Development of beta-cell mass in fetuses of rats deprived of protein and/or energy in last trimester of pregnancy

Fetal malnutrition is now proposed as a risk factor of later obesity and type II diabetes. We previously analyzed the long-term impact of reduced protein and/or energy intake strictly limited to the last week of pregnancy in Wistar rats. Three protocols of gestational malnutrition were used: 1) low-protein isocaloric diet (5 instead of 15%) with pair feeding to the mothers receiving the control diet, 2) restricted diet (50% of control diet), and 3) low protein-restricted diet (50% of low-protein diet). Only isolated protein restriction induced a long-term beta-cell mass decrease. In the present study, we used the same protocols of food restriction to analyze their short-term impact (on day 21.5 of pregnancy) on beta-cell mass development. A 50% beta-cell mass decrease was present in the three restricted groups, but low-protein diet, either associated or not to energy restriction, increased fetal beta-cell insulin content. Among all the parameters analyzed to further explain our results, we found that the fetal plasma level of taurine was lowered by low-protein diet and was the main predictor of the fetal plasma insulin level (r = 0.63, P < 0.01). In conclusion, rat fetuses exposed to protein and/or energy restriction during the third part of pregnancy have a similar dramatic decrease in beta-cell mass, and their ability to recover beta-cell mass development retardation depends on the type of malnutrition used. Moreover, our results support the hypothesis that taurine might play an important role in fetal beta-cell mass function.     

Active peptidic mimics of the second intracellular loop of the V(1A) vasopressin receptor are structurally related to the second intracellular rhodopsin loop: a combined 1H NMR and biochemical study

Vasopressin (VP) receptors belong to the widespread G protein-coupled receptor family. The crucial role of VP receptor intracellular loops in the coupling with the heterotrimeric G proteins was previously demonstrated by construction of a vasopressin receptor chimera. Yet, no fine structural data are available concerning the receptor molecular determinants involved in their interactions with G proteins. In this study, we synthesized both a linear and a cyclic form of the second intracellular loop (i2) of the human V(1a) vasopressin receptor isoform that is important for the interaction between the alphaq/alpha11 G protein and the receptor. These two peptides are biologically active. They specifically inhibit vasopressin binding to the V(1a) receptor, suggesting that the corresponding endogenous peptides contribute to the structure of the vasopressin binding site via intra- or intermolecular interactions with the core of the V(1a) receptor. The i2 peptide structures were determined by (1)H NMR. Both exhibit a helix and helical elements in their N- and C-terminal parts, respectively, separated by a turn imposed by a proline residue. More interestingly, the central Pro-Leu motif conserved in many GPCRs and thought to be important for coupling to G proteins can adopt different conformations. The "U" shape structure of the i2 loop is compatible with its anchoring to transmembrane domains III and IV and is very similar to the shape of bovine rhodopsin i2. Altogether, these data contribute to a better understanding of the structure of a not yet crystallized GPCR using the mimetic peptide approach.     

Replication of the chicken beta-globin locus: early-firing origins at the 5' HS4 insulator and the rho- and betaA-globin genes show opposite epigenetic modifications

Chromatin structure is believed to exert a strong effect on replication origin function. We have studied the replication of the chicken beta-globin locus, whose chromatin structure has been extensively characterized. This locus is delimited by hypersensitive sites (HSs) that mark the position of insulator elements. A stretch of condensed chromatin and another HS separate the beta-globin domain from an adjacent folate receptor (FR) gene. We demonstrate here that in erythroid cells that express the FR but not the globin genes, replication initiates at four sites within the beta-globin domain, one at the 5' HS4 insulator and the other three near the rho- and beta(A)-globin genes. Three origins consist of G+C-rich sequences enriched in CpG dinucleotides. The fourth origin is A+T rich. Together with previous work, these data reveal that the insulator origin has unmethylated CpGs, hyperacetylated histones H3 and H4, and lysine 4-methylated histone H3. In contrast, opposite modifications are observed at the other G+C-rich origins. We also show that the whole region, including the stretch of condensed chromatin, replicates early in S phase in these cells. Therefore, different early-firing origins within the same locus may have opposite patterns of epigenetic modifications. The role of insulator elements in DNA replication is discussed.     

Regulation of MHC class II expression and antigen processing in murine and human mesenchymal stromal cells by IFN-gamma, TGF-beta, and cell density

Mesenchymal stromal cells (MSC) possess immunosuppressive properties, yet when treated with IFN-gamma they acquire APC functions. To gain insight into MSC immune plasticity, we explored signaling pathways induced by IFN-gamma required for MHC class II (MHC II)-dependent Ag presentation. IFN-gamma-induced MHC II expression in mouse MSC was enhanced by high cell density or serum deprivation and suppressed by TGF-beta. This process was regulated by the activity of the type IV CIITA promoter independently of STAT1 activation and the induction of the IFN regulatory factor 1-dependent B7H1/PD-L1 encoding gene. The absence of direct correlation with the cell cycle suggested that cellular connectivity modulates IFN-gamma responsiveness for MHC II expression in mouse MSC. TGF-beta signaling in mouse MSC involved ALK5 and ALK1 TGF-beta RI, leading to the phosphorylation of Smad2/Smad3 and Smad1/Smad5/Smad8. An opposite effect was observed in human MSC where IFN-gamma-induced MHC II expression occurred at the highest levels in low-density cultures; however, TGF-beta reduced IFN-gamma-induced MHC II expression and its signaling was similar as in mouse MSC. This suggests that the IFN-gamma-induced APC features of MSC can be modulated by TGF-beta, serum factors, and cell density in vitro, although not in the same way in mouse and human MSC, via their convergent effects on CIITA expression.     

