepsis, T-cell immune response changes with time, and the level of SOCE may change with time. Therefore, more time points are needed to clarify its changing law in the CLP model. In this study, we found that the level of SOCE of CD4+ T cells in sepsis was slightly decreased, while the proliferation of CD4+ T cells was significantly decreased.The reason is that the proliferation ability is not only affected by SOCE, but also by other factors, such as apoptosis and immunosuppression.
Calcium SOCs compose of poreforming Ca2+ channel subunits Orai1, Orai2, and Orai3 as well as ER Ca2+ sensors STIM1 and STIM2 [32, 33]. As major components of SOCs, Orai1 appear to be both necessary and sufficient to reconstitute SOCE [13, 14]. In septic mice, the protein expression of Orai1 was decreased compared with that in the sham group. Interestingly, Upregulation of Orai1 not only can improve immune function of T cell in sepsis and reduce the mortality and organ damage in septic mice but also can partially recovery of SOCE in sepsis. These results proved that SOCE channels were suppressed during sepsis, and this may be involved in sepsis-induced immunosuppression of T lymphocytes and acute organ dysfunction. However, a study has shown that Orai1-deficient mice had normal Treg development; this is presumably because of residual SOCE in naive CD4+ T cells, which is likely to be mediated by Orai2 and/or Orai3 [34]. We will further validate this mechanism by knocking out Orai1 in septic mice in future studies.
Taken together, the skewing of the Treg/Th17 along with Th2/Th1 may contribute to the pathogenesis of sepsis. SOCE channels were suppressed during sepsis, Orai1 can regulate SOCE. Lower SOCE levels may be associated with the sepsis-induced immunosuppression of T lymphocytes and acute organ dysfunction. Upregulation of Orai1 can partially block this effect. This will be a potential target for treatment of sepsis.