The role of heat shock protein 70 in mediating age-dependent mortality in sepsis

Sepsis is primarily a disease of the aged, with increased incidence and mortality occurring in aged hosts. Heat shock protein (HSP) 70 plays an important role in both healthy aging and the stress response to injury. The purpose of this study was to determine the role of HSP70 in mediating mortality and the host inflammatory response in aged septic hosts. Sepsis was induced in both young (6- to 12-wk-old) and aged (16- to 17-mo-old) HSP70(-/-) and wild-type (WT) mice to determine whether HSP70 modulated outcome in an age-dependent fashion. Young HSP70(-/-) and WT mice subjected to cecal ligation and puncture, Pseudomonas aeruginosa pneumonia, or Streptococcus pneumoniae pneumonia had no differences in mortality, suggesting HSP70 does not mediate survival in young septic hosts. In contrast, mortality was higher in aged HSP70(-/-) mice than aged WT mice subjected to cecal ligation and puncture (p = 0.01), suggesting HSP70 mediates mortality in sepsis in an age-dependent fashion. Compared with WT mice, aged septic HSP70(-/-) mice had increased gut epithelial apoptosis and pulmonary inflammation. In addition, HSP70(-/-) mice had increased systemic levels of TNF-α, IL-6, IL-10, and IL-1β compared with WT mice. These data demonstrate that HSP70 is a key determinant of mortality in aged, but not young hosts in sepsis. HSP70 may play a protective role in an age-dependent response to sepsis by preventing excessive gut apoptosis and both pulmonary and systemic inflammation.     

Overexpression of Bcl-2 in transgenic mice decreases apoptosis and improves survival in sepsis

In sepsis there is extensive apoptosis of lymphocytes, which may be beneficial by down-regulating the accompanying inflammation. Alternatively, apoptosis may be detrimental by impairing host defense. We studied whether Bcl-2, a potent antiapoptotic protein, could prevent lymphocyte apoptosis in a clinically relevant model of sepsis. Transgenic mice in which Bcl-2 was overexpressed in T cells had complete protection against sepsis-induced T lymphocyte apoptosis in thymus and spleen. Surprisingly, there was also a decrease in splenic B cell apoptosis in septic Bcl-2 overexpressors compared with septic HeJ and HeOuJ mice. There were marked increases in TNF-alpha, IL-1beta, and IL-10 in thymic tissue in sepsis in the three species of mice, and the increase in TNF-alpha and IL-10 in HeOuJ mice was greater than that in Bcl-2 mice. Mitotracker, a mitochondrial membrane potential indicator, demonstrated a sepsis-induced loss of membrane potential in T cells in HeJ and HeOuJ mice but not in Bcl-2 mice. Importantly, Bcl-2 overexpressors also had improved survival in sepsis. To investigate the potential impact of loss of lymphocytes on survival in sepsis, Rag-1-/- mice, which are totally deficient in mature T and B cells, were also studied. Rag-1-/- mice had decreased survival compared with immunologically normal mice with sepsis. We conclude that overexpression of Bcl-2 provides protection against cell death in sepsis. Lymphocyte death may be detrimental in sepsis by compromising host defense.     

Murine Lung Cancer Increases CD4+ T Cell Apoptosis and Decreases Gut Proliferative Capacity in Sepsis

Background

Mortality is significantly higher in septic patients with cancer than in septic patients without a history of cancer. We have previously described a model of pancreatic cancer followed by sepsis from Pseudomonas aeruginosa pneumonia in which cancer septic mice have higher mortality than previously healthy septic mice, associated with increased gut epithelial apoptosis and decreased T cell apoptosis. The purpose of this study was to determine whether this represents a common host response by creating a new model in which both the type of cancer and the model of sepsis are altered.

Methods

C57Bl/6 mice received an injection of 250,000 cells of the lung cancer line LLC-1 into their right thigh and were followed three weeks for development of palpable tumors. Mice with cancer and mice without cancer were then subjected to cecal ligation and puncture and sacrificed 24 hours after the onset of sepsis or followed 7 days for survival.

