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1.

Background

Several meta-analyses have demonstrated that the rs662 polymorphism in Paraoxonase 1 gene (PON1) gene is associated with coronary heart disease (CHD). However, it is still uncertain whether this polymorphism is associated with the plasma levels of oxidized low-density lipoprotein (Ox-LDL) and lipids. This meta-analysis is aimed to clarify the relationships between the rs662 polymorphism and plasma levels of Ox-LDL and lipids.

Methods

By searching in PubMed, Google Scholar, Web of Science, Cochrane Library, Wanfang, VIP and CNKI databases, 5 studies (1369 subjects) and 85 studies (46,740 subjects) were respectively identified for Ox-LDL association analysis and lipid association analysis. Standardized mean difference (SMD) was used to estimate the effects of the rs662 polymorphism on plasma Ox-LDL and lipid levels.

Results

The carriers of the variant R allele had higher levels of Ox-LDL (SMD?=?0.23, 95% CI?=?0.10–0.36, P?<? 0.01), triglyceride (TG) (SMD?=?0.06, 95% CI?=?0.01–0.11, P?=?0.02), total cholesterol (TC) (SMD?=?0.04, 95% CI?=?0.00–0.07, P?=?0.05) and low-density lipoprotein cholesterol (LDL-C) (SMD?=?0.04, 95% CI?=?0.00–0.08, P?=?0.04) than the non-carriers.

Conclusions

This meta-analysis suggests that the association between the PON1 rs662 polymorphism and CHD may partly be mediated by abnormal Ox-LDL and lipid levels caused by the R allele.
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2.

Purpose

The consumption of Brazil nuts has been associated with benefits to lipid metabolism and reductions in total cholesterol and LDL concentrations. They are the richest natural source of selenium which has essential functions in human physiology. Genetic polymorphisms in Selenoprotein P could impair lipid and glucose metabolisms. The aim of this work was to verify the influence of polymorphisms in genes for selenoproteins on blood lipid levels after dietary supplementation with Brazil nuts in healthy adults.

Methods

The study included 130 healthy volunteers selected at the University of São Paulo, Brazil. They were supplemented with one nut a day for 8 weeks, followed by 8 weeks without intervention. The following analyses were performed: anthropometric measurements, serum fasting glucose, lipid profile, C-reactive protein and plasma MDA levels. The volunteers were genotyped for SNPs rs1050450, rs3811699, rs1800699, rs713041, rs3877899, rs7579, rs34713741, and rs5845 in genes for selenoproteins.

Results

The concentrations of total cholesterol and fasting glucose levels decreased after 8 weeks of supplementation (p < 0.05). Glucose levels were modulated by rs3877899 in SEPP1, with significantly lower levels observed for individuals with the GA + AA genotype (p = 0.025). In addition, rs7579 was associated with cholesterol concentrations, which were significantly lower for individuals with the GG genotype (p = 0.053).

Conclusions

Supplementation with one Brazil nut a day for 8 weeks reduced total cholesterol and glucose levels. Furthermore, our results suggest that rs3877899 might be associated with glucose concentrations and rs7579 with cholesterol concentrations. Therefore, the effect of genetic variations should be considered in future nutritional interventions evaluating the response to Brazil nut supplementation.
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3.

Objective

Telomere length shortening is modulated not only by aging, but also by both genetic and environmental factors. The aim of this study was to investigate the interactions between antioxidant nutrient metabolism-related gene single nucleotide polymorphisms (the genetic factors) and nutrient intake (the environmental factors) in their effects on telomere length shortening.

Setting & participants

Data were collected on the relative telomere lengths (RTLs) of buccal cells and the habitual food intake of 70 healthy Japanese adults.

Measurements

All subjects were genotyped for two common single nucleotide polymorphisms: rs6564851 in the β-carotene-15,15’-mono-oxygenase 1 (BCMO1) gene and rs362090 in the intestine-specific homeobox (ISX) gene.

