IAP is involved in the maintenance of normal gut microbial homeostasis and may have therapeutic potential against dysbiosis and pathogenic infections.

Ramasamy, S., S. Singh, P. Taniere, M. J. S. Langman, and M. C. Eggo. Sulfide-detoxifying enzymes in the human colon are decreased in cancer and upregulated in differentiation. Am J Physiol Gastrointest Liver Physiol 291: G288 -G296, 2006. First published February 23, 2006 doi:10.1152/ajpgi.00324.2005.-H 2S is highly toxic and selectively inhibits butyrate oxidation in colonocytes. Ineffective detoxification may result in mucosal insult, inflammation, and ultimately in colorectal cancer (CRC). Rhodanese can detoxify H 2S and is comprised of two isoenzymes: thiosulfate sulfurtransferase (TST) and mercaptopyruvate sulfurtransferase (MST). Using specific antisera to discriminate TST from MST, we found that only TST could detoxify H2S. In sections of normal colon, both enzymes were located on the luminal mucosal surface, and they were expressed in the colonocytes but not in the mucin-secreting goblet cells. Expression of both enzymes was focally lost in ulcerative colitis and markedly reduced in advanced colon cancer, the disease progression correlating with the decreased expression of MST and TST. In HT-29 cells, a human colon cancer cell line, TST activity and expression were significantly increased by butyrate and by histone deacetylase inhibition, agents that promote HT-29 cell differentiation. Sulfide (0.1 mM) also increased TST activity, but higher sulfide concentrations (0.3-3 mM) were toxic. Preincubation in butyrate to increase TST expression, decreased sensitivity of the cells to sulfide toxicity. We conclude that decreased expression of TST (or MST) is a tumor marker for CRC. TST expression is increased in colonocyte differentiation. Dysregulation of TST expression and activity resulting in inability to effectively detoxify could be a factor in the cell loss and inflammation that accompany ulcerative colitis and ultimately then in CRC. colorectal cancer; thiosulfate sulfurtransferase; butyrate; HT-29 IN THE HUMAN COLON, anaerobic bacteria produce hydrogen sulfide (H 2 S) (25), which inhibits the oxidation of short-chain fatty acids (SCFA) (18,19). SCFAs are the main energy source (Ͼ70% ) for colonocytes (18) and also contribute 5-10% of the total energy supplies in humans. Inhibition of their oxidation in the colon will result in colonocyte starvation and death, resulting in inflammation. This, the "energy deficiency" hypothesis of Roediger (18), is postulated to lead to ulcerative colitis (UC). Many studies have shown that patients with UC have increased colorectal cancer (CRC) risk, with Prior et al. (16) estimating this as an 11-fold excess. On the basis of these studies, we suggest that the expression of enzymes important in sulfide detoxification is essential to preserve a healthy mucosa and that derangements in expression may accompany UC and CRC.We recently showed that colonic mucosal rhodanese can detoxify hydrogen sulfide (15) and thus might protect against mucosal injury. Rhodanese is a mitochondrial enzyme, present in all living organisms from bacteria to humans, and is thought to play a cen...

Intestinal alkaline phosphatase (IAP) is a small intestinal brush border enzyme that has been shown to function as a gut mucosal defense factor, but its precise mechanism of action remains unclear. We investigated the effects of IAP on specific bacteria and bacterial components to determine its molecular targets. Purulent fluid from a cecal ligation and puncture model, specific live and heat-killed bacteria (Escherichia coli, Salmonella typhimurium, and Listeria monocytogenes), and a variety of proinflammatory ligands (LPS, CpG DNA, Pam-3-Cys, flagellin, and TNF) were incubated with or without calf IAP (cIAP). Phosphate release was determined by using a malachite green assay. The various fluids were applied to target cells (THP-1, parent HT-29, and IAP-expressing HT-29 cells) and IL-8 secretion measured by ELISA. cIAP inhibited IL-8 induction by purulent fluid in THP-1 cells by >35% (P < 0.005). HT29-IAP cells had a reduced IL-8 response specifically to gram-negative bacteria; >90% reduction compared with parent cells (P < 0.005). cIAP had no effect on live bacteria but attenuated IL-8 induction by heat-killed bacteria by >40% (P < 0.005). cIAP exposure to LPS and CpG DNA caused phosphate release and reduced IL-8 in cell culture by >50% (P < 0.005). Flagellin exposure to cIAP also resulted in reduced IL-8 secretion by >40% (P < 0.005). In contrast, cIAP had no effect on TNF or Pam-3-Cys. The mechanism of IAP action appears to be through dephosphorylation of specific bacterial components, including LPS, CpG DNA, and flagellin, and not on live bacteria themselves. IAP likely targets these bacterially derived molecules in its role as a gut mucosal defense factor.

