Effects of enteropathogenic bacteria & lactobacilli on chemokine secretion & Toll like receptor gene expression in two human colonic epithelial cell lines

Background & objectives: The intestinal epithelium is part of the innate immune system responding to contact with pathogenic or commensal bacteria. The objective of this study was to compare innate responses of intestinal epithelial cell lines to pathogenic bacteria and to lactobacilli. Methods: Two human intestinal epithelial cell lines, HT29 (enterocyte-like) and T84 (crypt-like), were exposed to pathogenic bacteria representative of non invasive (Vibrio cholerae O1 and O139), adherent (enterohaemorrhagic Escherichia coli, EHEC) or invasive (Salmonella Typhimurium and Shigella flexneri) phenotypes and to non pathogenic Lactobacillus rhamnosus GG or Lactobacillus plantarum. Interleukin-8 (IL-8) was measured in culture supernatant by ELISA, while mRNA from cells was subjected to quantitative reverse transcriptase PCR for several other chemokines (CXCL1, CCL5 and CXCL5) and for Toll-like receptors (TLR) 2, 4, 5 and 9. Results: V. cholerae, S. Typhimurium, S. flexneri and EHEC induced IL-8 secretion from epithelial cells into the medium. Salmonella, Shigella and EHEC, but not V. cholerae, significantly increased mRNA expression of CXCL1. None of the pathogens induced CCL5 or CXCL5. Salmonella and Vibrio significantly increased TLR4 expression, while Vibrio and EHEC decreased TLR5 expression. EHEC also decreased TLR9 expression. Lactobacilli attenuated the IL-8 response of the cell lines to V. cholerae, Salmonella, and EHEC but did not significantly change the IL-8 response to Shigella. Interpretation amp; conclusions: Distinct patterns of epithelial cell chemokine responses were induced by the bacterial pathogens studied and these were modulated by commensal lactobacilli. Alterations in TLR expression by these pathogens are likely to be important in pathogenesis

Evaluation of Hepatoprotective and Antioxidant Effect of Solanum Seaforthianum Leaf Extract against Ccl4 - Induced Li~1

INTRODUCTION: LIVER-INTRODUCTION:The liver has pivotal rule in human metabolism, including detoxification, protein synthesis, and the production of bio-chemicals necessary for digestion. The liver is necessary for survival; there is no currently way to compensate for the absence of liver function long term, however liver cell can regenerate. It produces and secretes bile that stored in gallbladder which used to break down and digest fatty acids in food. 1. It also produces prothrombin and fibrinogen, both are blood-clotting factors. 2. It converts sugar into glycogen, which it stores until the muscle cells need energy and it is secreted into the blood stream as glucose. 3. It synthesizes protein and cholesterol and convert carbohydrates and protein into fats, which is stored for later uses. 4. It also produces blood protein and enzymes needed for digestion and other bodily functions in body. 5. The liver stores critical trace elements such as iron, copper, as well as vitamins A, D, and B12. 6. The liver is responsible for detoxifying of poisonous substances in the body. AIM OF PRESENT STUDY:In recent year there has been a tremendous increase in demand for herbal drugs due to its safety, efficacy and better therapeutic results and also due to its economic pricing as compared to synthetic or allopathic drugs, which have several therapeutic complications. Looking to the scope of herbal drugs and increasing its demand especially in disease of liver, hypertension, diabetes, cancer, arthritis and skin disease etc, it was planned to study Solanum seaforthianum in detail due its effectiveness for liver problems, especially for jaundice. Solanum seaforthianum, selected for study was made on the basis of its, a) Easy availability b) Folklore claiming its therapeutic use as liver protection and for jaundice. c) Degree of research work which is not done. Very less pharmacological studies have been carried out on the Solanum seaforthianum. Hence, we have decided to submit the project on Solanum seaforthianum which includes detailed Pharmacognostical, Preliminary Phytochemical and Pharmacological (Acute oral toxicity study, Hepatoprotective activity with antioxidant effect) studies to provide scientific validation to its folklore claims. SUMMARY AND CONCLUSION: The present study was undertaken to determine the hepatoprotective and antioxidant effect of methanolic extract from leaf of Solanum seaforthianum The Pharmacognostical studies made on the powdered leaves of Solanum seaforthianum like ash values, extractive value, loss on drying gave valuable information. This will help for correct identification of the plant. The preliminary phytochemical investigation showed the presence of, alkaloids, glycosides, flavonoids, and steroids, carbohydrate, fatty acids and proteins and phenolic compounds.in methanol extract. Histopathological studies on isolated liver revealed that methanolic extract of Solanum seaforthianum reversed the liver damage caused by CCl4. The normal pattern of histology of liver was observed. Phytochemical screening revealed the presence of flavonoids. Flavonoids have been documented to possess potent antioxidant and free radical scavenging effect. Example; Solanum seaforthianum contain 2-di-Cglcosylflavones of vicenin and lucenin type, anthocyanin-cynidin-3- glucoside,luteolin-7-glycoside and mono-C-glcosyl flavones – vitexin and orientin Based on the results obtained from the present study, it can be concluded methanolic extract of Solanum seaforthianum is found to be more potent and effective hepatoprotective and antioxidant activity