Results

Cancer septic mice had a higher mortality than previously healthy septic mice (60% vs. 18%, p = 0.003). Cancer septic mice had decreased number and frequency of splenic CD4+ lymphocytes secondary to increased apoptosis without changes in splenic CD8+ numbers. Intestinal proliferation was also decreased in cancer septic mice. Cancer septic mice had a higher bacterial burden in the peritoneal cavity, but this was not associated with alterations in local cytokine, neutrophil or dendritic cell responses. Cancer septic mice had biochemical evidence of worsened renal function, but there was no histologic evidence of renal injury.

Conclusions

Animals with cancer have a significantly higher mortality than previously healthy animals following sepsis. The potential mechanisms associated with this elevated mortality differ significantly based upon the model of cancer and sepsis utilized. While lymphocyte apoptosis and intestinal integrity are both altered by the combination of cancer and sepsis, the patterns of these alterations vary greatly depending on the models used.     

IL-10 mediation of activation-induced TH1 cell apoptosis and lymphoid dysfunction in polymicrobial sepsis

Recent studies suggest that increased activation-induced lymphocyte apoptosis (AICD) is detected in mouse splenocytes during polymicrobial sepsis which may contribute to lymphocyte immune dysfunction [i.e., decreased interleukin (IL-)2 and interferon-gamma (IFN-gamma) production] leading to the associated morbidity seen in those animals. Thus, we wanted to examine the hypothesis that immune suppressive agents, such as IL-4, IL-10 or prostaglandin E2(PGE2), known to be elevated in septic animals, also contribute to this increase in AICD. Here we demonstrate that the inclusion of monoclonal antibody (mAb) to IL-10, but not anti-IL-4 or ibuprofen (IBU), blunted this sepsis induced increase in splenocyte AICD. Additionally, septic mice deficient in the IL-10 gene product (-/-) showed neither an increase in AICD nor a loss of IL-2/IFN-gamma release capacity. Interestingly, mAb to IL-10 did not altered the extent of AICD in a Th2-cell line, but exogenous IL-10 did potentiate Th1-like cell line AICD. This was consistent with the finding that the increased AICD seen in septic mouse splenocytes was restricted largely to the CD4+ cells producing IL-2 (Th1-cells) and that mAb to IL-10 treatment suppressed this change. Furthermore, IL-10 appears to mediate its AICD effect by upregulation of the Fas receptor and Fas receptor signaling protein components, but not by altered expression of Bcl/Bax/Bad family members, in septic mouse splenocytes. To the extent that these processes contribute in a pathological fashion to the animal's capacity to survive sepsis we have previously observed that in vivo post-treatment of mice with mAb IL-10 markedly attenuated septic mortality. Collectively, these data indicate that in the septic mouse the Th2 cytokine IL-10 not only serves to actively induce Th1 lymphocyte immune dysfunction but also plays a role in their apoptotic depletion. These processes in turn appear to contribute to the animal's inability to ward off lethal septic challenge.     

Intestine-Specific Deletion of Microsomal Triglyceride Transfer Protein Increases Mortality in Aged Mice

Background

Mice with conditional, intestine-specific deletion of microsomal triglyceride transfer protein (Mttp-IKO) exhibit a complete block in chylomicron assembly together with lipid malabsorption. Young (8–10 week) Mttp-IKO mice have improved survival when subjected to a murine model of Pseudomonas aeruginosa-induced sepsis. However, 80% of deaths in sepsis occur in patients over age 65. The purpose of this study was to determine whether age impacts outcome in Mttp-IKO mice subjected to sepsis.

Methods

Aged (20–24 months) Mttp-IKO mice and WT mice underwent intratracheal injection with P. aeruginosa. Mice were either sacrificed 24 hours post-operatively for mechanistic studies or followed seven days for survival.

Results

In contrast to young septic Mttp-IKO mice, aged septic Mttp-IKO mice had a significantly higher mortality than aged septic WT mice (80% vs. 39%, p = 0.005). Aged septic Mttp-IKO mice exhibited increased gut epithelial apoptosis, increased jejunal Bax/Bcl-2 and Bax/Bcl-X_L ratios yet simultaneously demonstrated increased crypt proliferation and villus length. Aged septic Mttp-IKO mice also manifested increased pulmonary myeloperoxidase levels, suggesting increased neutrophil infiltration, as well as decreased systemic TNFα compared to aged septic WT mice.