Results

Univariate analysis revealed that buccal RTL was not significantly modulated by either age or gender. Then, we subdivided the study population into four groups based on combinations of the rs6564851 and rs362090 genotypes. After this subdivision, we showed a positive effect of daily α- or β-carotene intake on buccal RTL in the ISX rs362090 G-allele carrier + BCMO1 rs6564851 GG-genotype group (p = 0.026). Additionally, daily intake of another antioxidative fat-soluble vitamin, α-tocopherol, was positively associated with buccal RTL in the ISX rs362090 AA-homozygote + BCMO1 rs6564851 T-allele carrier group (p = 0.037).

Conclusion

Our study clearly indicates that high dietary intake of the antioxidants α, β-carotene and α-tocopherol protects buccal cells from RTL shortening, depending on the genetic background of antioxidant vitamin-related genes.
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4.

Purpose

Xenobiotic metabolism is related to the interplay between diet and breast cancer (BC) risk. This involves detoxification enzymes, which are polymorphic and metabolise various dietary metabolites. An important characteristic of this pathway is that chemoprotective micronutrients can act not only as substrates but also as inducers for these enzymes. We investigated whether functional GSTP1 (p.Ile105Val-rs1695), NAT2 (590G>A-rs1799930) SNPs and GSTM1 and GSTT1 deletion polymorphisms could modulate the effect of the Mediterranean diet (MD) on BC risk, in Greek-Cypriot women.

Methods

Genotyping was performed on women from the MASTOS case–control study of BC in Cyprus. A 32-item food-frequency questionnaire was used to obtain dietary intake information. A dietary pattern, which closely resembles the MD (high loadings of vegetables, fruit, legumes and fish), was previously derived with principal component analysis and was used as our dietary variable.

Results

GSTT1 null genotype increased BC risk compared with the homozygous non-null GSTT1 genotype (OR 1.21, 95 % CI 1.01–1.45). Increasing adherence to the MD reduced BC risk in women with at least one GSTP1 Ile allele (OR for Ile/Ile = 0.84, 95 % CI 0.74–0.95, for Ile/Val = 0.73, 95 % CI 0.62–0.85) or one NAT2 590G allele (OR for 590 GG = 0.73, 95 % CI 0.63–0.83, for 590 GA = 0.81, 95 % CI 0.70–0.94). p interaction values were not, however, statistically significant.

Conclusion

The homozygous null GSTT1 genotype could be a risk allele for BC among Greek-Cypriot women. The anticarcinogenic effects of the high adherence to MD against BC risk could also be further enhanced when combined with the wild-type alleles of the detoxification GSTP1 or NAT2 SNPs.
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5.

Objectives

RNF213 p.R4810K is a founder polymorphism that confers genetic susceptibility to moyamoya disease in East Asia. Only a few studies have investigated the symptoms and disease histories of RNF213 p.R4810K carriers in Japan. This study investigated the frequency of RNF213 p.R4810K in the general Japanese population and the health characteristics of the carriers.

Methods

Through a health-promotion campaign in the city of Uji, Japan, 519 subjects (120 males and 399 females) of the general Japanese population were genotyped for RNF213 p.R4810K and interviewed to determine health characteristics.

Results

Nine RNF213 p.R4810K heterozygous carriers (GA genotype) and no RNF213 p.R4810K homozygous carriers (AA genotype) were found among the 519 individuals. The estimates of the genotypes and allele frequencies for RNF213 p.R4810K were 1.73 and 0.87 %, respectively. There were no obvious differences in age, gender ratio, body mass index, hypertension, dyslipidemia, diabetes, kidney disease, liver disease, heart disease, or drinking or smoking habits between carriers and non-carriers. Interestingly, one patient with moyamoya disease was found among the nine RNF213 p.R4810K carriers.

Conclusions

This study showed the genotypes and allele frequencies of RNF213 p.R4810K in the general Japanese population to be similar to results of previous reports.
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6.

