Genetic polymorphisms in the hypothalamic pathway in relation to subsequent weight change--the DiOGenes study

Background

Single nucleotide polymorphisms (SNPs) in genes encoding the components involved in the hypothalamic pathway may influence weight gain and dietary factors may modify their effects.

Aim

We conducted a case-cohort study to investigate the associations of SNPs in candidate genes with weight change during an average of 6.8 years of follow-up and to examine the potential effect modification by glycemic index (GI) and protein intake.

Methods and Findings

Participants, aged 20–60 years at baseline, came from five European countries. Cases (‘weight gainers’) were selected from the total eligible cohort (n = 50,293) as those with the greatest unexplained annual weight gain (n = 5,584). A random subcohort (n = 6,566) was drawn with the intention to obtain an equal number of cases and noncases (n = 5,507). We genotyped 134 SNPs that captured all common genetic variation across the 15 candidate genes; 123 met the quality control criteria. Each SNP was tested for association with the risk of being a ‘weight gainer’ (logistic regression models) in the case-noncase data and with weight gain (linear regression models) in the random subcohort data. After accounting for multiple testing, none of the SNPs was significantly associated with weight change. Furthermore, we observed no significant effect modification by dietary factors, except for SNP rs7180849 in the neuromedin β gene (NMB). Carriers of the minor allele had a more pronounced weight gain at a higher GI (P = 2×10⁻⁷).

Conclusions

We found no evidence of association between SNPs in the studied hypothalamic genes with weight change. The interaction between GI and NMB SNP rs7180849 needs further confirmation.     

Interleukin 12B (IL12B) genetic variation and pulmonary tuberculosis: a study of cohorts from The Gambia, Guinea-Bissau, United States and Argentina

We examined whether polymorphisms in interleukin-12B (IL12B) associate with susceptibility to pulmonary tuberculosis (PTB) in two West African populations (from The Gambia and Guinea-Bissau) and in two independent populations from North and South America. Nine polymorphisms (seven SNPs, one insertion/deletion, one microsatellite) were analyzed in 321 PTB cases and 346 controls from Guinea-Bissau and 280 PTB cases and 286 controls from The Gambia. For replication we studied 281 case and 179 control African-American samples and 221 cases and 144 controls of European ancestry from the US and Argentina. First-stage single locus analyses revealed signals of association at IL12B 3′ UTR SNP rs3212227 (unadjusted allelic p = 0.04; additive genotypic p = 0.05, OR = 0.78, 95% CI [0.61–0.99]) in Guinea-Bissau and rs11574790 (unadjusted allelic p = 0.05; additive genotypic p = 0.05, OR = 0.76, 95% CI [0.58–1.00]) in The Gambia. Association of rs3212227 was then replicated in African-Americans (rs3212227 allelic p = 0.002; additive genotypic p = 0.05, OR = 0.78, 95% CI [0.61–1.00]); most importantly, in the African-American cohort, multiple significant signals of association (seven of the nine polymorphisms tested) were detected throughout the gene. These data suggest that genetic variation in IL12B, a highly relevant candidate gene, is a risk factor for PTB in populations of African ancestry, although further studies will be required to confirm this association and identify the precise mechanism underlying it.     

Glucose Metabolism and Regulation in Lactating Mink (Mustela Vison) - Effects of Low Dietary Protein Supply

