[Ethical issues of vaccination against Sars-CoV-2]. / Enjeux éthiques de la vaccination contre le Sars-CoV-2.

As a tool for public health, the vaccination policy is based on the analysis of benefits and risks. Thus, the National Consultative Ethics Committee has been at the heart of the orientations taken in terms of the deployment of the vaccination against severe acute respiratory syndrome coronavirus 2, by contributing its reflections on the associated ethical issues.

The Contribution of Intestinal Gluconeogenesis to Glucose Homeostasis Is Low in 2-Day-Old Pigs.

Background: Active gluconeogenesis is essential to maintain blood glucose concentrations in neonatal piglets because of the high glucose requirements after birth. In several adult mammals, the liver, kidney, and possibly the gut may exhibit gluconeogenesis during fasting and insulinopenic conditions. During the postnatal period, the intestine expresses all of the gluconeogenic enzymes, suggesting the potential for gluconeogenesis. Galactose in milk is a potential gluconeogenic precursor for newborns.Objective: Our aim was to quantify the rate of intestinal glucose production from galactose in piglets compared with the overall rate of glucose production.Methods: A single bolus of [U-14C]-galactose was injected into 2-d-old piglets (females and males; mean ± SEM weight: 1.64 ± 0.07 kg) through a gastric catheter. Galactosemia, glycemia, and glucose turnover rate (assessed by monitoring d-[6-3H]-glucose) were monitored. Intestinal glucose production from [U-14C]-galactose was calculated from [U-14C]-glucose appearance in the blood and isotopic dilution. Galactose metabolism was also investigated in vitro in enterocytes isolated from 2-d-old piglets that were incubated with increasing concentrations of galactose.Results: In piglet enterocytes, galactose metabolism was active (mean ± SEM maximum rate of reaction: 2.26 ± 0.45 nmol · min-1 · 106 cells-1) and predominantly oriented toward lactate and pyruvate production (74.0% ± 14.5%) rather than glucose production (26.0% ± 14.5%). In conscious piglets, gastric galactose administration led to an increase in arterial galactosemia (from 0 to 1.0 ± 0.8 mmol/L) and glycemia (35% ± 12%). The initial increase in arterial glycemia after galactose administration was linked to an increase in glucose production rate (33% ± 15%) rather than to a decrease in glucose utilization rate (3% ± 6%). The contribution of intestinal glucose production from galactose was <10% of total glucose production in 2-d-old piglets.Conclusion: Our results indicate that there is a low contribution to glucose homeostasis from intestinal gluconeogenesis in 2-d-old piglets.

Chapitre 2. La loi de bioéthique : un enjeu de démocratie sanitaire.

The law of bioethics defines the judicial framework for governing medical practices and research involving the human body and the embryo. Does any scientific or technologic innovation warrant a modification of the law? To respond to this question, a preliminary reflection is necessary so as to define new equilibria which respect ethical principles, the promotion of solidarity with respect for the autonomy of each person.The CCNE is charged with organizing public debate, in the form of a general assembly of bioethics, with the support of regional spaces of ethical reflection. This extensive experience took place from January to June 2018 and lead to several lessons, notably the strong expression of a need for information and a critical vision of the notion of medical progress.Then, the parliamentary instruction and hearings that accompanied it, from 24 July 2019 to 2 August 2021, contributed developments to the initial bill. Thus, the periodic review of the law of bioethics is now based on citizen participation in the construction of legislative arbitration, demonstrating the French way of social participation.

Chapitre 2. La loi de bioéthique : un enjeu de démocratie sanitaire.

The law of bioethics defines the judicial framework for governing medical practices and research involving the human body and the embryo. Does any scientific or technologic innovation warrant a modification of the law? To respond to this question, a preliminary reflection is necessary so as to define new equilibria which respect ethical principles, the promotion of solidarity with respect for the autonomy of each person.The CCNE is charged with organizing public debate, in the form of a general assembly of bioethics, with the support of regional spaces of ethical reflection. This extensive experience took place from January to June 2018 and lead to several lessons, notably the strong expression of a need for information and a critical vision of the notion of medical progress.Then, the parliamentary instruction and hearings that accompanied it, from 24 July 2019 to 2 August 2021, contributed developments to the initial bill. Thus, the periodic review of the law of bioethics is now based on citizen participation in the construction of legislative arbitration, demonstrating the French way of social participation.

Oleate metabolism in pig enterocytes is characterized by an increased oxidation rate in the presence of a high esterification rate within two days after birth.

