Background Intestinal microbiota influences the progression of colitis-associated colorectal cancer (CAC). With diet being a key determinant of the gut microbial ecology, dietary interventions are an attractive avenue for the prevention of CAC. Curcumin is the most active constituent of the ground rhizome of the Curcuma Longa plant, which has been demonstrated to have anti-inflammatory, anti-oxidative and anti-proliferative properties. Methods Il10−/− mice on 129/SvEv background were used as a model of CAC. Starting at 10 weeks of age, WT or Il10−/− mice received six weekly i.p. injections of azoxymethane (AOM) or saline, and were started on either a control or curcumin-supplemented diet. Stools were collected every 4 weeks for microbial community analysis. Mice were sacrificed at 30 weeks of age. Results Curcumin-supplemented diet increased survival, decreased colon weight/length ratio, and at 0.5%, entirely eliminated tumor burden. Although colonic histology indicated improvement with curcumin, no effects of mucosal immune responses have been observed in PBS/Il10−/− mice, and limited effects were seen in AOM/Il10−/− mice. In WT and in Il10−/− mice, curcumin increased bacterial richness, prevented age-related decrease in alpha diversity, increased the relative abundance of Lactobacillales, and decreased Coriobacterales order. Taxonomic profile of AOM/Il10−/− mice receiving curcumin was more similar to those of wild-type mice than those fed control diet. Conclusions In AOM/Il10−/− model, curcumin reduced or eliminated colonic tumor burden with limited effects on mucosal immune responses. The beneficial effect of curcumin on tumorigenesis was associated with the maintenance of a more diverse colonic microbial ecology.

. Reduced colonic microbial diversity is associated with colitis in NHE3-deficient mice.

SUMMARYNOD-like receptor (NLR) proteins are intracellular innate immune sensors/receptors that regulate immunity. This work shows that NLRX1 serves as a tumor suppressor in colitis-associated cancer (CAC) and sporadic colon cancer by keeping key tumor promoting pathways in check. Nlrx1−/− mice were highly susceptible to CAC, showing increases in key cancer-promoting pathways including nuclear factor κB (NF-κB), mitogen-activated protein kinase (MAPK), signal transducer and activator of transcription 3 (STAT3), and interleukin 6 (IL-6). The tumor-suppressive function of NLRX1 originated primarily from the non-hematopoietic compartment. This prompted an analysis of NLRX1 function in the Apcmin/+ genetic model of sporadic gastrointestinal cancer. NLRX1 attenuated Apcmin/+ colon tumorigenesis, cellular proliferation, NF-κB, MAPK, STAT3 activation, and IL-6 levels. Application of anti-interleukin 6 receptor (IL6R) antibody therapy reduced tumor burden, increased survival, and reduced STAT3 activation in Nlrx1−/−Apcmin/+ mice. As an important clinical correlate, human colon cancer samples expressed lower levels of NLRX1 than healthy controls in multiple patient cohorts. These data implicate anti-IL6R as a potential personalized therapy for colon cancers with reduced NLRX1.

Background & Aims Dysregulated Ca2+ homeostasis likely contributes to the etiology of IBD-associated loss of bone mineral density (BMD). Experimental colitis leads to decreased expression of Klotho, a protein which supports renal Ca2+ reabsorption by stabilizing TRPV5 channel on the apical membrane of distal tubule epithelial cells. Methods Colitis was induced in mice via administration of 2,4,6-trinitrobenzenesulfonic acid (TNBS) or transfer of CD4+IL10−/− and CD4+, CD45RBhi T cells. We investigated changes in bone metabolism, renal processing of Ca2+, and expression of TRPV5. Results Mice with colitis had normal serum levels of Ca2+ and parathormone. Computed tomography analysis demonstrated decreased density of cortical and trabecular bone, and there was biochemical evidence for reduced bone formation and increased bone resorption. Increased fractional urinary excretion of Ca2+ was accompanied by reduced levels of TRPV5 protein in distal convoluted tubules, with a concomitant increase in TRPV5 sialylation. In mIMCD3 cells transduced with TRPV5 adenovirus, the inflammatory cytokines tumor necrosis factor (TNF), interferon (IFN)γ, and interleukin 1β reduced levels of TRPV5 on the cell surface, leading to its degradation. Cytomix induced interaction between TRPV5 and UBR4, an E3 ubiquitin ligase; knockdown of UBR4 with small interfering RNAs prevented cytomix-induced degradation of TRPV5. The effects of cytokines on TRPV5 were not observed in cells stably transfected with membrane-bound Klotho; TRPV5 expression was preserved when colitis was induced with TNBS in transgenic mice that overexpress Klotho or in mice with T-cell transfer colitis injected with soluble recombinant Klotho. Conclusion Following induction of colitis in mice via TNBS administration or T-cell transfer, TNF and IFNγ reduce expression and activity of Klotho, which would otherwise protect TRPV5 from hyper-sialylation and cytokine-induced TRPV5 endocytosis, UBR4-dependent ubiquitination, degradation, and urinary wasting of Ca2+.