Eighteen lactating mink raising litters of 6 to 7 kits were fed ad libitum from parturition on diets with 32% of ME derived from protein and decreasing fat:carbohydrate ratios [high fat:low carbohydrate (HFLC): 67:1, medium fat:medium carbohydrate (MFMC): 52:16, low fat:high carbohydrate (LFHC): 37:31]. Four weeks post partum the dams were fitted with a jugular vein catheter, and the experiment started with a 3 hours fasting period, after which the dams were fed 210kJ ME of the experimental diets. Blood samples were collected 10 and 5min before feeding and 30, 60, 90, 120, 150 and 180min postprandially. Two hours postprandially a single dose of 50µCi U-¹⁴C-labelled glucose was administered to each dam and blood samples were collected 5, 10, 20, 30, 45 and 60min after the tracer administration. Plasma concentrations of glucose and insulin 30 to 120min postprandially were higher in dams fed the LFHC diet, than in dams fed the HFLC diet, values for dams fed the MFMC diet being intermediate. Plasma glucagon concentrations were not significantly affected by dietary treatment. The glucagon:insulin ratios decreased postprandially in all dams, the response being significant in dams fed the LFHC diet. Plasma concentrations of urea were not significantly affected by dietary treatment. Plasma FFA concentrations tended to increase postprandially in dams fed the HFLC diet. Glucose turnover rates were approximately 4.0% permin in all dams, irrespective of dietary treatment. However, the daily glucose flux was lower in dams fed the HFLC diet than in dams fed the LFHC diet, and tended to be lower than in dams fed the MFMC diet. In conclusion, a dietary protein supply of 32% of ME simultaneously with a carbohydrate supply of 16% or 31% of ME had no adverse effects on glucose homeostasis or glucose metabolism in lactating mink.     

Systemic Inflammation in Progressive Multiple Sclerosis Involves Follicular T-Helper,Th17- and Activated B-Cells and Correlates with Progression

Pathology studies of progressive multiple sclerosis (MS) indicate a major role of inflammation including Th17-cells and meningeal inflammation with ectopic lymphoid follicles, B-cells and plasma cells, the latter indicating a possible role of the newly identified subset of follicular T-helper (T_FH) cells. Although previous studies reported increased systemic inflammation in progressive MS it remains unclear whether systemic inflammation contributes to disease progression and intrathecal inflammation. This study aimed to investigate systemic inflammation in progressive MS and its relationship with disease progression, using flow cytometry and gene expression analysis of CD4⁺ and CD8⁺T-cells, B-cells, monocytes and dendritic cells. Furthermore, gene expression of cerebrospinal fluid cells was studied. Flow cytometry studies revealed increased frequencies of ICOS⁺T_FH-cells in peripheral blood from relapsing-remitting (RRMS) and secondary progressive (SPMS) MS patients. All MS subtypes had decreased frequencies of Th1 T_FH-cells, while primary progressive (PPMS) MS patients had increased frequency of Th17 T_FH-cells. The Th17-subset, interleukin-23-receptor⁺CD4⁺T-cells, was significantly increased in PPMS and SPMS. In the analysis of B-cells, we found a significant increase of plasmablasts and DC-SIGN⁺ and CD83⁺B-cells in SPMS. ICOS⁺T_FH-cells and DC-SIGN⁺B-cells correlated with disease progression in SPMS patients. Gene expression analysis of peripheral blood cell subsets substantiated the flow cytometry findings by demonstrating increased expression of IL21, IL21R and ICOS in CD4⁺T-cells in progressive MS. Cerebrospinal fluid cells from RRMS and progressive MS (pooled SPMS and PPMS patients) had increased expression of T_FH-cell and plasmablast markers. In conclusion, this study is the first to demonstrate the potential involvement of activated T_FH-cells in MS. The increased frequencies of Th17-cells, activated T_FH- and B-cells parallel findings from pathology studies which, along with the correlation between activated T_FH- and B-cells and disease progression, suggest a pathogenic role of systemic inflammation in progressive MS. These observations may have implications for the treatment of progressive MS.     

