Masaoki Yonezawa

A randomized controlled trial on the efficacy of synbiotic versus probiotic or prebiotic treatment to improve the quality of life in patients with ulcerative colitis.

OBJECTIVE Studies suggest that synbiotic therapy could prove more effective in the treatment of ulcerative colitis (UC) than therapies limited to probiotics or prebiotics. This study compared the effect of each of these therapies in the treatment of UC. METHODS One hundred twenty outpatients with UC were randomly sorted into three groups of 40 patients each for probiotic, prebiotic, or synbiotic therapy. The probiotic group ingested one daily capsule consisting of Bifidobacterium longum 2 x 10(9) colony-forming units and the prebiotic group ingested daily 8.0-g doses of psyllium. The synbiotic group underwent both treatments. All patients completed Inflammatory Bowel Disease Questionnaires (IBDQs) at the onset of the trial, at the 2-wk midpoint, and at the 4-wk end of the trial. Blood variables were also evaluated in a subset of 32 patients randomly selected from all groups and values were compared with IBDQ scores. RESULTS Thirty-one patients in the probiotic group, 31 in the prebiotic group, and 32 in the synbiotic group qualified for analyses. The remaining 26 patients had incomplete questionnaires. Total IBDQ scores improved within groups by the end of the trial (probiotics 162 to 169, NS; prebiotics 174 to 182, NS; synbiotics 168 to 176, P = 0.03). Individual scores improved as follows: probiotics, emotional function (P = 0.03); prebiotics, bowel function (P = 0.04); and synbiotics, systemic and social functions (P = 0.008 and P = 0.02). C-reactive protein decreased significantly only with synbiotic therapy (from 0.59 to 0.14 mg/dL, P = 0.04). There were no adverse events. CONCLUSION Patients with UC on synbiotic therapy experienced greater quality-of-life changes than patients on probiotic or prebiotic treatment. These data suggest that synbiotic therapy may have a synergistic effect in the treatment of UC.

Prevention of nonsteroidal anti-inflammatory drug–induced small-intestinal injury by prostaglandin: a pilot randomized controlled trial evaluated by capsule endoscopy

BACKGROUND There is no known preventive agent against nonsteroidal anti-inflammatory drug (NSAID) induced small-intestinal injury. OBJECTIVE To evaluate by capsule endoscopy whether coadministration of prostaglandin (PG) can prevent small-intestinal damage induced by short-term NSAID treatment. DESIGN Single-blind, randomized, controlled trial. SETTING All procedures were performed at Nippon Medical School. SUBJECTS Thirty-four healthy male volunteers. METHODS All subjects were randomly assigned to 2 groups: an NSAID-control group, who underwent NSAID (diclofenac sodium, 25 mg 3 times daily) and omeprazole (20 mg once daily) treatment, and an NSAID-PG group, who received PG (misoprostol, 200 microg 3 times daily) in addition to the same NSAID-omeprazole treatment. Eligible subjects, 15 per group, underwent capsule endoscopy before and 14 days after treatment. MAIN OUTCOME MEASUREMENTS The number of mucosal breaks at capsule endoscopy. RESULTS NSAID treatment significantly increased the mean (SD) number of mucosal breaks per subject, from a basal level of 0.1 +/- 0.3 up to 2.9 +/- 6.3 lesions in the NSAID-control group (P = .012). In contrast, there was no significant change in the mean number of mucosal breaks before and after PG cotreatment (P = 0.42). Thus, the mean number of posttreatment mucosal breaks per subject was significantly higher in the NSAID-control group than in the NSAID-PG group (P = .028). There was a significant increase in the percentage of subjects in the NSAID-control group, with at least 1 mucosal break after treatment (from 6.7% to 53.3%), whereas there was no change in the incidence of mucosal breaks in the NSAID-PG group, which remained at 13.3%. (P = .002). LIMITATIONS Single-center, open-label study. CONCLUSIONS PG cotherapy reduced the incidence of small-intestinal lesions induced by a 2-week administration of diclofenac sodium.