Conclusions

Blocking intestinal chylomicron secretion alters mortality following sepsis in an age-dependent manner. Increases in gut apoptosis and pulmonary neutrophil infiltration, and decreased systemic TNFα represent potential mechanisms for why intestine-specific Mttp deletion is beneficial in young septic mice but harmful in aged mice as each of these parameters are altered differently in young and aged septic WT and Mttp-IKO mice.     

Scavenger Receptor Class A Plays a Central Role in Mediating Mortality and the Development of the Pro-Inflammatory Phenotype in Polymicrobial Sepsis

Sepsis is a frequent complication in critical illness. The mechanisms that are involved in initiation and propagation of the disease are not well understood. Scavenger receptor A (SRA) is a membrane receptor that binds multiple polyanions such as oxidized LDL and endotoxin. Recent studies suggest that SRA acts as a pattern recognition receptor in the innate immune response. The goal of the present study was to determine the role of SRA in polymicrobial sepsis. SRA deficient (SRA^−/−) and C57BL/6JB/6J (WT) male mice were subjected to cecal ligation and puncture (CLP) to induce polymicrobial sepsis. NFκB activity, myeloperoxidase activity, and co-association of SRA with toll like receptor (TLR) 4 and TLR2 was analyzed in the lungs. Spleens were analyzed for apoptosis. Serum cytokines and chemokines were assayed. Blood and peritoneal fluid were cultured for aerobic and anaerobic bacterial burdens. Long term survival was significantly increased in SRA^−/− septic mice (53.6% vs. 3.6%, p<0.05) when compared to WT mice. NFκB activity was 45.5% lower in the lungs of SRA^−/− septic mice versus WT septic mice (p<0.05). Serum levels of interleukin (IL)-5, IL-6, IL-10 and monocyte chemoattractant protein −1 were significantly lower in septic SRA^−/− mice when compared to septic WT mice (p<0.05). We found that SRA immuno-precipitated with TLR4, but not TLR2, in the lungs of WT septic mice. We also found that septic SRA^−/− mice had lower bacterial burdens than WT septic mice. SRA deficiency had no effect on pulmonary neutrophil infiltration or splenocyte apoptosis during sepsis. We conclude that SRA plays a pivotal, and previously unknown, role in mediating the pathophysiology of sepsis/septic shock in a murine model of polymicrobial sepsis. Mechanistically, SRA interacts with TLR4 to enhance the development of the pro-inflammatory phenotype and mediate the morbidity and mortality of sepsis/septic shock.     

Akt decreases lymphocyte apoptosis and improves survival in sepsis

Sepsis induces extensive death of lymphocytes that may contribute to the immunosuppression and mortality of the disorder. The serine/threonine kinase Akt is a key regulator of cell proliferation and death. The purpose of this study was to determine whether overexpression of Akt would prevent lymphocyte apoptosis and improve survival in sepsis. In addition, given the important role of Akt in cell signaling, T cell Th1 and Th2 cytokine production was determined. Mice that overexpress a constitutively active Akt in lymphocytes were made septic, and survival was recorded. Lymphocyte apoptosis and cytokine production were determined at 24 h after surgery. Mice with overexpression of Akt had a marked improvement in survival compared with wild-type littermates, i.e., 94 and 47% survival, respectively, p < 0.01. In wild-type littermates, sepsis caused a marked decrease in IFN-gamma production, while increasing IL-4 production >2-fold. In contrast, T cells from Akt transgenic mice had an elevated production of IFN-gamma at baseline that was maintained during sepsis, while IL-4 had little change. Akt overexpression also decreased sepsis-induced lymphocyte apoptosis via a non-Bcl-2 mechanism. In conclusion, Akt overexpression in lymphocytes prevents sepsis-induced apoptosis, causes a Th1 cytokine propensity, and improves survival. Findings from this study strengthen the concept that a major defect in sepsis is impairment of the adaptive immune system, and suggest that strategies to prevent lymphocyte apoptosis represent a potential important new therapy.     