ADP Protects Cardiac Mitochondria under Severe Oxidative Stress

ADP is not only a key substrate for ATP generation, but also a potent inhibitor of mitochondrial permeability transition pore (mPTP). In this study, we assessed how oxidative stress affects the potency of ADP as an mPTP inhibitor and whether its reduction of reactive oxygen species (ROS) production might be involved. We determined quantitatively the effects of ADP on mitochondrial Ca²⁺ retention capacity (CRC) until the induction of mPTP in normal and stressed isolated cardiac mitochondria. We used two models of chronic oxidative stress (old and diabetic mice) and two models of acute oxidative stress (ischemia reperfusion (IR) and tert-butyl hydroperoxide (t-BH)). In control mitochondria, the CRC was 344 ± 32 nmol/mg protein. 500 μmol/L ADP increased CRC to 774 ± 65 nmol/mg protein. This effect of ADP seemed to relate to its concentration as 50 μmol/L had a significantly smaller effect. Also, oligomycin, which inhibits the conversion of ADP to ATP by F₀F₁ATPase, significantly increased the effect of 50 μmol/L ADP. Chronic oxidative stress did not affect CRC or the effect of 500 μmol/L ADP. After IR or t-BH exposure, CRC was drastically reduced to 1 ± 0.2 and 32 ± 4 nmol/mg protein, respectively. Surprisingly, ADP increased the CRC to 447 ± 105 and 514 ± 103 nmol/mg protein in IR and t-BH, respectively. Thus, it increased CRC by the same amount as in control. In control mitochondria, ADP decreased both substrate and Ca²⁺-induced increase of ROS. However, in t-BH mitochondria the effect of ADP on ROS was relatively small. We conclude that ADP potently restores CRC capacity in severely stressed mitochondria. This effect is most likely not related to a reduction in ROS production. As the effect of ADP relates to its concentration, increased ADP as occurs in the pathophysiological situation may protect mitochondrial integrity and function.     

Response to multi‐generational selection under elevated [CO2] in two temperature regimes suggests enhanced carbon assimilation and increased reproductive output in Brassica napus L.

Functional plant traits are likely to adapt under the sustained pressure imposed by environmental changes through natural selection. Employing Brassica napus as a model, a multi‐generational study was performed to investigate the potential trajectories of selection at elevated [CO₂] in two different temperature regimes. To reveal phenotypic divergence at the manipulated [CO₂] and temperature conditions, a full‐factorial natural selection regime was established in a phytotron environment over the range of four generations. It is demonstrated that a directional response to selection at elevated [CO₂] led to higher quantities of reproductive output over the range of investigated generations independent of the applied temperature regime. The increase in seed yield caused an increase in aboveground biomass. This suggests quantitative changes in the functions of carbon sequestration of plants subjected to increased levels of CO₂ over the generational range investigated. The results of this study suggest that phenotypic divergence of plants selected under elevated atmospheric CO₂ concentration may drive the future functions of plant productivity to be different from projections that do not incorporate selection responses of plants. This study accentuates the importance of phenotypic responses across multiple generations in relation to our understanding of biogeochemical dynamics of future ecosystems. Furthermore, the positive selection response of reproductive output under increased [CO₂] may ameliorate depressions in plant reproductive fitness caused by higher temperatures in situations where both factors co‐occur.     

Separate and Joint Associations of Occupational and Leisure-Time Sitting with Cardio-Metabolic Risk Factors in Working Adults: A Cross-Sectional Study

Background

The workplace is a main setting for prolonged sitting for some occupational groups. Convincing evidence has recently accumulated on the detrimental cardio-metabolic health effects of leisure-time sitting. Yet, much less is known about occupational sitting, and the potential health risk attached compared to leisure-time sitting.

Objective

To explore the separate and joint associations of occupational and leisure-time sitting with cardio-metabolic risk factors in working adults.

Methods

All working adults (N = 2544) from the Health2006, a Danish population-based study, were included in this cross-sectional study. Participants reported hours of sitting during work, during leisure-time along with socio-demographic and behavioral characteristics, including physical activity. Cardio-metabolic risk factors (waist circumference, body mass index, body fat percentage, total cholesterol, HDL cholesterol, LDL cholesterol, triglycerides, insulin, hemoglobin A1c and plasma glucose) were measured. Associations were explored by linear regression for leisure-time, occupational, and overall sitting time.

Results

Statistically significant (p<.05) detrimental associations of leisure-time sitting were observed with all cardio-metabolic risk factors, except hemoglobin A1c and plasma glucose. Similarly, occupational sitting time was significantly detrimentally associated with HDL cholesterol, triglycerides, and insulin. For categories of sitting time, a joint adverse association of sitting much during both work-time and leisure-time was observed.