Objectives

Many East Asians have the genetic polymorphisms rs1229984 in alcohol dehydrogenase 1B (ADH1B) and rs671 in aldehyde dehydrogenase 2 (ALDH2). Here we analyzed the relationships of the two genotypes with alcohol sensitivity, drinking behavior and problem drinking among older and younger men living in rural areas of Japan.

Methods

The subjects were 718 Japanese men aged 63.3 ± 10.8 (mean ± SD), categorized into the older (≥65 years, n = 357) and younger (<65 years, n = 361) groups. Facial flushing frequency, drinking behavior and positive CAGE results were compared among the genotypes using Bonferroni-corrected χ2 test and a multivariate logistic regression analysis adjusting for age, BMI and lifestyle factors.

Results

The frequency of ‘always’ facial flushing among the ADH1B*1/*2 carriers was significantly lower than that among the ADH1B*2/*2 carriers in the older group (P < 0.01). The alcohol consumption (unit/day) in the ADH1B*1/*2 carriers tended to be higher compared with that in the ADH1B*2/*2 carriers among the older group (P = 0.050). In the younger group, no significant differences in alcohol sensitivity and drinking habits were generally found among the ADH1B genotypes. The ADH1B*1/*1 genotype tended to be positively associated with problem drinking in the older group (P = 0.080) but not in the younger group. The ALDH2 genotypes consistently and strongly affected the alcohol sensitivity, drinking behavior and problem drinking in both the younger and older group.

Conclusions

We for the first time observed a significant difference in alcohol sensitivity between ADH1B*1/*2 and ADH1B*2/*2 in older men aged 65 and above.
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7.

Purpose

To examine the effect of asbestos exposure on global DNA methylation and determine whether lung function and inflammatory and fibrosis biomarkers are correlated with the methylation state.

Methods

A total of 26 healthy subjects without asbestos exposure (Group 1), 47 healthy subjects with exposure (Group 2), and 52 subjects with benign asbestos-related disorders (ARDs) (Group 3) participated in this cross-sectional study. Blood global 5-methylcytosine (5mC) and serum TNF-α, collagen IV, CCL5 and CC16 concentrations were analyzed using enzyme-linked immunosorbent assay-like assays. Spirometric maneuvers were performed to assess lung function.

Results

Decreased 5mC levels were observed in Groups 2 and 3 compared to Group 1, irrespective of lung function (p?<?0.01). There was no significant change in 5mC between Groups 2 and 3. Overall, 5mC was negatively correlated with CCL5 and collagen IV (p?<?0.05), but no significant inverse relationship was found between 5mC and CCL5 or collagen IV in each group. Additionally, both 5mC and CC16 were inversely associated with FEV1/FVC% (p?=?0.001, adjusted R 2?=?0.145) for non-smokers, and consistently significant inverse relationships were found between CC16 and FEV1/FVC%, independent of asbestos exposure.

Conclusions

Asbestos exposure causes global DNA hypomethylation. DNA hypomethylation has no influence on serum biomarkers and lung function in asbestos-exposed population with or without pleural and pulmonary parenchymal abnormalities.
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8.
9.
10.

Background

Diarrhea is a major contributor to the burden of morbidity and mortality in children; it accounts for a median of 11% of all deaths among children aged less than 5 years, amounting to approximately 0.8 million deaths per year. Currently there is a dearth of literature exploring the effectiveness of antibiotics for diarrhea due to Cholera, Shigella and cryptosporidiosis in children.

Methods

We reviewed the literature reporting the effect of antibiotics for the treatment of diarrhea due to Cholera, Shigella and Cryptosporidium in children under five years. We used a standardized abstraction and grading format and performed meta-analyses to determine the effect of the treatment with various antibiotics on mortality and rates of clinical and bacteriological/parasitological failure. The CHERG Standard Rules were applied to determine the final effect of treatment with antibiotics on diarrhea morbidity and mortality.