A3 adenosine receptor activation decreases mortality and renal and hepatic injury in murine septic peritonitis

The role of A3 adenosine receptors (ARs) in sepsis and inflammation is controversial. In this study, we determined the effects of A3AR modulation on mortality and hepatic and renal dysfunction in a murine model of sepsis. To induce sepsis, congenic A3AR knockout mice (A3AR KO) and wild-type control (A3AR WT) mice were subjected to cecal ligation and double puncture (CLP). A3AR KO mice had significantly worse 7-day survival compared with A3AR WT mice. A3AR KO mice also demonstrated significantly higher elevations in plasma creatinine, alanine aminotransferase, aspartate aminotransferase, keratinocyte-derived chemokine, and TNF-alpha 24 h after induction of sepsis compared with A3AR WT mice. Renal cortices from septic A3AR KO mice exhibited increased mRNA encoding proinflammatory cytokines and enhanced nuclear translocation of NF-kB compared with samples from A3AR WT mice. A3AR WT mice treated with N6-(3-iodobenzyl)ADO-5'N-methyluronamide (IB-MECA; a selective A3AR agonist) or 3-ethyl-5-benzyl-2-methyl-4-phenylethynyl-6-phenyl-1,4-(+/-)-dihydropyridine-3,5-dicarboxylate (MRS-1191; a selective A3AR antagonist) had improved or worsened 7-day survival after induction of sepsis, respectively. Moreover, A3AR WT mice treated with IB-MECA or MRS-1191 showed acutely improved or worsened, respectively, renal and hepatic function following CLP. IB-MECA significantly reduced mortality in mice lacking the A1AR or A2aAR but not the A3AR, demonstrating specificity of IB-MECA in activating A3ARs and mediating protection against sepsis-induced mortality. We conclude that endogenous or exogenous A3AR activation confers significant protection from murine septic peritonitis primarily by attenuating the hyperacute inflammatory response in sepsis.     

Accelerated lymphocyte death in sepsis occurs by both the death receptor and mitochondrial pathways

Patients with sepsis are immune compromised, as evidenced by their failure to clear their primary infection and their propensity to develop secondary infections with pathogens that are often not particularly virulent in normal healthy individuals. A potential mechanism for immunosuppression in sepsis is lymphocyte apoptosis, which may occur by either a death receptor or a mitochondrial-mediated pathway. A prospective study of blood samples from 71 patients with sepsis, 55 nonseptic patients, and 6 healthy volunteers was undertaken to quantitate lymphocyte apoptosis and determine cell death pathways and mechanisms of apoptosis. Apoptosis was evaluated by flow cytometry and Western blotting. Lymphocyte apoptosis was increased in CD4 and CD8 T cells, B cells (CD20), and NK cells (CD56) in septic vs nonseptic patients. Samples taken sequentially from 10 patients with sepsis showed that the degree of CD3 T cell apoptosis correlated with the activity of his/her sepsis. In septic patients, apoptotic lymphocytes were positive for active caspases 8 and 9, consistent with death occurring by both mitochondrial-mediated and receptor-mediated pathways. In support of the concept that both death pathways were operative, lymphocyte apoptosis occurred in cells with markedly decreased Bcl-2 (an inhibitor of mitochondrial-mediated apoptosis) as well as cells with normal concentrations of Bcl-2. In conclusion, apoptosis occurs in a broad range of lymphocyte subsets in patients with sepsis and correlates with the activity of the disease. Lymphocyte loss occurs by both death receptor and mitochondrial-mediated apoptosis, suggesting that there may be multiple triggers for lymphocyte apoptosis.     

TAT-BH4 and TAT-Bcl-xL peptides protect against sepsis-induced lymphocyte apoptosis in vivo

Apoptosis is a key pathogenic mechanism in sepsis that induces extensive death of lymphocytes and dendritic cells, thereby contributing to the immunosuppression that characterizes the septic disorder. Numerous animal studies indicate that prevention of apoptosis in sepsis improves survival and may represent a potential therapy for this highly lethal disorder. Recently, novel cell-penetrating peptide constructs such as HIV-1 TAT basic domain and related peptides have been developed to deliver bioactive cargoes and peptides into cells. In the present study, we investigated the effects of sepsis-induced apoptosis in Bcl-x(L) transgenic mice and in wild-type mice treated with an antiapoptotic TAT-Bcl-x(L) fusion protein and TAT-BH4 peptide. Lymphocytes from Bcl-x(L) transgenic mice were resistant to sepsis-induced apoptosis, and these mice had a approximately 3-fold improvement in survival. TAT-Bcl-x(L) and TAT-BH4 prevented Escherichia coli-induced human lymphocyte apoptosis ex vivo and markedly decreased lymphocyte apoptosis in an in vivo mouse model of sepsis. In conclusion, TAT-conjugated antiapoptotic Bcl-2-like peptides may offer a novel therapy to prevent apoptosis in sepsis and improve survival.     

