The effects of physiological adaptations to calorie restriction on global cell proliferation rates

Calorie restriction (CR) reduces the rate of cell proliferation in mitotic tissues. It has been suggested that this reduction in cell proliferation may mediate CR-induced increases in longevity. However, the mechanisms that lead to CR-induced reductions in cell proliferation rates remain unclear. To evaluate the CR-induced physiological adaptations that may mediate reductions in cell proliferation rates, we altered housing temperature and access to voluntary running wheels to determine the effects of food intake, energy expenditure, percent body fat, and body weight on proliferation rates of keratinocytes, liver cells, mammary epithelial cells, and splenic T-cells in C57BL/6 mice. We found that ～20% CR led to a reduction in cell proliferation rates in all cell types. However, lower cell proliferation rates were not observed with reductions in 1) food intake and energy expenditure in female mice housed at 27°C, 2) percent body fat in female mice provided running wheels, or 3) body weight in male mice provided running wheels compared with ad libitum-fed controls. In contrast, reductions in insulin-like growth factor I were associated with decreased cell proliferation rates. Taken together, these data suggest that CR-induced reductions in food intake, energy expenditure, percent body fat, and body weight do not account for the reductions in global cell proliferation rates observed in CR. In addition, these data are consistent with the hypothesis that reduced cell proliferation rates could be useful as a biomarker of interventions that increase longevity.     

BglII and MunI: what a difference a base makes

Restriction endonucleases are resilient to alterations in their DNA-binding specificities. Structures of the BglII and MunI endonucleases bound to their palindromic DNA sites, which differ by only their outer base pairs from the recognition sequences of BamHI and EcoRI, respectively, have recently been determined. A comparison of these complexes reveals surprising differences and similarities in structure, and provides a basis for understanding the immutability of restriction endonucleases.     

Inferring population structure and relationship using minimal independent evolutionary markers in Y-chromosome: a hybrid approach of recursive feature selection for hierarchical clustering

Inundation of evolutionary markers expedited in Human Genome Project and 1000 Genome Consortium has necessitated pruning of redundant and dependent variables. Various computational tools based on machine-learning and data-mining methods like feature selection/extraction have been proposed to escape the curse of dimensionality in large datasets. Incidentally, evolutionary studies, primarily based on sequentially evolved variations have remained un-facilitated by such advances till date. Here, we present a novel approach of recursive feature selection for hierarchical clustering of Y-chromosomal SNPs/haplogroups to select a minimal set of independent markers, sufficient to infer population structure as precisely as deduced by a larger number of evolutionary markers. To validate the applicability of our approach, we optimally designed MALDI-TOF mass spectrometry-based multiplex to accommodate independent Y-chromosomal markers in a single multiplex and genotyped two geographically distinct Indian populations. An analysis of 105 world-wide populations reflected that 15 independent variations/markers were optimal in defining population structure parameters, such as F_ST, molecular variance and correlation-based relationship. A subsequent addition of randomly selected markers had a negligible effect (close to zero, i.e. 1 × 10⁻³) on these parameters. The study proves efficient in tracing complex population structures and deriving relationships among world-wide populations in a cost-effective and expedient manner.     

Protein mapping of Chaetomium globosum,a potential biological control agent through proteomics approach

Chaetomium globosum is a ubiquitous filamentous fungus having biological control properties. The potential isolates mycoparasitize the pathogen and produce antifungal metabolites which suppress the growth of pathogenic fungi. A proteomics approach was undertaken to separate and identify proteins from a mycoparasitic strain Cg1 of C. globosum under normal and heat shock conditions in order to identify differentially expressed proteins. We developed and standardized the procedure for extraction of total proteins and 2D gel electrophoresis, which resulted in profiling of more than 100 protein spots. 48 proteins were identified by a combination of matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF) and liquid chromatography mass spectrometry (LCMS/MS). Out of total proteins identified, 79 % were hypothetical proteins and 21 % proteins were functionally characterized. Out of total 79 % hypothetical proteins 24 % proteins matched with C. globosum while 18 % proteins matched with Aspergillus spp., 13 % with Coprinopsis cinerea, 10 % with Giberrella zaea, 8 % with Magnaporthe grisea and 5 % with Neurospora crassa and Lodderomyces elonisporus. Some of the functionally characterized proteins included MAP kinase, maltose permease, GTP binding protein, dyenin heavy chain, HET- C2, vacuolar Dig A protein, polyketide synthase, peptide prolyl cis trans isomerase and translation elongation factor. This study has generated a protein reference map for Chaetomium globosum, and being the first report on proteomics studies would greatly help to unravel biocontrol mechanism and its survival under heat stress conditions.     