Results

For Cholera; the evidence was weak to recommend any effect on mortality. For Shigella; there was no data on mortality; either all-cause or cause specific, hence we used clinical failure rates as a proxy for Shigella deaths and propose that treatment of Shigella dysentery with antibiotics can result in a 82% reduction in diarrhea mortality due to Shigella. For cryptosporidiosis; there was data on all-cause mortality but the evidence was weak hence we used clinical failure rates as a proxy for mortality to estimate that antimicrobial treatment of diarrhea due to cryptosporidiosis can result in a 54% reduction in mortality.

Conclusions

There is evidence to recommend antibiotic use for reduction of morbidity and mortality due to Cholera, Shigella and Cryptosporidium. We recommend that more clinical trials should be conducted to evaluate the efficacy and safety of first- and second- line drugs currently in use for treatment for diarrhea and dysentery in both developing and developed countries.
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11.

Background

Heterocyclic aromatic amines (HAAs) may confer prostate cancer risk; however, the evidence is inconclusive and the activity of HAA-metabolizing enzymes is modulated by gene variants. The purpose of our study was to determine whether there was evidence of an association between HAA intake, polymorphisms in NAT2, CYP1A1, and CYP1A2 and prostate cancer risk in Japanese men.

Methods

Secondary data analysis of an observational case control study was performed. Among 750 patients with prostate cancer and 870 healthy controls, 351 cases and 351 age-matched controls were enrolled for analysis. HAA intake was estimated using a food frequency questionnaire and genotypes were scored by TaqMan real-time PCR assay. Logistic regression analysis was conducted according to affected/control status.

Results

We found that high HAA intake was significantly associated with an increased risk of prostate cancer (odds ratio (OR), 1.90; 95% confidence interval (95% CI), 1.40–2.59). The increased risk of prostate cancer was observed among individuals with the NAT2 slow acetylator phenotype (OR, 1.65; 95% CI, 1.04–2.61), CYP1A1 GA + GG genotype (OR, 1.27; 95% CI, 1.02–1.59), and CYP1A2 CA + AA genotype (OR, 1.43; 95% CI, 1.03–2.00). In addition, CYP1A1 GA + GG genotypes were associated with increased cancer risk in low (OR, 2.05; 95% CI, 1.19–3.63), moderate (OR, 1.72; 95% CI, 1.07–2.76), and high (OR, 2.86; 95% CI, 1.83–4.47) HAA intake groups.

Conclusions

Our results suggest that high HAA intake is a risk factor of prostate cancer, and genotypes related to HAA metabolic enzymes can modulate the degree of the risk.
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12.

Background

Nowadays, the bacterial contamination in the hospital environment is of particular concern because the hospital-acquired infections (HAIs), also known as nosocomial infections, are responsible for significant morbidity and mortality. This work evaluated the capability of Enterococcus hirae to form biofilm on different surfaces and the action of two biocides on the produced biofilms.

Methods

The biofilm formation of E. hirae ATCC 10541 was studied on polystyrene and stainless steel surfaces through the biomass quantification and the cell viability at 20 and 37 °C. The effect of LH IDROXI FAST and LH ENZYCLEAN SPRAY biocides on biomasses was expressed as percentage of biofilm reduction. E. hirae at 20 and 37 °C produced more biofilm on the stainless steel in respect to the polystyrene surface. The amount of viable cells was greater at 20 °C than with 37 °C on the two analyzed surfaces. Biocides revealed a good anti-biofilm activity with the most effect for LH ENZYCLEAN SPRAY on polystyrene and stainless steel at 37 °C with a maximum biofilm reduction of 85.72 and 86.37%, respectively.

Results

E. hirae is a moderate biofilm producer depending on surface material and temperature, and the analyzed biocides express a remarkable antibiofilm action.

Conclusion

The capability of E. hirae to form biofilm can be associated with its increasing incidence in hospital-acquired infections, and the adoption of suitable disinfectants is strongly recommended.
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13.

