Biomedical subjects
Publications and source records attributed to Philippe Gérard.
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Intensity of the cholesterol-to-coprostanol conversion in the intestine, as assessed by the coprostanol-to-cholesterol ratio in faeces, was found highly variable among 15 human volunteers, ranging from absent to almost complete cholesterol conversion. The number of coprostanoligenic bacteria in the same faecal samples, as estimated by the most probable number method, was found to be less than 10(6) cellsg-1 of fresh stools in the low-to-inefficient converters and at least 10(8) cellsg-1 of fresh stools in the highest converters, indicating that the population level of cultivable faecal coprostanoligenic bacteria correlated with the intensity of cholesterol-to-coprostanol conversion in the human gut. Microbial communities of the samples were profiled by temporal temperature gradient gel electrophoresis (TTGE) of bacterial 16S rRNA gene amplicons. Dendrogram analysis of the TTGE profiles using the Pearson product moment correlation coefficient and a unweighted pair group method with arithmetic averages (UPGMA) algorithm clearly separated banding patterns from low-to-inefficient and high converters in two different clusters suggesting a relationship between TTGE profiles and coprostanoligenic activity. Principal components analysis further demonstrated that a large subset of bands rather than some individual bands contributed to this clustering.
Ursodeoxycholic acid-producing bacteria are of clinical and industrial interest due to the multiple beneficial effects of this bile acid on human health. This work reports the first isolation of 7-epimerizing bacteria from feces of a healthy volunteer, on the basis of their capacity to epimerize the primary bile acid, chenodeoxycholic acid, to ursodeoxycholic acid. Five isolates were found to be active starting from unconjugated chenodeoxycholic acid and its tauro-conjugated homologue, but none of these strains could epimerize the glyco-conjugated form. Biochemical testing and 16S ribosomal DNA sequencing converged to show that all five isolates were closely related to Clostridium baratii (99% sequence similarity), suggesting that this bacterial species could be responsible at least partially, for this bioconversion in the human gut.
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BACKGROUND/AIMS: The respective influence of folate and vitamin B12 deficiency on MTR activity and transcription, and on DNA methylation is not clearly established. The aim of this study was to assess the respective influence of folate and vitamin B12 deficiency on MTR transcription and activity, and on DNA methylation. METHODS: Sixty-one rats were administered normal diet or diet deficient in choline, methionine, folic acid and vitamin B12. Forty-seven of them underwent total gastrectomy or ileal resection. RESULTS: Low vitamin B12 was observed only in gastrectomized rats. Low folate was observed in rats under deficient diet. Total MTR activity (holo- + apoenzyme) was lowered only with vitamin B12 level <200 pmol/l (p=0.0002), while the ratios of total vs. holo-MTR activity and of transcripts MTR vs. GAPDH (RT-PCR) were unchanged. Vitamin B12 was the single determinant of low MTR (lower quartile, odds ratio=15.75, p=0.0017). Low MTR and low vitamin B12 were the two determinants of DNA hypomethylation (lower quartile) (odds ratio=17.07, p=0.0006, and odds ratio=7.31, p=0.006, respectively). CONCLUSION: Vitamin B12 affects MTR expression by a non-transcriptional mechanism different from a protective effect on MTR proteolysis. It is also a strong determinant of DNA hypomethylation.
FtsW and RodA are homologous integral membrane proteins involved in bacterial cell division and cell growth, respectively. Both proteins from Streptococcus pneumoniae were overexpressed in Escherichia coli as N-terminal His-tagged fusions. Their membrane addressing in E. coli was demonstrated by cell fractionation and was confirmed for FtsW by immunolocalization. Recombinant FtsW and RodA were solubilized from membranes using 3-(laurylamido)-N,N'-dimethylaminopropylamine oxide (LAPAO). The detergent was exchanged to polyoxyethylene 8 lauryl ether (C12E8) during one-step purification procedure by Co(2+)-affinity chromatography. This procedure yielded 50-150 microg protein per litre of culture. Both proteins are likely to be folded as they are resistant to trypsin digestion and could be incorporated into reconstituted lipid vesicles.
Deficiencies of the major dietary sources of methyl groups, methionine and choline, lead to the formation of liver cancer in rodents. The most widely investigated hypothesis has been that dietary methyl insufficiency results in abnormal DNA methylation. Vitamin B12 and folate also play important roles in DNA methylation since these two coenzymes are required for the synthesis of methionine and S-adenosyl methionine, the common methyl donor required for the maintenance of methylation patterns in DNA. The aim of this study was to review the effects of methyl-deficient diets on DNA methylation and liver carcinogenesis in rats, and to evaluate the role of vitamin B12 status in defining carcinogenicity of a methyl-deficient diet. Several studies have shown that a methyl-deficient diet influences global DNA methylation. Evidence from in vivo studies has not clearly established a link between vitamin B12 and DNA methylation. We reported that vitamin B12 and low methionine synthase activity were the two determinants of DNA hypomethylation. Choline- or choline/methionine-deficient diets have been shown to cause hepatocellular carcinoma in 20-50% of animals after 12-24 months. In contrast, the effect of vitamin B12 withdrawal, in addition to choline, methionine and folate, induced hepatocellular carcinoma in less than 5% of rats.
The topology of FtsW from Streptococcus pneumoniae, an essential membrane protein involved in bacterial cell division, was predicted by computational methods and probed by the alkaline phosphatase fusion and cysteine accessibility techniques. Consistent results were obtained for the seven N-terminal membrane-spanning segments. However, the results from alkaline phosphatase fusions did not confirm the hydropathy analysis of the C-terminal part of FtsW, whereas the accessibility of introduced cysteine residues was in agreement with the theoretical prediction. Based on the combined results, we propose the first topological model of FtsW, featuring 10 membrane-spanning segments, a large extracytoplasmic loop, and both N and C termini located in the cytoplasm.
Hyperhomocysteinemia has been associated with several pregnancy complications. We have investigated the variation of plasma total homocysteine (tHcys) during the 2 last trimesters of normal pregnancy and related it to blood vitamin B12 and folate and to the excretion of the degraded intrinsic factor receptor (IFCR) in urine, in a follow-up study of 15 cases. A significant rise in tHcys was observed between the beginning of the second trimester and the third trimester with respective values (median) 6.1, 5.8 and 6.7 micromol/l (p = 0.038). The tHcys/albumin ratio also increased significantly, while no correlation was found between albumin and folate blood concentration. In contrast, a significant decrease in vitamin B12 was observed (279, 225 and 199 pmol/l, between the 4th and 6th, and the 6th and 9th month, respectively (p = 0.017-0.002)). A significant negative correlation was found between tHcys between the 4th and 9th month of pregnancy and the ratio of vitamin B12 between the 4th and 9th month of pregnancy (r = 0.55, p = 0.037). The urine excretion of IFCR was increased and was not related to vitamin B12 and tHcys. In conclusion, we have observed a rise in tHcys between the beginning of the second trimester and the third trimester of pregnancy which was related to the decreased blood level of vitamin B12. Subclinical deficiency of vitamin B12 should be further investigated in pregnant women who remain on inadequate diet.