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Interaction of the neuropeptide substance P (SP) and its neurokinin-1 receptor (NK-1R) plays an important role in the pathophysiology of intestinal inflammation. SP is known to stimulate production of interleukin (IL)-6 and IL-8 in the U-373-MG human astrocytoma cell line via activation of p38 MAPK (mitogen-activated protein kinase) and nuclear factor (NF)-kappaB, respectively. However, the signalling mechanisms by which SP-NK-1R interaction induces NF-kappaB activation and IL-8 expression are still not clear. In this study we demonstrate that SP stimulates IL-8 secretion and IL-8 promoter activity in the NCM460 non-transformed human colonic epithelial cell line transfected with NK-1R cDNA. Our results indicate that inhibition of endogenous Rho family proteins (RhoA, Rac1 and Cdc42) by their respective dominant negative mutants significantly decreases SP-induced IL-8 secretion and IL-8 promoter activity. We also demonstrate that SP rapidly activates RhoA, Rac1 and Cdc42 and that co-expression of the dominant negative mutants of RhoA, Rac1 and Cdc42 in NK-1R cDNA-transfected NCM460 cells significantly inhibits SP-induced NF-kappaB-dependent gene expression. These results demonstrate that Rho family small GTPases RhoA, Rac1 and Cdc42 are novel signal transducers for SP-stimulated IL-8 expression.

Background Non-alcoholic fatty liver disease (NAFLD) results from over-consumption and is a significant and increasing cause of liver failure. The type of diet that is conducive to the development of this disease has not been established and evidence-based treatment options are currently lacking. We hypothesized that the onset of hepatic steatosis is linked to the consumption of a diet with a high fat content, rather than related to excess caloric intake. In addition, we also hypothesized that fully manifested hepatic steatosis could be reversed by reducing the fat percentage in the diet of obese mice. Methods C57Bl/6J male mice were either fed a purified rodent diet containing 10% fat or a diet with 60% of calories derived from fat. A pair-feeding design was used to distinguish the effects of dietary fat content and caloric intake on dietary-induced hepatic lipid accumulation and associated injury. Livers were analyzed by quantitative RT-PCR for lipid metabolism-related gene expression. Results After 9 weeks, mice on the 60% fat diet exhibited more weight gain, insulin resistance and hepatic steatosis, compared to mice on a 10% fat diet with equal caloric intake. Furthermore, mice with established metabolic syndrome at 9 weeks showed reversal of hepatic steatosis, insulin resistance and obesity when switched to a 10% fat diet for an additional 9 weeks, independent of caloric intake. Quantitative RT-PCR revealed that transcripts related to both de novo lipogenesis and increased uptake of free fatty acids were significantly upregulated in mice pair-fed a 60% fat diet, compared to 10% fat-fed animals. Conclusion Dietary fat content, independent from caloric intake, is a crucial factor in the development of hepatic steatosis, obesity and insulin resistance in the C57Bl/6J diet-induced obesity model caused by increased uptake of free fatty acids and de novo lipogenesis. In addition, once established, all these features of the metabolic syndrome can be successfully reversed after switching obese mice to a diet low in fat. Low fat diets deserve attention in the investigation of a potential treatment of patients with NAFLD.

We report here that human monocytic͞macrophage THP-1 cells express the neurokinin 1 receptor (NK-1R), and that exposure of these cells to the proinflammatory cytokine IL-1␤ increased the expression of the NK-1R gene at the mRNA and protein levels.

Summary Women approaching advanced maternal age have extremely poor outcomes with both natural and assisted fertility. Moreover, the incidence of chromosomal abnormalities and birth defects increases with age. As of yet, there is no effective and practical strategy for delaying ovarian aging or improving oocyte quality. We demonstrate that the lifelong consumption of a diet rich in omega-3 fatty acids prolongs murine reproductive function into advanced maternal age, while a diet rich in omega-6 fatty acids is associated with very poor reproductive success at advanced maternal age. Furthermore, even short-term dietary treatment with a diet rich in omega-3 fatty acids initiated at the time of the normal age-related rapid decline in murine reproductive function is associated with improved oocyte quality, while short-term dietary treatment with omega-6 fatty acids results in very poor oocyte quality. Thus, omega-3 fatty acids may provide an effective and practical avenue for delaying ovarian aging and improving oocyte quality at advanced maternal age.