Oleate (OLE) is the principle fatty acid (FA) in mammalian colostrum, but its role in the energy supply in enterocytes after birth remains unknown. We investigated the metabolic fate of OLE in pig enterocytes at birth (d0) and after 2 d of suckling (d2). Cellular TG and phospholipids (PL) and FA composition were analyzed. Metabolic end-products of [1-¹4C]OLE were measured in enterocyte incubations. We characterized intestinal carnitine palmitoyltransferase 1 (CPT1), the key enzyme of mitochondrial FA oxidation. The TG content was 6.6-fold higher in enterocytes from pigs on d 2 than in those obtained on d 0, whereas the PL content did not differ. The level of OLE in TG and PL increased from 15 and 11% of total FA, respectively, in enterocytes from newborn piglets to 30 and 17%, respectively, in those from d2 pigs. The capacity for OLE utilization was 2.8-fold greater in d2 than in d0 pig enterocytes. The oxidation and esterification rates were enhanced in enterocytes from piglets on d 2 compared to those obtained on d 0, by 4- and 2.6-fold, respectively. The predominant OLE fate was the esterification pathway, representing >85% of OLE metabolized in both groups. The limited OLE oxidation observed at d 2 may result from the presence of a highly malonyl-CoA-sensitive CPT1A, because the half maximal inhibitory concentration for malonyl-CoA was 162 ± 25 nmol/L. This study highlighted the high esterification capacity for OLE in the newborn pig intestine, which may preserve this major colostrum FA for delivery to other tissues.

Microbiota matures colonic epithelium through a coordinated induction of cell cycle-related proteins in gnotobiotic rat.

Previous studies have suggested that intestinal microbiota modulates colonic epithelium renewal. The objective of our work was to study the effects of microbiota on colonic epithelium structure and cell cycle-related proteins by using gnotobiotic rats. Colonic crypts and amount of cell cycle-related proteins were compared between germ-free (GF), conventional (CV), and conventionalized rats by histochemistry and Western blot. Ki67 and proliferating cell nuclear antigen (PCNA) were used as surrogates for proliferative cells; p21(cip1) and p27(kip1) were markers of cell cycle arrest; anti- and proapoptotic proteins, Bcl2 and Bax, respectively, were also studied. We observed 40% increase of the crypt proliferative area 2 days after inoculation of GF rats with a complex microbiota. This recruitment of proliferative cells may account for the 30% increase of crypt depth observed between CV and GF rats. The hyperproliferative boost induced by microbiota was compensated by a fourfold increase of p21(cip1) and p27(kip1) involved in cell cycle arrest and a 30% drop of antiapoptotic Bcl2 protein while Bax was unchanged. Inductions of p21(cip1), p27(kip1), and PCNA protein were not paralleled by an increase of the corresponding mRNA. We also showed that p21(cip1) induction by microbiota was partially restored by Bacteroides thetaiotaomicron, Ruminococcus gnavus, and Clostridium paraputrificum. Colonization of the colon by a complex microbiota increases the crypt depth of colon epithelium. This event takes place in conjunction with a multistep process: a hyperproliferative boost accompanied by compensatory events as induction of p21(cip1) and p27(kip1) and decrease of Bcl2.

Morphological adaptation with preserved proliferation/transporter content in the colon of patients with short bowel syndrome.

In short bowel syndrome (SBS), although a remaining colon improves patient outcome, there is no direct evidence of a mucosal colonic adaptation in humans. This prospective study evaluates morphology, proliferation status, and transporter expression level in the epithelium of the remaining colon of adult patients compared with controls. The targeted transporters were Na+/H+ exchangers (NHE2 and 3) and oligopeptide transporter (PepT1). Twelve adult patients with a jejuno-colonic anastomosis were studied at least 2 yr after the last surgery and compared with 11 healthy controls. The depth of crypts and number of epithelial cells per crypt were quantified. The proliferating and apoptotic cell contents were evaluated by revealing Ki67, PCNA, and caspase-3. NHE2, NHE3, PepT1 mRNAs, and PepT1 protein were quantified by quantitative RT-PCR and Western blot, respectively. In patients with SBS compared with controls, 1) hyperphagia and severe malabsorption were documented, 2) crypt depth and number of cells per crypt were 35% and 22% higher, respectively (P < 0.005), whereas the proliferation and apoptotic levels per crypt were unchanged, and 3) NHE2 mRNA was unmodified; NHE3 mRNA was downregulated near the anastomosis and unmodified distally, and PepT1 mRNA and protein were unmodified. We concluded that, in hyperphagic patients with SBS with severe malabsorption, adaptive colonic changes include an increased absorptive surface with an unchanged proliferative/apoptotic ratio and well-preserved absorptive NHE2, NHE3, and PepT1 transporters. This is the first study showing a controlled nonpharmacological hyperplasia in the colon of patients with SBS.