Background The brush border enzyme intestinal alkaline phosphatase (IAP) functions as a gut mucosal defense factor and is protective against dextran sulfate sodium (DSS)-induced acute injury in rats. The present study evaluated the potential therapeutic role for orally administered calf IAP (cIAP) in two independent mouse models of chronic colitis: (1) DSS-induced chronic colitis, and (2) chronic spontaneous colitis in Wiskott-Aldrich Syndrome protein (WASP) deficient (knockout) mice that is accelerated by irradiation. Methods The wild-type (WT) and IAP knockout (IAP-KO) mice received 4 cycles of 2% DSS ad libitum for 7 days. Each cycle was followed by a 7-day DSS-free interval during which mice received either cIAP or vehicle in the drinking water. The WASP-KO mice received either vehicle or cIAP for 6 weeks beginning on the day of irradiation. Results Microscopic colitis scores of DSS-treated IAP-KO mice were higher than DSS-treated WT mice (52 ± 3.8 vs. 28.8 ± 6.6, respectively, P < 0.0001). cIAP treatment attenuated the disease in both groups (KO = 30.7 ± 6.01, WT = 18.7 ± 5.0, P < 0.05). In irradiated WASP-KO mice cIAP also attenuated colitis compared to control groups (3.3 ± 0.52 vs. 6.2 ± 0.34, respectively, P < 0.001). Tissue myeloperoxidase activity and pro-inflammatory cytokines were significantly decreased by cIAP treatment. Conclusions Endogenous IAP appears to play a role in protecting the host against chronic colitis. Orally administered cIAP exerts a protective effect in two independent mouse models of chronic colitis and may represent a novel therapy for human IBD.

Background and Aims Intestinal alkaline phosphatase (IAP) is a gut mucosal defense factor known to dephosphorylate lipopolysaccharide (LPS); however, the role of IAP in the gut response to luminal bacteria remains poorly defined. We investigated immune responses of wild-type (WT) and IAP-knockout (IAP-KO) mice to LPS and Salmonella typhimurium challenges. Methods Cryostat sectioning and standard indirect immunohistochemical staining for major histocompatibility complex (MHC) class II molecules were performed on liver tissue from WT and IAP-KO mice. WT and IAP-KO mice were orally gavaged with S. typhimurium; bacterial translocation to mesenteric nodes, liver, and spleen was determined by tissue homogenization and plating. In other experiments, WT and IAP-KO mice received intraperitoneal injections of LPS, with subsequent quantification of complete blood counts and serum interleukin (IL)-6 by enzyme-linked immunosorbent assay (ELISA). WT and IAP-KO whole blood were plated and stimulated with LPS and Pam-3-Cys, followed by cytokine assays. Results Immunohistologic liver examinations showed increased expression of MHC class II molecules in IAP-KO mice. Following S. typhimurium challenge, WT mice appeared moribund compared with IAP-KO mice, with increased bacterial translocation. WT mice had [50% decrease (P \ .005) in platelets and 1.8-fold (P \ .05) increased serum IL-6 compared with IAP-KO mice in response to LPS injections. IAP-KO whole-blood stimulation with LPS and Pam-3-Cys resulted in increased IL-6 and tumor necrosis factor (TNF)-alpha secretion compared with WT. Conclusions IAP-KO mice exhibit characteristics consistent with local LPS tolerance. Whole-blood response of IAP-KO mice did not reflect systemic tolerance. These data suggest that IAP is a local immunomodulating factor, perhaps regulating LPS–toll-like receptor 4 (TLR4) interaction between commensal microflora and intestinal epithelium.

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Peritoneal irrigation with cIAP significantly enhances survival in a mouse model of peritonitis, likely through reduction of local inflammation and remote organ damage. We suggest that intraperitoneal cIAP irrigation could be a novel therapy for intra-abdominal sepsis.

Background and Aims-Intestinal alkaline phosphatase (IAP) is a gut mucosal defense factor known to dephosphorylate lipopolysaccharide (LPS); however, the role of IAP in the gut response to luminal bacteria remains poorly defined. We investigated immune responses of wild-type (WT) and IAP-knockout (IAP-KO) mice to LPS and Salmonella typhimurium challenges. Methods-Cryostat sectioning and standard indirect immunohistochemical staining for major histocompatibility complex (MHC) class II molecules were performed on liver tissue from WT and IAP-KO mice. WT and IAP-KO mice were orally gavaged with S. typhimurium; bacterial translocation to mesenteric nodes, liver, and spleen was determined by tissue homogenization and plating. In other experiments, WT and IAP-KO mice received intraperitoneal injections of LPS, with subsequent quantification of complete blood counts and serum interleukin (IL)-6 by enzymelinked immunosorbent assay (ELISA). WT and IAP-KO whole blood were plated and stimulated with LPS and Pam-3-Cys, followed by cytokine assays. Results-Immunohistologic liver examinations showed increased expression of MHC class II molecules in IAP-KO mice. Following S. typhimurium challenge, WT mice appeared moribund compared with IAP-KO mice, with increased bacterial translocation. WT mice had [50% decrease (P \ .005) in platelets and 1.8-fold (P \ .05) increased serum IL-6 compared with IAP-KO mice in response to LPS injections. IAP-KO whole-blood stimulation with LPS and Pam-3-Cys resulted in increased IL-6 and tumor necrosis factor (TNF)-alpha secretion compared with WT. Conclusions-IAP-KO mice exhibit characteristics consistent with local LPS tolerance. Wholeblood response of IAP-KO mice did not reflect systemic tolerance. These data suggest that IAP is a local immunomodulating factor, perhaps regulating LPS-toll-like receptor 4 (TLR4) interaction between commensal microflora and intestinal epithelium.