Granulocytes mediates the Fas-L-associated apoptosis during lung metastasis of melanoma that determines the metastatic behaviour

The survival of tumour cells in a new tissue environment is crucial for tumour metastasis. Factors contributing to the death of tumour cells during metastasis are not completely understood. In murine melanoma model, activation of Fas (CD95, APO-1) signal in tumour cells reduces their lung metastasis potential, which may be associated with an induction of apoptosis in tumours. To elucidate the cellular mechanism, we used a Fas-ligand (Fas-L) specific ribozyme (Fas-Lribozyme) to suppress the expression of Fas-L but not Fas or TNF-α in B16F10 melanoma cells. The Fas-Lribozyme-carrying cells grew slightly faster in vitro with better viability than controls. Suppression of Fas-L in B16F10 melanoma cells by Fas-Lribozyme enhanced lung metastasis of the cells in C57BL/6 mice, and that was correlated with reductions in both apoptotic tumour cells and granulocytic infiltration. Mice depleted of granulocytes, but not CD4+ and CD8+ cells, showed a greatly elevated susceptibility to lung metastasis. Moreover, apoptosis in tumour cells was significantly reduced in granulocyte-depleted mice during the course of tumour formation. Taken together, our findings indicate that Fas-L-associated apoptosis in tumour cells determines the metastasis behaviour of melanoma in the lung and this apoptosis is primarily mediated by the cytotoxicity of recruited granulocytes

Much Ado About the TPP’s Effect on Pharmaceuticals

Ocular antigens are sequestered behind the blood-retina barrier and the ocular environment protects ocular tissues from autoimmune attack. The signals required to activate autoreactive T cells and allow them to cause disease in the eye remain in part unclear. In particular, the consequences of peripheral presentation of ocular antigens are not fully understood. We examined peripheral expression and presentation of ocular neo-self-antigen in transgenic mice expressing hen egg lysozyme (HEL) under a retina-specific promoter. High levels of HEL were expressed in the eye compared to low expression throughout the lymphoid system. Adoptively transferred naïve HEL-specific CD4+ T cells proliferated in the eye draining lymph nodes, but did not induce uveitis. By contrast, systemic infection with a murine cytomegalovirus (MCMV) engineered to express HEL induced extensive proliferation of transferred naïve CD4+ T cells, and significant uveoretinitis. In this model, wild-type MCMV, lacking HEL, did not induce overt uveitis, suggesting that disease is mediated by antigen-specific peripherally activated CD4+ T cells that infiltrate the retina. Our results demonstrate that retinal antigen is presented to T cells in the periphery under physiological conditions. However, when the same antigen is presented during viral infection, antigen-specific T cells access the retina and autoimmune uveitis ensues

Loss of Sex and Age Driven Differences in the Gut Microbiome Characterize Arthritis-Susceptible *0401 Mice but Not Arthritis-Resistant *0402 Mice

<div><h3>Background</h3><p>HLA-DRB1*0401 is associated with susceptibility, while HLA-DRB1*0402 is associated with resistance to developing rheumatoid arthritis (RA) and collagen-induced arthritis in humans and transgenic mice respectively. The influence of gut-joint axis has been suggested in RA, though not yet proven.</p> <h3>Methodology/Principal Findings</h3><p>We have used HLA transgenic mice carrying arthritis susceptible and -resistant HLA-DR genes to explore if genetic factors and their interaction with gut flora gut can be used to predict susceptibility to develop arthritis. Pyrosequencing of the 16S rRNA gene from the fecal microbiomes of DRB1*0401 and DRB1*0402 transgenic mice revealed that the guts of *0401 mice is dominated by a Clostridium-like bacterium, whereas the guts of *0402 mice are enriched for members of the <em>Porphyromonadaceae</em> family and <em>Bifidobacteria</em>. DRB1*0402 mice harbor a dynamic sex and age-influenced gut microbiome while DRB1*0401 mice did not show age and sex differences in gut microbiome even though they had altered gut permeability. Cytokine transcripts, measured by rtPCR, in jejuna showed differential TH17 regulatory network gene transcripts in *0401 and *0402 mice.</p> <h3>Conclusions/Significance</h3><p>We have demonstrated for the first time that HLA genes in association with the gut microbiome may determine the immune environment and that the gut microbiome might be a potential biomarker as well as contributor for susceptibility to arthritis. Identification of pathogenic commensal bacteria would provide new understanding of disease pathogenesis, thereby leading to novel approaches for therapy.</p> </div