Diallyl disulfide increases CDKN1A promoter-associated histone acetylation in human colon tumor cell lines.

Diallyl disulfide (DADS) is an organosulfur compound from garlic, which inhibits colon tumor cell proliferation. In a previous study, we have shown that in Caco-2 and HT-29 cells DADS (200 microM) increases global histone acetylation, CDKN1A mRNA, and p21(waf1) protein levels and induces G2/M cell cycle arrest. These results suggested that DADS could inhibit cell proliferation through at least in part a transcriptional activation of CDKN1A expression involving histone acetylation. In this study, using chromatin immunoprecipitation assays, we demonstrate that in Caco-2 and HT-29 cells histone H4 and/or H3 acetylation is increased within CDKN1A promoter after 3 and 6 h treatments with DADS. These results strongly suggest that histone acetylation, a molecular mechanism implicated in the regulation of gene expression, could account for the induction of CDKN1A expression and the antiproliferating effects of DADS in colon tumor cells.

Repetitive treatments of colon HT-29 cells with diallyl disulfide induce a prolonged hyperacetylation of histone H3 K14.

Diallyl disulfide (DADS) is a sulfur compound derived from garlic. Several studies carried out in rodents have revealed protective effects of DADS against colon carcinogenesis. The antipromoting effects of DADS may be partly related to its ability to inhibit tumoral cell proliferation. In a previous study, we have shown that in two human colon tumor cell lines (HT-29 and Caco-2) seeded at a low density (0.2 x 10(6) cells/100-mm petri dish), DADS antiproliferative effects were associated with a transient increase of histone H3 K14 acetylation. Moreover, DADS could inhibit nuclear histone deacetylase activity. Therefore, in the present study, we examined the possible effects of different experimental conditions (HT-29 cells at high density, repetitive treatments with DADS) on the pattern of DADS-induced histone hyperacetylation. Using HT-29 cells seeded at a higher density (5 x 10(6) cells/100-mm petri dish), we found that DADS induced histone H3 K14 hyperacetylation rapidly (3 h). When administrated as single treatments, the DADS effect on histone H3 K14 remained transient. In contrast, repetitive treatment with DADS resulted in a prolonged hyperacetylation of histone H3 K14. Whatever the cell culture conditions were, DADS had no effect on histone H4 acetylation. Thus, in vitro, the cell density and pattern of DADS treatment influenced the HT-29 nuclear response to DADS. DADS belongs to food-borne molecules that may play a role in chromatin remodeling and contribute to the nutritional modulation of gene expression.

Enterocyte metabolism during early adaptation after extensive intestinal resection in a rat model.

OBJECTIVE: A better knowledge of intestinal adaptation after resection is required to improve the nutritional support that is given to patients. The aim of this study was to understand the metabolic changes underlying early adaptation after massive intestinal resection. METHODS: Rats were assigned to either 80% intestinal resection or transection. All animals received the same intragastric nutrition. On day 8, plasma glutamine turnover was measured. Substrate use was determined on isolated enterocytes that were incubated in the presence of D-[U-(14)C] glucose (2 mmol/L), L-[U-(14)C] glutamine (2 mmol/L), L-[U-(14)C] arginine (1 mmol/L), or L-[1-(14)C] ornithine (1 mmol/L). RESULTS: Plasma glutamine turnover was similar in both groups. The rate of enterocyte glutamine use was significantly increased in the resection group, although the maximal glutaminase activity was unchanged. Glutathione generation was enhanced 3-fold in remnant intestine as compared with transected intestine (P <.05). L-ornithine decarboxylation was increased markedly in resected animals (P <.05), without any detectable change of maximal ornithine decarboxylase activity. CONCLUSION: The early phase of intestinal adaptation after resection induces changes in enterocyte glutamine and ornithine metabolism that may be related, in part, to increased de novo polyamine synthesis. This observation suggests that a supplementation of artificial nutrition by nutrients that lead to the generation of trophic agents may be of potential interest.

Drastic changes in fecal and mucosa-associated microbiota in adult patients with short bowel syndrome.