(IBD). Vitamin D 3 has been considered a viable adjunctive therapy in IBD. However, vitamin D 3 plays a pleiotropic role in bone modeling and regulates the bone formation-resorption balance, depending on the physiological environment, and supplementation during active IBD may have unintended consequences. We evaluated the effects of vitamin D 3 supplementation during the active phase of disease on colonic inflammation, BMD, and bone metabolism in an adoptive IL-10 Ϫ/Ϫ CD4 ϩ T cell transfer model of chronic colitis. High-dose vitamin D 3 supplementation for 12 days during established disease had negligible effects on mucosal inflammation. Plasma vitamin D 3 metabolites correlated with diet, but not disease, status. Colitis significantly reduced BMD. High-dose vitamin D 3 supplementation did not affect cortical bone but led to a further deterioration of trabecular bone morphology. In mice fed a high vitamin D3 diet, colitis more severely impacted bone formation markers (osteocalcin and bone alkaline phosphatase) and increased bone resorption markers, ratio of receptor activator of NF-B ligand to osteoprotegrin transcript, plasma osteoprotegrin level, and the osteoclast activation marker tartrate-resistant acid phosphatase (ACp5). Bone vitamin D receptor expression was increased in mice with chronic colitis, especially in the high vitamin D3 group. Our data suggest that vitamin D3, at a dose that does not improve inflammation, has no beneficial effects on bone metabolism and density during active colitis or may adversely affect BMD and bone turnover. These observations should be taken into consideration in the planning of further clinical studies with high-dose vitamin D3 supplementation in patients with active IBD. inflammatory bowel disease; bone mineral density; 1,25-dihydroxyvitamin D3; bone turnover INFLAMMATORY BOWEL DISEASE (IBD) is characterized by an inappropriate and persistent immune response against commensal intestinal microbiota and by intestinal inflammation and mucosal damage. However, this chronic inflammatory disease can also affect bone metabolism and bone mineral density (BMD), with osteopenia and osteoporosis as the two major extraintestinal symptoms. Low BMD has been reported in Crohn's disease (CD) and ulcerative colitis (UC). The relative risk of fracture is 40% higher in the IBD patient than in the general population (3), and although this has not been systematically studied, it is expected that children with IBD do not reach the optimal peak bone mass in early adulthood. The prevalence of osteopenia and osteoporosis in IBD varies depending on the population studied, geographic location, and study design, with a range of 22-77% for osteopenia and 17-41% for osteoporosis (3). Vitamin D 3 increases intestinal and renal Ca 2ϩ and P i absorption and is typically considered an anabolic hormone that positively affects osteoblast differentiation and bone matrix synthesis. Moreover, its immunomodulatory effects, including promotion of macrophage antimicrobial responses to pathogens, regulation of ...