Distribution of small intestinal mucosal injuries as a result of NSAID administration.

Eur J Clin Invest 2010; 40 (6): 504–510

Monocyte chemoattractant protein 1 and macrophage cyclooxygenase 2 expression in colonic adenoma.

Background and aims: Cyclooxygenase 2 (COX-2) expression in subepithelial macrophages of colorectal adenoma has been suggested as the first in a series of steps leading to colorectal tumorigenesis. We tested the hypothesis that chemokines released from human colorectal adenoma epithelium might be involved in COX-2 expression in macrophages of the lamina propria. Methods: Endoscopic samples of sporadic colorectal adenomas were tested by enzyme linked immunosorbent assay for chemokines involved in macrophage chemotaxis. Localisation of adenoma macrophage chemoattractant protein 1 (MCP-1) and COX-2 were determined by immunohistochemistry. The effects of MCP-1, in the presence or absence of celecoxib, on COX-2 expression, and prostaglandin (PG) E2 and vascular endothelial growth factor (VEGF) release, were examined in human macrophages isolated from peripheral blood. Results: MCP-1 levels were markedly higher in adenoma with mild-moderate dysplasia (129.7 (19.9) pg/mg protein) and severe dysplasia (227.9 (35.4) pg/mg protein) than in normal colonic mucosa (55.8 (4.2) pg/mg protein). Other chemokine levels, macrophage inflammatory proteins (MIP)-1α and MIP-1β, and the chemokine regulated on activation of normal T cell expressed and secreted (RANTES) did not vary significantly between adenoma and normal mucosa. MCP-1 levels in both adenoma and normal colonic mucosa increased significantly three hours after tissue cultivation in vitro. MCP-1 immunoreactivity was restricted to the adenoma epithelium, with no reactivity seen in adjacent normal epithelial cells. MCP-1 stimulated COX-2 expression and PGE2 and VEGF release in human macrophages. Celecoxib, a selective COX-2 inhibitor, inhibited MCP-1-induced PGE2 and VEGF release in macrophages. Addition of exogenous PGE2 reversed this inhibitory effect on VEGF release, suggesting that MCP-1 in adenoma epithelial cells might be involved in COX-2 expression and subsequent macrophage activation. Conclusions: MCP-1 in colorectal adenoma epithelial cells might be involved in macrophage migration and COX-2 expression, leading to the subsequent development of colonic adenoma.

Microsomal prostaglandin E synthase protein levels correlate with prognosis in colorectal cancer patients

The aim of this study is to investigate the expression of three prostaglandin E synthase (PGES) isomers in colorectal cancer (CRC) tissue and to evaluate their relationship to clinicopathological factors and patient prognosis. Microsomal PGES (mPGES)-1, mPGES-2, cytosolic PGES (cPGES) and cyclooxygenase (COX)-2 protein expression were analyzed by real-time polymerase chain reaction and Western blot. The localization of each PGES and COX-2 protein was examined by immunohistochemistry in 155 surgical resections and correlated to clinicopathological factors and patient prognosis. mPGES-1 mRNA and protein levels were significantly higher in CRC than in paired normal tissues. mPGES-1 immunoreactivity localized in cancer cells in 43% of cases. mPGES-2 immunoreactivity was significantly more pronounced in cancer cells than in adjacent normal epithelium in 36% of cases. cPGES immunoreactivity was homogeneous in cancer cells and thus determined constitutive. mPGES-1 and mPGES-2 correlated with significantly worse prognosis in stage I–III patients. These results indicate that mPGES-1 and mPGES-2 may each play a role in CRC progression.