Short bowel syndrome (SBS) is observed in Humans after a large resection of gut. Since the remnant colon and its associated microbiota play a major role in the outcome of patients with SBS, we studied the overall qualitative and quantitative microbiota composition of SBS adult patients compared to controls. The population was composed of 11 SBS type II patients (with a jejuno-colonic anastomosis) and 8 controls without intestinal pathology. SBS patients had 38 +/- 30 cm remnant small bowel length and 66 +/- 19% of residual colon. The repartition of proteins, lipids, carbohydrates and fibres was expressed as % of total oral intake in patients and controls. The microbiota was profiled from stool and biopsy samples with temporal temperature gradient gel electrophoresis and quantitative PCR. We show here that microbiota of SBS patients is unbalanced with a high prevalence of Lactobacillus along with a sub-dominant presence and poor diversity of Clostridium leptum, Clostridium coccoides and Bacteroidetes. In addition, Lactobacillus mucosae was detected within the fecal and mucosa-associated microbiota of SBS patients, whereas it remained undetectable in controls. Thus, in SBS the microbial composition was deeply altered in fecal and mucosal samples, with a shift between dominant and sub-dominant microbial groups and the prevalence of L. mucosae.

Apple proanthocyanidins do not reduce the induction of preneoplastic lesions in the colon of rats associated with human microbiota.

Since the gut microbiota metabolizes various dietary constituents unabsorbed by the small intestine and modulates colon function, it plays an essential role in colon carcinogenesis. First, we have developed a model of human microbiota-associated rats (HMA), fed a human-type diet and injected with 1-2,dimethylhydrazine (DMH). We observed that the number and size of DMH-induced aberrant crypt foci (ACF) were significantly higher in HMA rats than in germ-free or conventional rats. Second, we used this model to assess the protective effect of an apple proanthocyanidin-rich extract (APE) on colon carcinogenesis. In this model, ACF number and multiplicity were not reduced by APE at 0.001% and 0.01% in drinking water. They were higher with APE 0.1% than with APE 0.01%. Therefore, the cross-talk between human microbiota and the colon epithelium should be taken into account in carcinogenesis models. Moreover, attention should be paid prior to using proanthocyanidin extracts as dietary supplements for humans.

In vivo treatment by diallyl disulfide increases histone acetylation in rat colonocytes.

Diallyl disulfide (DADS) is an organosulfur compound from garlic which exhibits various anticarcinogenic properties including inhibition of tumor cell proliferation. DADS antiproliferative effects were previously associated with an increase in histone acetylation in two human tumor colon cell lines, suggesting that DADS-induced histone hyperacetylation could be one of the mechanisms involved in its protective properties on colon carcinogenesis. The effects of DADS on histone H4 and H3 acetylation levels were investigated in vivo in colonocytes isolated from non-tumoral rat. Administrated by intracaecal perfusion or gavage, DADS increases histone H4 and H3 acetylation in colonocytes. Moreover, data generated using cDNA expression arrays suggest that DADS could modulate the expression of a subset of genes. These results suggest the involvement of histone acetylation in modulation of gene expression by DADS in normal rat colonocytes, which might play a role in its biological effects as well as in its anticarcinogenic properties in vivo.

Different activation patterns of rat xenobiotic metabolism genes by two constituents of garlic.

Diallyl sulfide (DAS) and diallyl disulfide (DADS) are natural components that could account for the anticarcinogenic properties of garlic, at least in part, through the activation of xenobiotic detoxifying metabolism. The aim of this work was to describe the effect of DAS and DADS on xenobiotic-related gene expressions and to study molecular mechanisms relaying DAS effect. We describe the different effects of DAS and DADS on hepatic CYP2B1/2, CYP3A and epoxide hydrolase (EpH) mRNAs in rats, in terms of activation profile, doses and kinetics. The activation profile varied with the mode of chemical administration, i.e. gastric infusion or intraperitoneal (i.p.) injection. Using gastric infusion, DAS and DADS proved different efficiencies at enhancing the mRNA level of the three drug-metabolizing enzymes. After an i.p. administration, we observed a specific activation of CYP2B1/2 gene by DAS. The DAS-mediated CYP2B1/2 activation occurred at transcriptional level and through an okadaic acid-sensitive pathway. In rat livers, a short sequence (NR1) derived from the CYP2B1/2 promoter was stimulated by DAS and we observed a nuclear accumulation of a DNA-protein complex binding NR1. Because constitutively activated receptor (CAR) is a major transcription factor driving the xenobiotic-induced stimulation of CYP2B1/2 through NR1, the role of CAR as a preferential mediator of DAS effect is discussed.

Changes induced in colonocytes by extensive intestinal resection in rats.

