A comparison of type 2 diabetes risk allele load between African Americans and European Americans

The prevalence of type 2 diabetes (T2D) is greater in populations of African descent compared to European-descent populations. Genetic risk factors may underlie the disparity in disease prevalence. Genome-wide association studies (GWAS) have identified >60 common genetic variants that contribute to T2D risk in populations of European, Asian, African and Hispanic descent. These studies have not comprehensively examined population differences in cumulative risk allele load. To investigate the relationship between risk allele load and T2D risk, 46 T2D single nucleotide polymorphisms (SNPs) in 43 loci from GWAS in European, Asian, and African-derived populations were genotyped in 1,990 African Americans (n = 963 T2D cases, n = 1,027 controls) and 1,644 European Americans (n = 719 T2D cases, n = 925 controls) ascertained and recruited using a common protocol in the southeast United States. A genetic risk score (GRS) was constructed from the cumulative risk alleles for each individual. In African American subjects, risk allele frequencies ranged from 0.024 to 0.964. Risk alleles from 26 SNPs demonstrated directional consistency with previous studies, and 3 SNPs from ADAMTS9, TCF7L2, and ZFAND6 showed nominal evidence of association (p < 0.05). African American individuals carried 38–67 (53.7 ± 4.0, mean ± SD) risk alleles. In European American subjects, risk allele frequencies ranged from 0.084 to 0.996. Risk alleles from 36 SNPs demonstrated directional consistency, and 10 SNPs from BCL11A, PSMD6, ADAMTS9, ZFAND3, ANK1, CDKN2A/B, TCF7L2, PRC1, FTO, and BCAR1 showed evidence of association (p < 0.05). European American individuals carried 38–65 (50.9 ± 4.4) risk alleles. African Americans have a significantly greater burden of 2.8 risk alleles (p = 3.97 × 10^?89) compared to European Americans. However, GRS modeling showed that cumulative risk allele load was associated with risk of T2D in European Americans, but only marginally in African Americans. This result suggests that there are ethnic-specific differences in genetic architecture underlying T2D, and that these differences complicate our understanding of how risk allele load impacts disease susceptibility.     

Molecular evidence to reconcile taxonomic instability in mahseer species (Pisces: Cyprinidae) of India

The mahseers are an important group of fishes endemic to Asia with most species considered threatened. Conservation plans to save declining wild populations are hindered by unstable taxonomy, and detailed systematic review could form a solid platform for future management and conservation. D-loop and cytochrome c oxidase I (COI) mtDNA sequences were examined in nine mahseer species of Tor, Neolissochilus, and Naziritor. Pseudogenes amplified in a portion of the species limited the utility of the D-loop region. ABGD analysis, NJ, ML, and MP methods and genetic distance (TrN?+?I?+?G) using COI data revealed concordant species delimiting patterns. The three genera were monophyletic, separated as distinct clades (TrN?+?I?+?G 0.064 to 0.106), and Naziritor was flagged as a separate genus, distinct from Puntius (TrN?+?I?+?G 0.196). Out of seven nominal species known for Tor cogeners from India, only five were recovered with mtDNA data (TrN?+?I?+?G 0.000 to 0.037) and two species could not be distinguished with the molecular data set employed. Tor mosal, synonymized as Tor putitora, was rediscovered as a distinct species (TrN?+?I?+?G 0.031) based on its type locality. Tor mussulah was confirmed as a separate species (TrN?+?I?+?G 0.019 to 0.026). Two valid species, Tor macrolepis and T. mosal mahanadicus, were not distinct from T. putitora (TrN?+?I?+?G 0.00). The high divergence with mtDNA data failed to validate T. mosal mahanadicus as a subspecies of T. mosal (TrN?+?I?+?G 0.031). Morphological outliers discovered within the distribution range of Tor tor (TrN?+?I?+?G 0.022 to 0.025) shared the same lineage with T. putitora (TrN?+?I?+?G 0.002 to 0.005), indicating a new extended distribution of the Himalayan mahseer T. putitora in the rivers of the Indian central plateau. The findings indicate the need for integrating molecular and morphological tools for taxonomic revision of the Tor and Naziritor genera, so that taxa are precisely defined for accurate in situ and ex situ conservation decisions.     