Heregulin-Induced VEGF Expression via the ErbB3 Signaling Pathway in Colon Cancer

Background/Aims: Heregulin (HRG/NRG) ligation to erbB3/4 promotes their respective heterodimerization with erbB2, and consequent erbB2 tyrosine phosphorylation. Although HRG has been shown to be expressed in a variety of cancer tissues, its expression and role in colon cancer have yet to be clarified. We therefore examined the link between the expression of these erbB receptors, and the relationship between HRG and vascular endothelial growth factor (VEGF) expression in colon cancer. Methods: We analyzed the effects of HRG on VEGF secretion in 6 colorectal cancer cell lines by enzyme-linked immunosorbent assay, and HRG-induced p85 subunit of phosphatidylinositol 3-kinase (p85 PI-3K), Akt, extracellular signal-regulated kinase 1/2 (ERK1/2), and p38 mitogen-activated protein kinase (p38 MAPK) activation in Caco-2 colon cancer cell lines by Western blot. We also examined HRG and VEGF mRNA expression in 16 colon cancer biopsy samples by real-time PCR. The localization of HRG and VEGF protein expression in colon cancer tissue was detected by immunohistochemistry. Results: Exogenous HRG stimulated VEGF secretion in all cell lines examined, and VEGF mRNA expression in Caco-2 cells. HRG also activated p85 PI-3K, Akt, ERK1/2, and p38 MAPK. VEGF secretion was inhibited by both specific p38 MAPK inhibitor and proteasome inhibitor that inhibit nuclear factor kappa B (NF-κB) activation. In colon cancer biopsy samples, HRG mRNA expression correlated with VEGF mRNA expression. HRG immunoreactivity was observed both in cancer cells and in mesenchymal cells in colon cancer tissues. Conclusion: These data suggest that HRG might affect colon cancer growth by regulating VEGF secretion via the erbB3 signaling pathway through autocrine and paracrine mechanisms.

Tomoregulin ectodomain shedding by proinflammatory cytokines

The shedding mechanism for the tomoregulin (TR) ectodomain, which contains two follistatin modules and a single epidermal growth factor (EGF)-like domain, remains unclear. Our study provides the first evidence that proinflammatory cytokines, IL-1beta and TNF-alpha, induce TR-ectodomain shedding in cultured A172 human glioma cells, without affecting TR mRNA expression. In addition, it appears that this shedding process is induced via activation of the NF-kappaB signaling pathway; with consequent increase in the production of metalloproteinases. Furthermore, since due to erbB4 tyrosine phosphorylation TR may have functions similar to EGF/neuregulin (NRG) family growth factors, our results suggest that following inflammation-induced injury, increases in TR shedding may contribute to tissue growth and repair in the central nervous system.

Prevention of traditional NSAID-induced small intestinal injury: recent preliminary studies using capsule endoscopy.

Capsule endoscopy and balloon endoscopy, advanced modalities that now allow for full investigation of the entire small intestine, have revealed that non-steroidal anti-inflammatory drugs (NSAIDs) can cause a variety of abnormalities in the small intestine. Traditional NSAIDs can induce small intestinal injuries in over 50% of patients. Several studies have shown that the preventive effect of proton pump inhibitors does not extend to the small intestine, suggesting that concomitant therapy may be required to prevent small intestinal side effects associated with traditional NSAIDs use. Recently, several randomized controlled trials used capsule endoscopy to evaluate the preventive effect of certain drugs on NSAID-induced small intestinal injuries. These studies show that misoprostol and rebamipide have a preventive effect for NSAID-induced small intestinal mucosal injuries. However, these studies included only a small series of healthy volunteers and tested short-term NSAID treatment. Therefore, further extensive studies are clearly required to ascertain the beneficial effect of these drugs.