Analyzing the Information Sharing Practices and Barriers in Supply Chain of Automotive Industries

Information sharing and technology remain one of the key factors of coordination amongst the parties in a supply chain. Supply chain efficiency is highly important as today�s competition is no longer between companies, but between supply chains. Information sharing can increase supply chain efficiency by reducing inventories and smoothing the production. The purpose of this study is to assess the status of the supply chain information sharing practices in Indian automotive industries. This study is focused on identifying the types of information shared, determining the level of information sharing , usage of Information Technology (IT) tools and identifying the barriers of information sharing. Data for this study is collected from the respondents of Original Equipment Manufacturer (OEM), Tier 1, Tier 2 and Tier 3 Indian automobile industries. Frequency analysis is employed to derive the results from the survey

Clostridium leptum group bacteria abundance and diversity in the fecal microbiota of patients with inflammatory bowel disease: a case–control study in India

BACKGROUND: Alterations in the fecal bacterial flora occur in inflammatory bowel disease (IBD). We examined the abundance and diversity of Clostridium leptum group, an important group of carbohydrate-fermenting bacteria, in the feces of patients with IBD and compared them with healthy controls. METHODS: Seventeen healthy controls (HC), 20 patients with Crohn’s disease (CD) and 22 patients with ulcerative colitis (UC) participated in the study. DNA extracted from fecal samples was amplified by PCR targeting 16S rRNA gene sequences specific to C. leptum group. The PCR product was subjected to temporal temperature gradient electrophoresis (TTGE) and the number and position of individual bands were noted and diversity was estimated. The identity of bands at different positions was confirmed by cloning and sequencing. Real time quantitative PCR with Mesa Green, targeted at specific 16S rRNA gene sequences, was used to quantitate C. leptum group and its most prominent constituent, Faecalibacterium prausnitzii. RESULTS: Twenty five different operational taxonomic units (OTUs, equivalent to species) were identified constituting the C. leptum group in these participants. Their sequences were deposited in GenBank [accession numbers GQ465348 to GQ465370]. OTU number was significantly reduced in CD (7.7±3.7, mean±SD) and UC (9.0±3.0) compared to HC (11.9±2.2) (P=0.0005). The Simpson D index of alpha diversity was not significantly different between the three groups. Total numbers of C. leptum group bacteria and F. prausnitzii were reduced in both CD and UC compared to HC (P=0.0036 and P<0.0001 respectively). Disease activity did not influence numbers of C. leptum or F. prausnitzii in patients with CD or UC. CONCLUSION: C. leptum numbers and diversity were significantly reduced in both CD and UC suggesting that alterations noted were not specific to one disease. This could contribute to reduced short chain fatty acid production in IBD

Effect of Vibrio cholerae on Chemokine Gene Expression in HT29 cells and its Modulation by Lactobacillus GG