Purpose

Gestational diabetes mellitus (GDM) represents the most frequent metabolic disorder in pregnancy. Since dietary intake plays an important role in obesity and type 2 diabetes development, it is likely to be for the susceptibility to GDM too. Food preferences, driving partly the diet composition, are changing during pregnancy. Taste and genetic variability in taste receptors is an important factor in determining food preferences. Aims of our study were (1) to characterize dietary habits of pregnant women and to find possible differences in food preferences between healthy pregnant women and those with GDM and (2) to ascertain possible association of several single nucleotide polymorphisms (SNPs) in taste receptor (TR) genes with GDM.

Methods

A total of 363 pregnant women (293 with GDM and 70 with physiologic pregnancy) were included in the study. Dietary pattern spanning the period of approx. 6 months preceding the time of GDM screening was assessed using a semi-quantitative food frequency questionnaire. A total of five SNPs in TR genes were selected for genotyping based on their functionality or previous associations.

Results

Women with GDM exhibited significantly more frequent meat consumption (esp. poultry, pork and smoked meat), dairy products and sweet beverages consumption. The legumes consumption was found to be inversely correlated with fasting glycaemia (P = 0.007, Spearman). CC genotype in TAS2R9 gene (SNP rs3741845) was significantly associated with GDM (P = 0.0087, Chi-square test).

Conclusions

Our study showed differences in dietary intake of selected food items between healthy pregnant women and those with GDM and genetic association of bitter taste receptor allele with GDM.
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14.

Purpose

According to the vast body of evidence demonstrating that the intestinal microbiota is undoubtedly linked with overall health, including cancer risk, searching for functional foods and novel prebiotic influencing on beneficial bacteria is necessary. The present study aimed to investigate the potential of polysaccharides from 53 wild-growing mushrooms to stimulate the growth of Lactobacillus acidophilus and Lactobacillus rhamnosus and to determine the digestibility of polysaccharide fractions.

Methods

Mushroom polysaccharides were precipitated with ethanol from aqueous extracts. Determination of growth promoting activity of polysaccharides was performed in U-shaped 96-plates in an ELISA reader in relation to the reference strain of L. acidophilus and two clinical strains of L. rhamnosus. The digestibility of mushroom polysaccharides was investigated in vitro by exposing them to artificial human gastric juice.

Results

Obtained results revealed that fungal polysaccharides stimulate the growth of Lactobacillus strains stronger than commercially available prebiotics like inulin or fructooligosaccharides. Moreover, selected polysaccharides were subjected to artificial human gastric juice and remain undigested in more than 90%.

Conclusion

Obtained results indicate that mushroom polysaccharides are able to pass through the stomach unchanged, reaching the colon and stimulating the growth of beneficial bacteria. Majority of 53 polysaccharide fractions were analysed for the first time in our study. Overall, our findings suggest that polysaccharide fractions from edible mushrooms might be useful in producing functional foods and nutraceuticals.
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15.

Purpose

To report total fluid intake (TFI) and the intake of different fluid types in adults (≥?18 years old) from Mexico, Argentina, Brazil and Uruguay. To compare intakes between countries and with recommended adequate intake (AI) of water from fluids.

Methods

Cross-sectional data were collected using a validated liquid intake 7-day record (Liq.In 7 ) in populations from Argentina (n?=?1089), Brazil (n?=?477), Mexico (n?=?1677) and Uruguay (n?=?554). Population characteristics, including age, gender, body mass index and socioeconomic level were recorded. Mean TFI was compared with the AI of water from fluids set by the USA Institute of Medicine.

Results

The lowest TFI was recorded in Mexican women (1748 mL/day) and the highest in Argentinean men (2318 mL/day). Median daily TFI was significantly different between countries; Uruguay and Argentina had higher values than Mexico and Brazil. In the former, plain water contributed to only 25% of TFI, the remainder being predominantly from hot beverages. Approximately, a third of adults did not drink enough fluid to meet the recommended AI. High SSB consumption was reported, which was significantly different between countries (p?<?0.05), the highest being in Mexico (median 25–75th percentiles): 531 (300–895 mL/day.