Glucan phosphate attenuates cardiac dysfunction and inhibits cardiac MIF expression and apoptosis in septic mice

Myocardial dysfunction is a major consequence of septic shock and contributes to the high mortality of sepsis. We have previously reported that glucan phosphate (GP) significantly increased survival in a murine model of cecal ligation and puncture (CLP)-induced sepsis. In the present study, we examined the effect of GP on cardiac dysfunction in CLP-induced septic mice. GP was administered to ICR/HSD mice 1 h before induction of CLP. Sham surgically operated mice served as control. Cardiac function was significantly decreased 6 h after CLP-induced sepsis compared with sham control. In contrast, GP administration prevented CLP-induced cardiac dysfunction. Macrophage migration inhibitory factor (MIF) has been implicated as a major factor in cardiomyocyte apoptosis and cardiac dysfunction during septic shock. CLP increased myocardial MIF expression by 88.3% (P < 0.05) and cardiomyocyte apoptosis by 7.8-fold (P < 0.05) compared with sham control. GP administration, however, prevented CLP-increased MIF expression and decreased cardiomyocyte apoptosis by 51.2% (P < 0.05) compared with untreated CLP mice. GP also prevented sepsis-caused decreases in phospho-Akt, phospho-GSK-3beta, and Bcl-2 levels in the myocardium of septic mice. These data suggest that GP treatment attenuates cardiovascular dysfunction in fulminating sepsis. GP administration also activates the phosphoinositide 3-kinase/Akt pathway, decreases myocardial MIF expression, and reduces cardiomyocyte apoptosis.     

Lymphocyte apoptosis in murine Pneumocystis pneumonia

Background

Apoptosis of lymphocytes is important in the termination of an immune response to infection but has also been shown to have detrimental effects in animal models of systemic infection and sepsis. We sought to characterize lymphocyte apoptosis in an animal model of pneumonia due to Pneumocystis murina, an infection localized to the lungs.

Methods

Control mice and mice depleted of CD4+ lymphocytes were inoculated with Pneumocystis. Apoptosis of lung and spleen lymphocytes was assayed by flow cytometry and PCR assay of apoptotic proteins.

Results

In control mice, apoptosis of lung lymphocytes was maximal just after the infection was cleared from lung tissue and then declined. However, in CD4-depleted mice, apoptosis was also upregulated in recruited lymphocytes in spite of progressive infection. In splenic lymphocytes, apoptosis was observed early at 1 week after inoculation and then declined. Apoptosis of lung lymphocytes in control mice was associated with a decrease in mRNA for Bcl-2 and an increase in mRNA for Bim. In CD4-depleted mice, lavaged CD8+ cells did change intracellular Bcl-2 but showed increased mRNA for Bim.

Conclusion

Apoptosis of both pulmonary and extrapulmonary lymphocytes is part of the normal host response to Pneumocystis but is also triggered in CD4-deficient animals with progressive infection. In normal mice apoptosis of pulmonary lymphocytes may serve to terminate the immune response in lung tissue. Apoptosis of lung lymphocytes takes place via both the intrinsic and extrinsic apoptotic pathways and is associated with changes in both pro- and anti-apoptotic proteins.     

The Bcl-2 Homology Domain 3 (BH3)-only Proteins Bim and Bid Are Functionally Active and Restrained by Anti-apoptotic Bcl-2 Family Proteins in Healthy Liver

