Objective: While type 1 programmed cell death (apoptosis) of T cells leads to immunosuppression in sepsis, a crosstalk between apoptosis and autophagy (type 2 programmed cell death) has not been shown. the gene expression of Bcl-2-like 11 and programmed cell death 1. Furthermore, mitochondrial accumulation in T cells occurred via a blockade of autophagy during sepsis. In BX-912 addition, interleukin-10 production in CD4+ T cells from the cecal ligation and punctureCoperated knockout mice was markedly increased. Consequently, deficiency of autophagy in T cells significantly decreased the survival rate in the murine sepsis model. Conclusions: We demonstrated that blocking autophagy accelerated apoptosis and increased mortality in concordance with the insufficient autophagy process in CD4+ T cells in the murine sepsis model, suggesting that T cell autophagy plays a protective role against apoptosis and immunosuppression in sepsis. test for continuous data and used two-way analysis of variance among different categoric independent variables. Statistical analyzes were conducted using the GraphPad Prism 6 (GraphPad Software, San Diego, CA). RESULTS Although Lymphocyte Autophagosomes are Increased by Septic Stimulation, the Process of Autophagy is Insufficient in a Murine Sepsis Model in CD4+ T Cells To evaluate the autophagy kinetics of lymphocytes in sepsis, we performed in vitro assay to replicate the condition at first. Lymphocytes from GFP-LC3 mice were stimulated with anti-CD3/CD28 or lipopolysaccharide (LPS) for 48 hours. Mean fluorescence intensity (MFI) of GFP-LC3, which represents autophagosomes, was measured by flow cytometry. As shown in Figure ?Figure11and = 0.016) (Fig. 4C). As shown in Figure 4C, the mortality rate for GFAP controls relative to the conditional knockout mice was a BX-912 more robust improvement in survival at 72 hours, that is, just after occurrence of the T cell apoptosis. DISCUSSION In this study, we demonstrated several findings regarding T cell autophagy in sepsis. First, an autophagy process in CD4+ T cells is insufficient during sepsis despite an increase of autophagosomes. Second, blockade of T cell autophagy accelerates apoptosis. Third, mitochondrial accumulation in T cells occurs via a blockade of autophagy during sepsis. Fourth, IL-10 production is increased in CD4+ T cells by blockade of autophagy, which may drive a further immunosuppressive state. Finally, a deficiency of autophagy in T cells decreases the survival rate in the murine sepsis model. These findings suggest that T cell autophagy plays a protective role against apoptosis and immunosuppression in the BX-912 murine sepsis model. As interest in the process of autophagy has developed, Crouser et al (17) have shed light on the interaction between autophagy and sepsis. In their report, they demonstrated that mitochondrial depletion was related to the removal of the damaged organelles by autophagy in a murine sepsis model. Furthermore, Watanabe et al (18) observed an increase of liver autophagosomes BX-912 in sepsis patients for the first time. Although the role of autophagy in sepsis had not been clearly understood, Takahashi et al (13) demonstrated a protective role of autophagy in liver using CLP-operated mice. However, the kinetics of autophagy in T cells during sepsis have still not been elucidated. BX-912 We used GFP-LC3 transgenic mice in which the autophagic activity can be monitored objectively over time using flow cytometry (19). Furthermore, we separated CD4+ T cells and confirmed the images of increased autophagosomes and autolysosomes in CLP relative to sham with our TEM data (Supplemental Fig. 1, Supplemental Digital Content 2, http://links.lww.com/CCM/C115; legend, Supplemental Digital Content 1, http://links.lww.com/CCM/C114). Also, we found remarkable mitochondrial damages in CD4+ T cells of CLP-operated mice. To our knowledge, ours is the only study in which T cell autophagy in sepsis was monitored in such detail to date. Our results demonstrated that mitochondria accumulated in the T cells of the CD4-Cre/Atg5f/f mice. It is consistent with the previous report that T cells lacking the autophagy-related genes Atg5 or Atg7 have inferior survival rates to those expressing the genes and contain expanded mitochondria (20, 21). As damaged organelles accumulate, cells are unable to maintain their physiologic functions. Although we did not demonstrate a direct relationship between apoptosis induction and mitochondrial accumulation, it has previously been shown that deletion of autophagy in T cells induces abnormal reactive oxygen species (ROS) production and apoptosis,.