Association of Common Genetic Variants with Lipid Traits in the Indian Population

Genome-wide association studies (GWAS) have been instrumental in identifying novel genetic variants associated with altered plasma lipid levels. However, these quantitative trait loci have not been tested in the Indian population, where there is a poorly understood and growing burden of cardiometabolic disorders. We present the association of six single nucleotide polymorphisms in 1671 sib pairs (3342 subjects) with four lipid traits: total cholesterol, triglycerides, high density lipoprotein cholesterol (HDL-C) and low density lipoprotein cholesterol (LDL-C). We also investigated the interaction effects of gender, location, fat intake and physical activity. Each copy of the risk allele of rs964184 at APOA1 was associated with 1.06 mmol/l increase in triglycerides (SE = 0.049; p = 0.006), rs3764261 at CETP with 1.02 mmol/l increase in both total cholesterol (SE = 0.042; p = 0.017) and HDL-C (SE = 0.041; p = 0.008), rs646776 at CELSR2-PSRC1-SORT1 with 0.96 mmol/l decrease in cholesterol (SE = 0.043; p = 0.0003) and 0.15 mmol/l decrease in LDL-C levels (SE = 0.043; p = 0.0003) and rs2954029 at TRIB1 with 1.02 mmol/l increase in HDL-C (SE = 0.039; p = 0.047). A combined risk score of APOA1 and CETP loci predicted an increase of 1.25 mmol/l in HDL-C level (SE = 0.312; p = 0.0007). Urban location and sex had strong interaction effects on the genetic association of most of the studied loci with lipid traits. To conclude, we validated four genetic variants (identified by GWAS in western populations) associated with lipid traits in the Indian population. The interaction effects found here may explain the sex-specific differences in lipid levels and their heritability. Urbanization appears to influence the nature of the association with GWAS lipid loci in this population. However, these findings will require replication in other Indian populations.     

Evaluation of Publicly Financed and Privately Delivered Model of Emergency Referral Services for Maternal and Child Health Care in India

Background

Emergency referral services (ERS) are being strengthened in India to improve access for institutional delivery. We evaluated a publicly financed and privately delivered model of ERS in Punjab state, India, to assess its extent and pattern of utilization, impact on institutional delivery, quality and unit cost.

Methods

Data for almost 0.4 million calls received from April 2012 to March 2013 was analysed to assess the extent and pattern of utilization. Segmented linear regression was used to analyse month-wise data on number of institutional deliveries in public sector health facilities from 2008 to 2013. We inspected ambulances in 2 districts against the Basic Life Support (BLS) standards. Timeliness of ERS was assessed for determining quality. Finally, we computed economic cost of implementing ERS from a health system perspective.

Results

On an average, an ambulance transported 3–4 patients per day. Poor and those farther away from the health facility had a higher likelihood of using the ambulance. Although the ERS had an abrupt positive effect on increasing the institutional deliveries in the unadjusted model, there was no effect on institutional delivery after adjustment for autocorrelation. Cost of operating the ambulance service was INR 1361 (USD 22.7) per patient transported or INR 21 (USD 0.35) per km travelled.

Conclusion

Emergency referral services in Punjab did not result in a significant change in public sector institutional deliveries. This could be due to high baseline coverage of institutional delivery and low barriers to physical access. Choice of interventions for reduction in Maternal Mortality Ratio (MMR) should be context-specific to have high value for resources spent. The ERS in Punjab needs improvement in terms of quality and reduction of cost to health system.     

RNA Expression Profiling of Human iPSC-Derived Cardiomyocytes in a Cardiac Hypertrophy Model

Cardiac hypertrophy is an independent risk factor for cardiovascular disease and heart failure. There is increasing evidence that microRNAs (miRNAs) play an important role in the regulation of messenger RNA (mRNA) and the pathogenesis of various cardiovascular diseases. However, the ability to comprehensively study cardiac hypertrophy on a gene regulatory level is impacted by the limited availability of human cardiomyocytes. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) offer the opportunity for disease modeling. Here we utilize a previously established in vitro model of cardiac hypertrophy to interrogate the regulatory mechanism associated with the cardiac disease process. We perform miRNA sequencing and mRNA expression analysis on endothelin 1 (ET-1) stimulated hiPSC-CMs to describe associated RNA expression profiles. MicroRNA sequencing revealed over 250 known and 34 predicted novel miRNAs to be differentially expressed between ET-1 stimulated and unstimulated control hiPSC-CMs. Messenger RNA expression analysis identified 731 probe sets with significant differential expression. Computational target prediction on significant differentially expressed miRNAs and mRNAs identified nearly 2000 target pairs. A principal component analysis approach comparing the in vitro data with human myocardial biopsies detected overlapping expression changes between the in vitro samples and myocardial biopsies with Left Ventricular Hypertrophy. These results provide further insights into the complex RNA regulatory mechanism associated with cardiac hypertrophy.     

Induction of interleukin-2 receptor by tumor necrosis factor α on cultured ovarian tumor-associated lymphocytes

Summary We have recently reported that autologous tumor-specific cytotoxic T lymphocyte (CTL) lines and clones can be developed from lymphocytes infiltrating ovarian malignant ascites (TAL). In this study, we investigated the biological effects of tumor necrosis factor (TNF) in the induction, expansion, long-term proliferation and lytic function of CD8⁺ TAL. TNF up-regulated the IL-2 receptor (IL-2R) chain (Tac antigen) on the surface of CD3⁺ CD8⁺ CD4^– TAL, enhanced the proliferation of autologous tumor-specific CTL, and potentiated their lytic function in long-term cultures. Furthermore, in the induction and expansion phase of CD8⁺ TAL, the presence of TNF was associated with a selective increase in CD8⁺ IL-2R⁺ (Tac⁺) cells, and subsequent decrease in CD4⁺ IL-2R⁺ (Tac⁺) cells. These results suggest that the observed facilitation of the outgrowth of CD8⁺ cells in TAL cultures may be due, at least in part, to the up-regulation of IL-2R, and indicate the usefulness of TNF in the analysis of signalling in autologous tumor-reactive CTL.