An intrinsic pathway of apoptosis is regulated by the B-cell lymphoma-2 (Bcl-2) family proteins. We previously reported that a fine rheostatic balance between the anti- and pro-apoptotic multidomain Bcl-2 family proteins controls hepatocyte apoptosis in the healthy liver. The Bcl-2 homology domain 3 (BH3)-only proteins set this rheostatic balance toward apoptosis upon activation in the diseased liver. However, their involvement in healthy Bcl-2 rheostasis remains unknown. In the present study, we focused on two BH3-only proteins, Bim and Bid, and we clarified the Bcl-2 network that governs hepatocyte life and death in the healthy liver. We generated hepatocyte-specific Bcl-xL- or Mcl-1-knock-out mice, with or without disrupting Bim and/or Bid, and we examined hepatocyte apoptosis under physiological conditions. We also examined the effect of both Bid and Bim disruption on the hepatocyte apoptosis caused by the inhibition of Bcl-xL and Mcl-1. Spontaneous hepatocyte apoptosis in Bcl-xL- or Mcl-1-knock-out mice was significantly ameliorated by Bim deletion. The disruption of both Bim and Bid completely prevented hepatocyte apoptosis in Bcl-xL-knock-out mice and weakened massive hepatocyte apoptosis via the additional in vivo knockdown of mcl-1 in these mice. Finally, the hepatocyte apoptosis caused by ABT-737, which is a Bcl-xL/Bcl-2/Bcl-w inhibitor, was completely prevented in Bim/Bid double knock-out mice. The BH3-only proteins Bim and Bid are functionally active but are restrained by the anti-apoptotic Bcl-2 family proteins under physiological conditions. Hepatocyte integrity is maintained by the dynamic and well orchestrated Bcl-2 network in the healthy liver.     

Decoy Receptor 3 Improves Survival in Experimental Sepsis by Suppressing the Inflammatory Response and Lymphocyte Apoptosis

Purpose

Unbalanced inflammatory response and lymphocyte apoptosis is associated with high mortality in septic patients. Decoy receptor 3 (DcR3), a member of the tumor necrosis factor receptor superfamily, is an anti-inflammatory and anti-apoptotic factor. Recently, DcR3 expression was found to be increased in septic patients. This study evaluated the therapeutic effect and mechanisms of DcR3 on cecal ligation and puncture (CLP)-induced sepsis in mice.

Methods

C57BL/6 mice were subjected to CLP-induced polymicrobial sepsis. DcR3 Fc was intravenously injected 30 min before and 6 h after CLP. Bacterial clearance, cytokine production, histology, lymphocyte apoptosis and survival were evaluated. Furthermore, we investigated the systemic effects of DcR3 in in vitro lymphocyte apoptosis regulation.

Results

Our results demonstrated that DcR3 protein treatments significantly improved survival in septic mice (p <0.05). Treatment with DcR3 protein significantly reduced the inflammatory response and decreased lymphocyte apoptosis in the thymus and spleen. Histopathological findings of the lung and liver showed milder impairment after DcR3 administration. In vitro experiments showed that DcR3 Fc inhibited Fas-FasL mediated lymphocyte apoptosis.

Conclusions

Treatment with the DcR3 protein protects mice from sepsis by suppressing the inflammatory response and lymphocyte apoptosis. DcR3 protein may be useful in treatment of sepsis.     

Deficiency of gammadelta T lymphocytes contributes to mortality and immunosuppression in sepsis

Studies have indicated that gammadelta T lymphocytes play an important role in the regulation of immune function and the clearance of intracellular pathogens. We have recently reported that intraepithelial lymphocytes (IEL), which are rich in gammadelta T cells, within the small intestine illustrated a significant increase in apoptosis and immune dysfunction in mice subjected to sepsis. However, the contribution of gammadelta T cells to the host response to polymicrobial sepsis remains unclear. In this study, we initially observed that after sepsis induced by cecal ligation and puncture (CLP), there was an increase in small intestinal IEL CD8+gammadelta+ T cells in control gammadelta+/+ mice. Importantly, we subsequently found an increased early mortality in mice lacking gammadelta T cells (gammadelta-/- mice) after sepsis. This was associated with decreases in plasma TNF-alpha, IL-6, and IL-12 levels in gammadelta-/- mice compared with gammadelta+/+ mice after sepsis. In addition, even though in vitro LPS-stimulated peritoneal macrophages showed a reduction in IL-6 and IL-12 release after CLP, these cytokines were less suppressed in macrophages isolated from gammadelta-/- mice. Alternatively, IL-10 release was not different between septic gammadelta+/+ and gammadelta-/- mice. Whereas T helper (Th)1 cytokine release by anti-CD3-stimulated splenocytes was significantly depressed in septic gammadelta+/+ mice, there was no such depression in gammadelta-/- mice. However, gammadelta T cell deficiency had no effect on Th2 cytokine release. These findings suggest that gammadelta T cells may play a critical role in regulating the host immune response and survival to sepsis, in part by alteration of the level of IEL CD8+gammadelta+ T cells and through the development of the Th1 response.     