Poly(ADP‐ribose) polymerases (PARPs), including its most abundant isoform PARP1, catalyzes the transfer of ADP‐ribose units from NAD+ to a number of target proteins. In vivo studies have demonstrated that genetic ablation or pharmacological inhibition of PARP1 ameliorates experimental colitis pathophysiological changes. However, the studies have failed to characterize the mechanism by which PARP1 inhibition is protective in colitis. We evaluated the effect of PARP1 deletion in T cell‐mediated colitis in the following scenarios: adoptive naïve CD4+CD45RBHigh T cell transfer from WT or Parp1‐/‐ mice into Rag2‐/‐ and Rag2‐/‐ x Parp1‐/‐ DKO mice and DSS‐induced colitis in WT, Parp1‐/‐, Rag2‐/‐, and Parp1‐/‐Rag2‐/‐ DKO mice. Compared to WT mice, Parp1‐/‐ mice were protected from DSS‐induced colitis. However, WT and Parp1‐/‐naïve T cells were equally colitogenic in Rag2‐/‐ recipients. DKO mice transferred with WT T cells or treated with DSS showed only marginal improvement. Our preliminary data indicate that Parp1 is dispensable in naïve and effector T‐cells, while Parp1‐/‐ mice are protected from colitis resulting from DSS‐induced epithelial damage. Moreover, the lack of significant protection in T cell transfer or DSS models of colitis in DKO mice suggests a pathogenic role of Parp1 in the innate immune cells and regulatory T cells during colitis. Grant Funding Source: NIH Grant 1K01DK099268‐01

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KGF (FGF-7), an epithelial cell growth factor, can lessen the injurious effects of post-BMT lung injury (IPS) and GVHD when given prior to conditioning of the recipient. We reported that mice treated with KGF pre-conditioning had elevated systemic levels of Th2-type cytokines at the time of BM infusion which was associated with less GVHD. In non-conditioned SCID mice that developed lethal GVHD from allogeneic T cells, KGF pretreatment also increased Th2 cytokines implying that KGF alone was responsible for the systemic Th2 milieu and not a response that was dependent on conditioning. In the post-BMT lung, KGF pretreatment did not alter the cellular composition of the inflammatory infiltrate but resulted in decreased levels of immune activation markers compared to non-treated control IPS mice. The mechanism of this immunomodulatory effect is unknown. Since Th2 responses are dependent on the STAT-6 transcription factor, we studied the effects of KGF in our established murine IPS model using lethally-conditioned STAT-6−/− mice (B6 background) as recipients of allogeneic B10.BR BM and T cells and compared to WT hosts. STAT-6−/− mice pretreated with KGF (5 mg/kg s.c. for 3 days) had an accelerated mortality rate and poorer survival compared to WT recipients. Examination of GVHD target organs revealed that KGF pretreatment ameliorated liver GVHD in WT and STAT-6−/− recipients. In contrast, the benefit of KGF for GVHD of the colon was only seen in the WT but not in STAT-6−/− recipients, consistent with post-BMT body weights and survival. Whole body plethysmograph was done to evaluate lung function at 30 cm water pressure on day 7 post-BMT, a time when acute IPS is manifested in this model. WT but not STAT-6−/− mice pretreated with KGF had improved lung compliance compared to their non-treated counterparts (p<0.05). KGF-pretreated STAT-6−/− recipients also had increased lung weights compared to control- or KGF-treated WT mice with IPS (p<0.05). Histological analysis of lung cryosections taken on day 7 post-BMT showed that all recipients of allogeneic BM and T cells had increased inflammatory cells in their lungs, consistent with acute IPS. However, STAT-6−/− hosts with IPS had elevated numbers of CD4+ T cells in their lungs compared to WT hosts with IPS (18.3%±1.5 vs 11.6%±2.7, p=0.01). Other cell phenotypes did not differ. However, KGF-pretreated STAT-6−/− mice had significantly elevated levels (10-fold) of IL-17, CCL2 and CCL8 in their bronchoalveolar lavage fluid on day 7 post-BMT compared to their WT KGF-treated counterparts implying a skewing to a Th17 inflammatory response in STAT-6−/− mice. These data indicate that the STAT-6 pathway plays a role in the cytoprotective function of KGF and this STAT-6-dependency is tissue-specific. To achieve the full benefits of KGF and other cytoprotective agents, it will be important to take into account the immune modulatory mechanisms operative in individual GVHD target organs.