Epithelial cells participate in the innate immune response to pathogenic bacteria by elaborating chemokines. This study examined the effect of Vibrio cholerae and Lactobacillus rhamnosus GG on inflammatory chemokine gene expression in the HT29 human intestinal epithelial cell line. HT29 cells were exposed to V. cholerae 0139, Lactobacillus or both for 2 h and cultured further thereafter for 4 h. RNA was extracted from the cells and expression of genes for chemokines and related molecules was quantitated by real time PCR using a pathway-focused PCR array. TLR4 was silenced using shRNA and output of interleukin-8 (IL-8) into the media quantitated with and without V. cholerae exposure. NF&#954;B and p38 MAP kinase activation were determined by immunoblotting for I&#954;B&#945; and phosphorylated p38. Vibrio cholerae significantly upregulated gene expression for the neutrophil chemoattractant CXCL chemokines, IL-8, CXCL and CXCL in HT29 cells, while downregulating the expression of macrophage-attracting C-C chemokines. TLR4 silencing did not reduce IL-8 output from HT29 cells in response to V. cholerae. I&#954;B&#945; degradation was noted in the HT29 cells soon after exposure to V. cholerae and this recovered over time after removal of bacteria. p38 MAP kinase activation was not noted. Vibrio cholerae upregulated the expression of neutrophil attractant chemokines, most prominently IL-8, in HT29 cells, but downregulated macrophage-attracting chemokines. Probiotic lactobacilli modulated the IL-8, but not the other chemokine gene changes, in response to V. cholerae

Common NOD2 mutations are absent in patients with Crohn's disease in India

Background: Crohn's disease is being increasingly diagnosed in the Indian subcontinent. Three apparently common mutations in the NOD2 gene are found in up to 30% of sporadic patients with Crohn's disease in western countries. We examined whether such mutations are also found in Indian patients with Crohn's disease. Methods: Venous blood was collected from 82 patients (age range: 7-65 years, 53 men) with Crohn's disease and 149 control subjects; DNA was extracted and subjected to polymerase chain reaction using specific primers. The amplified fragments of size 185, 163 and 151 bp for R702W, G908R and 1007fs, respectively, were digested with MspI, HhaI and ApaI, and the restriction pattern noted after electrophoresis. Results: Twenty-eight patients had ileocolonic disease, 26 ileal disease, 20 colonic disease and 8 had disease limited to proximal small bowel or stomach. None of the 82 patients showed any of the three NOD2 mutations. The control subjects (93 men) had a variety of chronic gastrointestinal disorders (ulcerative colitis 52, irritable bowel syndrome 30, intestinal tuberculosis 20, colon cancer 7, miscellaneous 37). None of the control subjects showed a mutation in any of the three NOD2 mutation analyses. Conclusion: The three NOD2 gene mutations described above are uncommon in Indian patients with Crohn's disease. This study complements information provided by recent studies on NOD2 mutations in Indians

Differential regulation of cholera toxin-inhibited Na-H exchange isoforms by butyrate in rat ileum

Electroneutral Na absorption occurs in the intestine via sodium-hydrogen exchanger (NHE) isoforms NHE2 and NHE3. Bicarbonate and butyrate both stimulate electroneutral Na absorption through NHE. Bicarbonate- but not butyrate-dependent Na absorption is inhibited by cholera toxin (CT). Long-term exposure to butyrate also influences expression of apical membrane proteins in epithelial cells. These studies investigated the effects of short- and long-term in vivo exposure to butyrate on apical membrane NHE and mRNA, protein expression, and activity in rat ileal epithelium that had been exposed to CT. Ileal loops were exposed to CT in vivo for 5 h and apical membrane vesicles were isolated. 22Na uptake was measured by using the inhibitor HOE694 to identify NHE2 and NHE3 activity, and Western blot analyses were performed. CT reduced total NHE activity by 70% in apical membrane vesicles with inhibition of both NHE2 and NHE3. Reduced NHE3 activity and protein expression remained low following removal of CT but increased to control values following incubation of the ileal loop with butyrate for 2 h. In parallel there was a 40% decrease in CT-induced increase in cAMP content. In contrast, NHE2 activity partially increased following removal of CT and was further increased to control levels by butyrate. NHE2 protein expression did not parallel its activity. Neither NHE2 nor NHE3 mRNA content were affected by CT or butyrate. These results indicate that CT has varying effects on the two apical NHE isoforms, inhibiting NHE2 activity without altering its protein expression and reducing both NHE3 activity and protein expression. Butyrate restores both CT-inhibited NHE2 and NHE3 activities to normal levels but via different mechanisms