Conclusions

This survey highlights the need to increase water consumption and reduce SSB intake in this region to avoid potential associated health risks. These findings may be useful information in monitoring public health policy strategies.
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16.

Purpose

This study aimed to identify and characterize patterns of fluid intake in children and adolescents from six countries: Argentina, Brazil, China, Indonesia, Mexico and Uruguay.

Methods

Data on fluid intake volume and type amongst children (4–9 years; N?=?1400) and adolescents (10–17 years; N?=?1781) were collected using the validated 7-day fluid-specific record (Liq.In7 record). To identify relatively distinct clusters of subjects based on eight fluid types (water, milk and its derivatives, hot beverages, sugar-sweetened beverages (SSB), 100% fruit juices, artificial/non-nutritive sweetened beverages, alcoholic beverages, other beverages), a cluster analysis (partitioning around k-medoids algorithm) was used. Clusters were then characterized according to their socio-demographics and lifestyle indicators.

Results

The six interpretable clusters identified were: low drinkers–SSB (n 523), low drinkers–water and milk (n 615), medium mixed drinkers (n 914), high drinkers–SSB (n 513), high drinkers–water (n 352) and very high drinkers–water (n 264). Country of residence was the dominant characteristic, followed by socioeconomic level, in all six patterns.

Conclusions

This analysis showed that consumption of water and SSB were the primary drivers of the clusters. In addition to country, socio-demographic and lifestyle factors played a role in determining the characteristics of each cluster. This information highlights the need to target interventions in particular populations aimed at changing fluid intake behavior and improving health in children and adolescents.
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17.

Background

There is little data on Trichomonas vaginalis infection in Ghana. This study evaluated the prevalence of trichomoniasis using different diagnostic methods and determined the risk factors for infection in patients.

Methods

A structured questionnaire was administered. Vaginal swabs, urethral swabs and urine specimens were obtained from consenting patients; and the samples processed following standard protocols. The presence of T. vaginalis was determined using wet mount microscopy and polymerase chain reaction (PCR) as gold standard. We also assessed the diagnostic performance the JD’s Trichomonas V® rapid antigen test to inform clinical practice.

Results

The PCR assay detected T. vaginalis positivity in 64 of 150 patients (42.6, 95%CI:35.0, 50.6) including all positive samples of wet mount microscopy and JD’s Trichomonas V® test. Wet mount microscopy showed low sensitivity (31.6%), high specificity (100%), moderate positive predictive value (75.0%), moderate positive likelihood ratio (3.0), and weak agreement (Cohen’s kappa, 0.283) with PCR assay. The JD’s Trichomonas V® test displayed lower sensitivity (25.0%), specificity (83.3%), and weaker measure of agreement (Cohen’s kappa, 0.233) with PCR. In multivariate analysis, the strongest independent predictor for T. vaginalis was female gender [adjusted odds ratio (AOR), 24.89; 95% confidence interval (CI): 10.58, 51.21; P-value<?0.001]. Knowledge of STI showed a protective effect against infection with the parasite (AOR, 0.13; 95%CI: 0.07, 0.29; P-value<?0.017).

Conclusion

The sensitivity of wet mount microscopy was low for T. vaginalis screening in our region. The JD’s Trichomonas V® test should not be considered as an alternative test. We recommend mandatory PCR assay for confirmation of negative wet mount results.
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18.

Background

Salvia hispanica seeds have been commonly used by people that seek healthy habits through natural foods to reduce cholesterol and triacylglycerides levels, however, the evidences that support this assumption are still scarce in literature. Here, we aimed to evaluate the lipid lowering effects of chia by using Caenorhabditis elegans as animal model, a nematode that has proven its usefulness for lipid metabolism studies.

Methods

We prepared hexane (HE) and Bligh-Dyer (BDE) extracts, evaluated and compared their safety, antioxidant potential and their lipid-lowering activity in the worms.