Enhanced dendritic cell survival attenuates lipopolysaccharide-induced immunosuppression and increases resistance to lethal endotoxic shock

Impaired immune function and associated immunosuppression are hallmarks of septic syndromes. As part of an overall deactivation of the immune system, profound depletion of dendritic cells (DCs) occurs in both septic patients and septic mice. Such depletion of DCs is potentially associated with immunosuppression and with failure to induce a protective Th1 immune response; it may equally be predictive of fatal outcome in septic patients. To evaluate the impact of enhanced DC survival on LPS-induced immunosuppression and on survival after LPS-induced septic shock, we created a transgenic mouse model specifically overexpressing the human form of the antiapoptotic protein Bcl-2 in DCs (DC-hBcl-2 mice). DCs derived from DC-hBcl-2 mice exhibited higher resistance to maturation-induced apoptosis after LPS treatment both in vitro and in vivo. Moreover, prolongation of DC survival diminished sublethal LPS-induced DC loss and immunosuppression, with maintenance of the differentiation potential of Th1 cells and enhanced T cell activation. Such modulation of the immune response appears to constitute a key feature of the attenuated mortality observed after LPS-induced shock in DC-hBcl-2 mice. Our study therefore identifies DC death as a key determinant of endotoxin-induced immunosuppression and mortality in mice.     

Female X-chromosome mosaicism for NOX2 deficiency presents unique inflammatory phenotype and improves outcome in polymicrobial sepsis

Cellular X-chromosome mosaicism, which is unique to females, may be advantageous during pathophysiological challenges compared with the single X-chromosome machinery of males, and it may contribute to gender dimorphism in the inflammatory response. We tested the hypothesis of whether cellular mosaicism for the X-linked gp91phox (NOX2) deficiency, the catalytic component of the superoxide anion-generating NADPH oxidase complex, is advantageous during polymicrobial sepsis. Deficient, wild-type (WT), and heterozygous/mosaic mice were compared following polymicrobial sepsis initiated by cecal ligation and puncture. Compared with WT littermates, sepsis-induced mortality was improved in deficient mice, as well as in mosaic animals carrying both deficient and WT phagocyte subpopulations. In contrast, blood bacterial counts were greatest in deficient mice. Consistent with poor survival, WT mice also showed the most severe organ damage following sepsis. In mosaic animals, the deficient neutrophil subpopulations displayed increased organ recruitment and elevated CD11b membrane expression compared with WT neutrophil subpopulations within the same animal. The dynamics of sepsis-induced blood and organ cytokine content and WBC composition changes, including lymphocyte subsets in blood and bone marrow, showed differences among WT, deficient, and mosaic subjects, indicating that mosaic mice are not simply the average of the deficient and WT responses. Upon oxidative burst, interchange of oxidants between WT and deficient neutrophil subpopulations occurred in mosaic mice. This study suggests that mice mosaic for gp91phox expression have multiple advantages compared with WT and deficient mice during the septic course.     

Preferential control of induced regulatory T cell homeostasis via a Bim/Bcl-2 axis

Apoptosis has an essential role in controlling T cell homeostasis, especially during the contraction phase of an immune response. However, its contribution to the balance between effector and regulatory populations remains unclear. We found that Rag1^−/− hosts repopulated with Bim^−/− conventional CD4⁺ T cells (Tconv) resulted in a larger induced regulatory T cell (iTreg) population than mice given wild-type (WT) Tconv. This appears to be due to an increased survival advantage of iTregs compared with activated Tconv in the absence of Bim. Downregulation of Bcl-2 expression and upregulation of Bim expression were more dramatic in WT iTregs than activated Tconv in the absence of IL-2 in vitro. The iTregs generated following Tconv reconstitution of Rag1^−/− hosts exhibited lower Bcl-2 expression and higher Bim/Bcl-2 ratio than Tconv, which indicates that iTregs were in an apoptosis-prone state in vivo. A significant proportion of the peripheral iTreg pool exhibits low Bcl-2 expression indicating increased sensitivity to apoptosis, which may be a general characteristic of certain Treg subpopulations. In summary, our data suggest that iTregs and Tconv differ in their sensitivity to apoptotic stimuli due to their altered ratio of Bim/Bcl-2 expression. Modulating the apoptosis pathway may provide novel therapeutic approaches to alter the balance between effector T cells and Tregs.