Clostridium difficile, the causative agent of antibiotic-associated colitis, mediates inflammatory diarrhea by releasing toxin A, a potent 308-kDa enterotoxin. Toxin A-induced inflammatory diarrhea involves many steps, including mucosal release of substance P (SP) corticotropin-releasing hormone (CRH) and neutrophil transmigration. Here we demonstrate that, compared with wild type, mice genetically deficient in CRH (Crh ؊/؊ ) have dramatically reduced ileal fluid secretion, epithelial cell damage, and neutrophil transmigration 4 h after intraluminal toxin A administration. This response is associated with diminished mucosal activity of the neutrophil enzyme myeloperoxidase compared with that of wildtype mice. In wild-type mice, toxin A stimulates an increase in intestinal SP content compared with buffer administration. In contrast, toxin A administration in Crh ؊/؊ mice fails to result in an increased SP content. Moreover, immunohistochemical experiments showed that CRH and SP are colocalized in some enteric nerves of wild-type mice, and this colocalization is more evident after toxin A administration. These results provide direct evidence for a major proinflammatory role for CRH in the pathophysiology of enterotoxin-mediated inflammatory diarrhea and indicate a SP-linked pathway.C orticotropin-releasing hormone (CRH), a 41-aa peptide, is a major peptide hormone that modulates the activity of the hypothalamic-pituitary-adrenal axis during stress, including inflammation (1-3). CRH mediates its effects by binding to its G protein-coupled receptor subtypes, CRH1 and CRH2 (4-6). CRH plays a dual role in the pathophysiology of inflammation. CRH secreted from the hypothalamus has indirect antiinflammatory effects via stimulation of glucocorticoid release, but CRH is also expressed peripherally, and its expression is increased in leukocytes, nerve fibers, and other cells involved in inflammatory reactions (7-12). Moreover, in vivo and in vitro studies indicate that CRH is a potent proinflammatory peptide (7,(12)(13)(14)(15).A limited number of studies indicate that CRH and its receptors play a role in the pathophysiology of intestinal inflammation. Thus, Van Tol et al. (11) found increased CRH expression in the rat cecum in colitis induced by injection of peptidoglycan-polysaccharide polymers. Increased expression of CRH-immunoreactive cells was also evident in the colonic mucosa of patients with inflammatory bowel disease (16). CRH gene expression is increased in hypothalamic neurons during colitis after intracolonic administration of 2,4,6-trinitrobenzenesulfonic acid (17) and appears to play a protective role in the worsening of experimental colitis induced by stress (18). We have recently shown that expression of CRH as well as CRH1 and CRH2 was dramatically increased in mouse ileum shortly after intraluminal administration of Clostridium difficile toxin A (19). Moreover, experiments with CRH receptor antagonists indicate that CRH plays a proinflammatory role in toxin A-induced intestinal secretion and inflammatio...

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Dietary strategies to alter the immune response to acute inflammation have the potential to improve outcomes in critically ill patients. A eucaloric ketogenic diet (EKD), composed predominantly of fat with very small amounts of carbohydrate, can provide adequate caloric support while minimizing spikes in blood glucose and reducing oxidative stress. The purpose of this study was to evaluate the effects of an EKD on glycemic control and the inflammatory response after acute endotoxemia in mice. Mice received either an EKD or a carbohydrate-based control diet (CD) for 4 weeks. Animals subsequently underwent either a 2-h fast (postprandial) or an overnight fast (postabsorptive), and half of the animals in each diet group were randomized to receive either intraperitoneal lipopolysaccharide (1 mg/kg) or an equivalent volume of saline. Glycemic response, insulin resistance, inflammatory cytokine levels, and the expression of key inflammatory and metabolic genes were measured. After endotoxin challenge, hypoglycemia was more frequent in mice fed a CD than an EKD in the postprandial period. This was due in part to the preservation of hepatic glycogen stores despite endotoxin exposure and prolonged fasting in mice fed an EKD. Furthermore, mice fed the CD had higher levels of IL-6 and TNF-α in the postabsorptive period, with a fivefold higher expression of hepatic NFκB compared to mice fed the EKD in both fasting periods. These results suggest that the unique metabolic state induced by an EKD can alter the response to acute inflammation in mice.