Results

The characterization of both extracts demonstrated that there were no differences in their lipid composition; however, BDE depicted better antioxidant potential. Both extracts reduced worm’s survival from 2%, and reproduction was reduced following treatment with both extracts, though a more notable effect was observed in HE-treated worms. In addition, the non-toxic concentration of both extracts (1%) increased stress resistance against paraquat toxicity in an antidote paradigm. Notably, this same concentration of both extracts reduced lipid accumulation in obese worms, which was not caused by food deprivation.

Conclusions

Taken together, our data demonstrate that both extraction methods from chia seeds result in oils that are rich in mono and polyunsaturated fatty acids, which may modulate lipid accumulation and provide antioxidant resistance in C. elegans.
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19.

Background

Vaccination coverage among medical students is often insufficient. Regarding vaccination coverage of students of other fields, there are no currently published data for Germany. Based on high international mobility, an insufficient or unknown vaccination status increases the health risk among students.

Objective

What is the vaccination status among students of different subjects? Are there group-based differences concerning knowledge of ones own vaccination status? Are there differences concerning attitudes towards vaccination?

Material and methods

The survey was conducted at the Technische Universität Dresden during the summer semester 2012 using a semi-standardized questionnaire. Data from 428 students (subjects: medicine, sociology, architecture and physics) were analyzed.

Results

Significantly less medical students reported an unknown vaccination status regarding all the nine vaccinations analyzed compared to students of sociology or technical subjects. Significantly more medical students reported complete vaccination status compared to students of other subjects (χ2-test: p ≤ 0.001; Z?test p ≤ 0.05). Significantly more medical students had the opinion that vaccinations were absolutely necessary compared to students of other subjects (χ2-test: p ≤ 0.001; Z?test p ≤ 0.05).

Conclusion

University students often have insufficient vaccination coverage. Students of subjects other than medicine are often not aware of their own vaccination status. Low-threshold opportunities to check vaccination status and to be vaccinated if necessary should be established, e.?g. at university and the workplace.
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20.

Background

In a previous study, we showed that consumption of diets enriched in saturated fatty acids causes changes in alternative splicing of pre-mRNAs encoding a number of proteins in rat skeletal muscle, including the one encoding skeletal muscle Troponin T (Tnnt3). However, whether saturated fatty acids act directly on muscle cells to modulate alternative pre-mRNA splicing was not assessed. Moreover, the signaling pathway through which saturated fatty acids act to promote changes in alternative splicing is unknown. Therefore, the objective of the present study was to characterize the signaling pathway through which saturated fatty acids act to modulate Tnnt3 alternative splicing.

Methods

The effects of treatment of L6 myotubes with saturated (palmitate), mono- (oleate), or polyunsaturated (linoleate) fatty acids on alternative splicing of pre-mRNA was assessed using Tnnt3 as a marker gene.

Results

Palmitate treatment caused a two-fold change (p?<?0.05) in L6 myotube Tnnt3 alternative splicing whereas treatment with either oleate or linoleate had minimal effects compared to control myotubes. Treatment with a downstream metabolite of palmitate, ceramide, had effects similar to palmitate on Tnnt3 alternative splicing and inhibition of de novo ceramide biosynthesis blocked the palmitate-induced alternative splicing changes. The effects of palmitate and ceramide on Tnnt3 alternative splicing were accompanied by a 40–50% reduction in phosphorylation of Akt on S473. However, inhibition of de novo ceramide biosynthesis did not prevent palmitate-induced Akt dephosphorylation, suggesting that palmitate may act in an Akt-independent manner to modulate Tnnt3 alternative splicing. Instead, pre-treatment with okadaic acid at concentrations that selectively inhibit protein phosphatase 2A (PP2A) blocked both palmitate- and ceramide-induced changes in Tnnt3 alternative splicing, suggesting that palmitate and ceramide act through PP2A to modulate Tnnt3 alternative splicing.

Conclusions

Overall, the data show that fatty acid saturation level and ceramides are important factors modulating alternative pre-mRNA splicing through activation of PP2A.
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