After massive intestinal resection, physiological compensatory events occur in the remnant small bowel and in the colon. The aim of our work was to study the propensity of the colon to evolve after a massive small bowel resection in rats. The resected group, where 80% of the small bowel length was removed, was compared with sham-operated rats (transected). During the 7 postoperative days, rats were fed orally or they received an elemental nutrition through a gastric catheter. PepT1 and NHE3 mRNAs encoding apical membrane transporters were not modified in the present experiment. However, two unexpected genes (I-FABP and UroR) were up-regulated in the colon following intestinal resection. These modifications occurred without an imbalance of cell cycle protein content and in a context of low short-chain fatty acid production.

[Dietary fatty acids and cancer: potential cellular and molecular mechanisms]. / Acides gras alimentaires et cancers: mécanismes d'action cellulaire et moléculaire.

Experimental evidence indicates that n-3 fatty acids, especially the long-chain polyunsaturated fatty acids, unlike n-6 fatty acids could prevent cancer development. This survey shows that fatty acids could act through several mechanisms including the production of reactive oxygen species, the modulation of gene expression and signal transduction pathways, or the eicosanoid biosynthesis. Human genetics has underlined several polymorphisms in genes identified as possible targets of fatty acids which suggests that the link between nutritional intake and cancer prevention, especially the eventual anti-carcinogenic effects of n-3 fatty acids, depends on the genetic background. Further studies are needed to evaluate the effects of fatty acids on angiogenesis which represents a marker of a poor prognosis in cancer. Finally, the use of genomic technologies combined with nutritional strategies could provide a more understanding of the effects of n-3 fatty acid intake on cancer prevention.

Diallyl disulfide increases rat h-ferritin, L-ferritin and transferrin receptor genes in vitro in hepatic cells and in vivo in liver.

Of the oil-soluble organosulfur compounds derived from garlic, diallyl disulfide (DADS) is one of the most abundant. We examined the effect of DADS on gene expression in rat liver. By suppressive subtractive hybridization, we identified the heavy (H)-ferritin gene as a DADS-stimulated gene in the rat liver epithelial (REL) cells. DADS stimulation of H- and L (light)-ferritin mRNA was analyzed in REL cells and in rat liver. Incubation of the REL cells in 10 micro mol/L DADS for 4 h increased H-ferritin 1.9 +/- 0.2-fold, n = 3) and light(L)-ferritin mRNA 1.5 +/- 0.2-fold, n = 3). Stimulation did not occur in the presence of an inhibitor of transcription, actinomycin D. Stimulation of ferritin at the RNA and protein levels was also found in rats administered a DADS-enriched oil solution intragastrically. There was a 3 +/- 1.1-fold increase in H- and 3 +/- 0.14-fold increase for L-ferritin mRNA 24 h after the end of the infusion in the presence of DADS, (n = 3). The expression of the transferrin receptor, an iron transporter, was also enhanced by DADS in rat liver. In conclusion, our data suggest that DADS could modify iron homeostasis through the modulation of ferritin and transferrin receptor gene expression.

Diallyl disulfide (DADS) increases histone acetylation and p21(waf1/cip1) expression in human colon tumor cell lines.

Diallyl disulfide (DADS) is a naturally occurring organosulfur compound, from garlic, which exerts pleiotropic biological effects. In rodents, DADS inhibits colon chemically induced carcinogenesis. DADS anti-promoting effect may partly result from its ability to inhibit tumoral cell proliferation in vivo and in vitro. As far as DADS may modulate the expression of a subset of genes, we investigated DADS effect on histone acetylation, in two human colon tumor cell lines. Our study demonstrates that in Caco-2 and HT-29 cells treated for 6 h, 200 microM DADS increases histone H3 acetylation (x2 and x1.4, respectively). In Caco-2 cells, we also observed histone H4 hyperacetylation, preferentially at the lysine residues 12 and 16. We explored the effects of DADS and one of its metabolites, allyl mercaptan (AM), on histone deacetylase (HDAC) activity: using nuclear extracts of Caco-2 cells, 200 microM DADS decreased HDAC activity by 29% and AM at the same concentration was more efficient (92% inhibition). We also observed that DADS induced an increase in p21(waf1/cip1) expression, at mRNA and protein levels, in both cell lines. This effect was associated with an accumulation of cells in the G2 phase of the cell cycle. Our results suggest that in Caco-2 and HT-29 cells, DADS could inhibit cell proliferation through the inhibition of HDAC activity, histone hyperacetylation and increase in p21(waf1/cip1) expression. The present study provides evidence for cellular and molecular responses triggered by DADS that could be linked to its effect on histone acetylation and play a role in its protective properties on colon carcinogenesis.