Apo alpha-lactalbumin and lysozyme are colocalized in their subsequently formed spherical supramolecular assembly

We have reported previously that the calcium-depleted form of bovine alpha-lactalbumin (apo alpha-LA) interacts with hen egg-white lysozyme (LYS) to form spherical supramolecular structures. These supramolecular structures contain an equimolar ratio of the two proteins. We further explore here the organization of these structures. The spherical morphology and size of the assembled LYS/apo alpha-LA supramolecular structures were demonstrated using confocal scanning laser microscopy and scanning electron microscopy. From confocal scanning laser microscopy experiments with labelled proteins, it was found that LYS and apo alpha-LA were perfectly colocalized and homogeneously distributed throughout the entire three-dimensional structure of the microspheres formed. The spatial colocalization of the two proteins was also confirmed by the occurrence of a fluorescence resonance energy transfer phenomenon between labelled apo alpha-LA and labelled LYS. Polarized light microscopy analysis revealed that the microspheres formed differ from spherulites, a higher order semicrystalline structure. As the molecular mechanism initiating the formation of these microspheres is still unknown, we discuss the potential involvement of a LYS/apo alpha-LA heterodimer as a starting block for such a supramolecular assembly.     

Easy DNA extraction method and optimisation of PCR-Temporal Temperature Gel Electrophoresis to identify the predominant high and low GC-content bacteria from dairy products

Molecular fingerprinting of bacterial ecosystems has recently increased in food microbiology. The aim of this work was to develop a rapid and easy method to extract DNA from various cheeses, and to optimize the separation of low and high GC-content bacteria by PCR-Temporal Temperature Gel Electrophoresis (PCR-TTGE). Seventy six strains belonging to 50 of the most frequently encountered bacterial species in dairy products were used to construct a database. Specific PCR-TTGE ladders containing 17 species forming a regular scale were created. Amplicons of these species were sequenced and the GC-content plotted against the migration distance: the correlation coefficients obtained were r(2)=0.97 and r(2)=0.99, respectively for high and low GC-contents. The extraction method developed did not use any harmful solvent such as phenol/chloroform. The concentrations of DNA extracted from hard cooked and pressed cheeses, quantified by picogreen molecular probes, were between 0.7 and 6 microg/g for core samples and 8 to 30 microg/g for rind samples. Experimental as well as commercial dairy products were analysed using the developed method and the reproducibility of the profiles was 89%. The method appears to be particularly efficient in the characterization of the ecosystem of cheese rinds.     

Health Behaviors: Is There Any Distinction for Teachers? A Cross-Sectional Nationwide Study

Background

Health behaviors, as important modifiable determinants of health, are consistently targeted by prevention messages. Teachers, as educators and role models, may play a key-role in bringing such messages to children and adolescents. It is not clear which areas of prevention could be improved in collaboration with teachers to promote healthy behaviors at the population level through health education in schools.

Methods

to evaluate teacher’s health awareness, we compared their health/risk behaviors to those of non-teachers, taking into account demographic and socioeconomic factors that could confound crude differences. We used data from the 2010 Health Barometer, a cross-sectional nationally-representative French survey conducted by telephone among 27,653 persons aged 15–85. Adjusting sequentially for potential confounders, we compared six indicators of lifestyle and risky conducts (at-risk drinking, current smoking, cannabis use, gambling, corpulence, sleep duration) between teachers (n = 725) and two comparison groups: other occupations (n = 12,483) on the one hand, and other intermediate and managerial/professional occupations (n = 6,026) on the other.

Results

In the fully-adjusted models, teachers were less likely than other occupations to smoke, to have used cannabis in the last 12 months, to gamble regularly and to be overweight or obese. When restricting the comparison group to other occupations belonging to the same socio-professional category, differences were attenuated, but remain highly significant for tobacco, cannabis and gambling. No significant differences were observed between teachers and non-teachers regarding alcohol use and sleep duration, once important confounders had been adjusted for.

Conclusions

Our results suggest that teachers behave on the whole more healthily than other adults with a similar demographic and socioeconomic profile. The absence of a teacher distinction toward at-risk drinking needs to be examined in more detail.     

Hesperetin stimulates differentiation of primary rat osteoblasts involving the BMP signalling pathway

Hesperidin found in citrus fruits has been reported to be a promising bioactive compound for maintaining an optimal bone status in ovariectomized rodent models. In this study, we examined the capacity of hesperetin (Hp) to affect the proliferation, differentiation and mineralization of rodent primary osteoblasts. Then, the impact of Hp on signalling pathways known to be implicated in bone formation was explored. We exposed osteoblasts to physiological concentrations of 1 μM Hp (Hp1) and 10 μM Hp (Hp10). Neither proliferation nor mineralization was affected by Hp at either dose during 19 days of exposure. Hp at both doses enhanced differentiation by significantly increasing alkaline phosphatase (ALP) activity from Day 14 of exposure (Day 19: Hp1: +9%, Hp10: +14.8% vs. control; P<.05). However, Hp did not induce an obvious formation of calcium nodules. The effect of Hp10 on ALP was inhibited by addition of noggin protein, suggesting a possible action of this flavanone through the bone morphogenetic protein (BMP) pathway. Indeed, Hp10 significantly induced (1.2- to 1.4-fold) mRNA expression of genes involved in this signalling pathway (i.e., BMP2, BMP4, Runx2 and Osterix) after 48 h of exposure. This was strengthened by enhanced phosphorylation of the complex Smad1/5/8. Osteocalcin mRNA level was up-regulated by Hp only at 10 μM (2.2 fold vs. control). The same dose of Hp significantly decreased osteopontin (OPN) protein level (50% vs. control) after 14 days of culture. Our findings suggest that Hp may regulate osteoblast differentiation through BMP signalling and may influence the mineralization process by modulating OPN expression.