Role of the Mitochondria in Immune-Mediated Apoptotic Death of the Human Pancreatic β Cell Line βLox5

Mitochondria are indispensable in the life and death of many types of eukaryotic cells. In pancreatic beta cells, mitochondria play an essential role in the secretion of insulin, a hormone that regulates blood glucose levels. Unregulated blood glucose is a hallmark symptom of diabetes. The onset of Type 1 diabetes is preceded by autoimmune-mediated destruction of beta cells. However, the exact role of mitochondria has not been assessed in beta cell death. In this study, we examine the role of mitochondria in both Fas- and proinflammatory cytokine-mediated destruction of the human beta cell line, βLox5. IFNγ primed βLox5 cells for apoptosis by elevating cell surface Fas. Consequently, βLox5 cells were killed by caspase-dependent apoptosis by agonistic activation of Fas, but only after priming with IFNγ. This beta cell line undergoes both apoptotic and necrotic cell death after incubation with the combination of the proinflammatory cytokines IFNγ and TNFα. Additionally, both caspase-dependent and -independent mechanisms that require proper mitochondrial function are involved. Mitochondrial contributions to βLox5 cell death were analyzed using mitochondrial DNA (mtDNA) depleted βLox5 cells, or βLox5 ρ0 cells. βLox5 ρ0 cells are not sensitive to IFNγ and TNFα killing, indicating a direct role for the mitochondria in cytokine-induced cell death of the parental cell line. However, βLox5 ρ0 cells are susceptible to Fas killing, implicating caspase-dependent extrinsic apoptotic death is the mechanism by which these human beta cells die after Fas ligation. These data support the hypothesis that immune mediators kill βLox5 cells by both mitochondrial-dependent intrinsic and caspase-dependent extrinsic pathways

Metabolic Regulation in Progression to Autoimmune Diabetes

Recent evidence from serum metabolomics indicates that specific metabolic disturbances precede β-cell autoimmunity in humans and can be used to identify those children who subsequently progress to type 1 diabetes. The mechanisms behind these disturbances are unknown. Here we show the specificity of the pre-autoimmune metabolic changes, as indicated by their conservation in a murine model of type 1 diabetes. We performed a study in non-obese prediabetic (NOD) mice which recapitulated the design of the human study and derived the metabolic states from longitudinal lipidomics data. We show that female NOD mice who later progress to autoimmune diabetes exhibit the same lipidomic pattern as prediabetic children. These metabolic changes are accompanied by enhanced glucose-stimulated insulin secretion, normoglycemia, upregulation of insulinotropic amino acids in islets, elevated plasma leptin and adiponectin, and diminished gut microbial diversity of the Clostridium leptum group. Together, the findings indicate that autoimmune diabetes is preceded by a state of increased metabolic demands on the islets resulting in elevated insulin secretion and suggest alternative metabolic related pathways as therapeutic targets to prevent diabetes

Gut Microbiota, Probiotics and Diabetes

Diabetes is a condition of multifactorial origin, involving several molecular mechanisms related to the intestinal microbiota for its development. In type 2 diabetes, receptor activation and recognition by microorganisms from the intestinal lumen may trigger inflammatory responses, inducing the phosphorylation of serine residues in insulin receptor substrate-1, reducing insulin sensitivity. In type 1 diabetes, the lowered expression of adhesion proteins within the intestinal epithelium favours a greater immune response that may result in destruction of pancreatic β cells by CD8+ T-lymphocytes, and increased expression of interleukin-17, related to autoimmunity. Research in animal models and humans has hypothesized whether the administration of probiotics may improve the prognosis of diabetes through modulation of gut microbiota. We have shown in this review that a large body of evidence suggests probiotics reduce the inflammatory response and oxidative stress, as well as increase the expression of adhesion proteins within the intestinal epithelium, reducing intestinal permeability. Such effects increase insulin sensitivity and reduce autoimmune response. However, further investigations are required to clarify whether the administration of probiotics can be efficiently used for the prevention and management of diabetes