Conclusion

The associations of occupational sitting time with cardio-metabolic risk factors were fewer and weaker compared to leisure-time sitting. Yet, the joint associations of occupational and leisure-time sitting with cardio-metabolic risk factors were higher than the separate. Our findings amplify the need for further focus in this area prior to making assumptions about equivalent health risks across sedentary behaviors. To our knowledge, this is the first study to contrast the deleterious associations of prolonged occupational and leisure-time sitting, both separately and jointly.     

Acute Myocardial Infarction and Pulmonary Diseases Result in Two Different Degradation Profiles of Elastin as Quantified by Two Novel ELISAs

Background

Elastin is a signature protein of the arteries and lungs, thus it was hypothesized that elastin is subject to enzymatic degradation during cardiovascular and pulmonary diseases. The aim was to investigate if different fragments of the same protein entail different information associated to two different diseases and if these fragments have the potential of being diagnostic biomarkers.

Methods

Monoclonal antibodies were raised against an identified fragment (the ELM-2 neoepitope) generated at the amino acid position ‘552 in elastin by matrix metalloproteinase (MMP) −9/−12. A newly identified ELM neoepitope was generated by the same proteases but at amino acid position ‘441. The distribution of ELM-2 and ELM, in human arterial plaques and fibrotic lung tissues were investigated by immunohistochemistry. A competitive ELISA for ELM-2 was developed. The clinical relevance of the ELM and ELM-2 ELISAs was evaluated in patients with acute myocardial infarction (AMI), no AMI, high coronary calcium, or low coronary calcium. The serological release of ELM-2 in patients with chronic obstructive pulmonary disease (COPD) or idiopathic pulmonary fibrosis (IPF) was compared to controls.

Results

ELM and ELM-2 neoepitopes were both localized in diseased carotid arteries and fibrotic lungs. In the cardiovascular cohort, ELM-2 levels were 66% higher in serum from AMI patients compared to patients with no AMI (p<0.01). Levels of ELM were not significantly increased in these patients and no correlation was observed between ELM-2 and ELM. ELM-2 was not elevated in the COPD and IPF patients and was not correlated to ELM. ELM was shown to be correlated with smoking habits (p<0.01).

Conclusions

The ELM-2 neoepitope was related to AMI whereas the ELM neoepitope was related to pulmonary diseases. These results indicate that elastin neoepitopes generated by the same proteases but at different amino acid sites provide different tissue-related information depending on the disease in question.     

Identification of a cyclin B1-derived CTL epitope eliciting spontaneous responses in both cancer patients and healthy donors

With the aim to identify cyclin B1-derived peptides with high affinity for HLA-A2, we used three in silico prediction algorithms to screen the protein sequence for possible HLA-A2 binders. One peptide scored highest in all three algorithms, and the high HLA-A2-binding affinity of this peptide was verified in an HLA stabilization assay. By stimulation with peptide-loaded dendritic cells a CTL clone was established, which was able to kill two breast cancer cell lines in an HLA-A2-dependent and peptide-specific manner, demonstrating presentation of the peptide on the surface of cancer cells. Furthermore, blood from cancer patients and healthy donors was screened for spontaneous T-cell reactivity against the peptide in IFN-γ ELISPOT assays. Patients with breast cancer, malignant melanoma, or renal cell carcinoma hosted powerful and high-frequency T-cell responses against the peptide. In addition, when blood from healthy donors was tested, similar responses were observed. Ultimately, serum from cancer patients and healthy donors was analyzed for anti-cyclin B1 antibodies. Humoral responses against cyclin B1 were frequently detected in both cancer patients and healthy donors. In conclusion, a high-affinity cyclin B1-derived HLA-A2-restricted CTL epitope was identified, which was presented on the cell surface of cancer cells, and elicited spontaneous T-cell responses in cancer patients and healthy donors.