A novel epidermal growth factor-like molecule containing two follistatin modules stimulates tyrosine phosphorylation of ERBB-4 in MKN28 gastric cancer cells

We have isolated a gene from stomach fibroblasts encoding novel proteins containing two follistatin modules which might bind TGF-beta-related growth factors and a single epidermal growth factor (EGF)-like domain which is closely related to EGF/Neuregulin (NRG) family growth factors. Sequence analysis revealed novel cDNA clones, the protein products of which were designated tomoregulin (TR) and consisted of at least three isoforms which were distinguished by their cytoplasmic domains. The cytoplasmic domains in all isoforms were short and contained potential G-protein activating motifs. Precursors of TR (Pro-TR) are glycosylated transmembrane proteins. Two secreted soluble forms resulting from proteolytic cleavage were distinguished by the presence or absence of the EGF-like domain. The EGF-like domain of TR was highly conserved compared to EGF/NRG family growth factors with the exception of an arginine to histidine substitution at position 39 (Arg --> His 39). Soluble TR stimulated erbB-4 tyrosine phosphorylation in MKN 28 gastric cancer cells, although it was weak compared to neuregulin-induced erbB-4 tyrosine phosphorylation; this suggests that TR might be a ligand for erbB-4- or erbB-4-related receptor tyrosine kinase. TR may have important roles in normal development of middle to late stages of embryos and maintenance of adult central nervous system tissues as high expression of TR mRNAs was observed in these tissues. The modular features suggest multiple roles for TR; these include functioning as a ligand for erbB- receptor, a regulator of TGF-beta-related growth factor signaling by direct interaction through the follistatin modules, and a G-protein-coupled receptor.

Localization of phosphorylated ErbB1-4 and heregulin in colorectal cancer.

BackgroundThe ErbB family consists of four proteins including (EGFR)/ErbB1, ErbB2, ErbB3, and ErbB4, and plays a crucial role in the promotion of multiple tumorigenic processes. In addition to the traditional pathways of EGFR signaling, EGFR translocates to the nucleus and acts as a transcription factor in the proliferation of cancer cells. Heregulin is known as both an ErbB3 and an ErbB4 ligand. This study aimed to investigate the expression of heregulin and its relevant EGFR family members as well as their phosphorylated forms in human colorectal cancer (CRC) tissues and to determine the relationship between their expression and clinicopathological factors including patient prognosis.MethodsWe analyzed the effects of exogenous heregulin on ErbB2, ErbB3 and ErbB4 phosphorylation in Caco-2, DLD-1, and HCT 116 colon cancer cell lines by western blot analysis. We examined 155 surgical resections from colorectomy patients. Cellular localization of ErbB1-4, their phosphorylated forms and heregulin protein was analyzed in CRC surgical resections by immunohistochemical analysis. Immunohistochemical results were compared with clinicopathological factors and patient prognosis.ResultsPhosphorylated ErbB2 (pErbB2) and phosphorylated ErbB3 (pErbB3) were detected in both nuclear and cytosolic fractions of Caco-2 and DLD-1 cells stimulated by exogenous heregulin. Whereas, phosphorylated ErbB4 (pErbB4) was detected only in cytosolic fractions of HCT 116 cells stimulated by exogenous heregulin. Phosphorylated EGFR (pEGFR) immunoreactivity was observed in the cytoplasm and nuclei of cancer cells, whereas the pattern of EGFR staining was membranous and cytoplasmic. Subcellular localization of pErbB2, cytoplasmic, membranous, or nuclear, varied among cases. pErbB3 immunoreactivity was exclusively observed in the nuclei of cancer cells. pErbB4 immunoreactivity was observed in the cell membrane of cancer cells. Statistically, heregulin immunoreactivity correlated with pErbB2 and pErbB4 expression. In multivariate analysis for disease free survival, lymph node status, pErbB3 and pErbB4 expression retained independent prognostic significance. In multivariate analysis for overall survival, lymph node status, pEGFR and pErbB4 retained independent prognostic significance.ConclusionsErbB2 and ErbB3 phosphorylated by heregulin localized in the nucleus of CRC cells. Phosphorylated ErbB1-4 and heregulin contribute to poorer patient prognosis in CRC. This heregulin-ErbB family member autocrine loop may be a candidate for targeted treatment of CRC.