Myeloid-derived suppressor cells and their role in CTLA-4 blockade therapy

Immune checkpoints are a series of inhibitory pathways that are crucial for modulating the intensity and duration of immune response. Among these checkpoints, cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) has been shown to be a key regulator of the early activation of naïve and memory T cells. Immune checkpoint blockade is emerging as one of the most promising therapeutic approaches directed toward the activation of the immune response against tumors. The first of these therapies that has been FDA approved is ipilimumab, a fully human monoclonal antibody that blocks CTLA-4. The in cis effects that CTLA-4 blockade has on T cells have been properly described, but there are still questions to be answered regarding the indirect or in trans effects. One of the alternative cellular populations that may play a role in the outcome of CTLA-4 blockade therapy is myeloid-derived suppressor cells (MDSCs), which have recently been associated with clinical outcome in advanced melanoma. In addition to this, MDSCs have been shown to be decreased in number and functional potential after treatment with ipilimumab. A better clarification of what effects CTLA-4 blockade may have on these cellular populations is likely to provide insights on possible predictive biomarkers for CTLA-4 blockade therapy.     

Distribution of ABCB1, CYP3A5, CYP2C19, and P2RY12 gene polymorphisms in a Mexican Mestizos population

The aim of the present study was to establish the gene frequency of six polymorphisms of the ABCB1, CYP3A5, CYP2C19, and P2RY12 genes in a population resident of Mexico City. The proteins encoded by these genes have been associated with the absorption, and biotransformation of clopidogrel. The ABCB1 T3435C, CYP3A5 V3* A6986G, P2RY12 G52T, P2RY12 C34T, CYP2C19 V2* and V3* (positions G681A and G636A, respectively), polymorphisms were analyzed by 5′ exonuclease TaqMan genotyping assays in a group of 269 healthy unrelated Mexican Mestizo individuals. The CYP2C19 V3* G636A polymorphism was not detected in the Mexican Mestizos population. However, the studied population presented significant differences (P < 0.05) in the distribution of the T3435C, A6986G, G681A, G52T and C34T polymorphisms when compared to reported frequencies of Amerindian of South America, Caucasian, Asian, and African populations. In summary, the distribution of the ABCB1, CYP3A5, CYP2C19, and P2RY12 gene polymorphisms distinguishes to the Mexican Mestizos population from other ethnic groups.     

Development of ARMS-PCR assay for genotyping of Pro12Ala SNP of PPARG gene: a cost effective way for case–control studies of type 2 diabetes in developing countries

Type 2 diabetes (T2D) is a prevalent metabolic disorder across the globe. Research is underway on various aspects including genetics to understand and control the global epidemic of diabetes. Recently, several SNPs in various genes have been associated with T2D. These association studies are mainly carried out in the developed countries through Genome Wide Association Scans, with follow-up replication/validation studies by high-throughput genotyping techniques (e.g. Taqman Technology). Although, similar studies could be conducted in developing countries, however, the limiting factors are the associated cost and expertise. These factors hamper research into the genetic association and replication studies from low-income countries to figure out the role of putatively associated SNPs in diabetes. Although, there are several SNP detection methods (e.g. Taqman assay, Dot-blot, PCR-RFLP, DGGE, SSCP) but these are either expensive or labor intensive or less sensitive. Hence, our aim was to develop a low-cost method for the validation of PPARG (Pro12Ala, CCA>GCA) SNP (rs1801282) for its association with T2D. Here, we developed a cost-effective and rapid amplification refractory mutation specific-PCR (ARMS-PCR) method for this SNP detection. We successfully genotyped PPARG SNPs (Pro12Ala) in human samples and the validity of this method was confirmed by DNA sequencing of a few representative samples for the three different genotypes. Furthermore, ARMS-PCR was applied to T2D patients and control samples for the screening of this SNP.     

MiR-199a is overexpressed in plasma of type 2 diabetes patients which contributes to type 2 diabetes by targeting GLUT4

Decreased GLUT4 expression and impaired GLUT4 cell membrane translocation are involved in type 2 diabetes mellitus (T2DM) pathogenesis so the factors impacting GLUT4 expression may be associated with T2DM. In this study, we identified four miRNAs: miR-31, miR-93, miR-146a, and miR-199a which suppress GLUT4 expression in HEK293T cells. Subsequently, we determined expression of these four miRNAs in plasma samples of T2DM patients, T2DM susceptible individuals, and healthy controls and found miR-199a was overexpressed in patients’ plasma compared with healthy control. Because the miR-199a binding site in GLUT4 3′UTR is highly conserved among vertebrates, we detected the glucose uptake in rat L6 myoblast cells through gain- and loss-of-function of miR-199a. We found that miR-199a can repress glucose uptake in L6 cells, which was rescued by GLUT4 overexpression. These results indicate that T2DM patients may have a high level miR-199a that reduce GLUT4 expression and contribute to the insulin resistance. Hence, miR-199a may be a novel biomarker for risk estimation and classification in T2DM patients.     

Reducing the bioactivity of Tannerella forsythia lipopolysaccharide by Porphyromonas gingivalis

Tannerella forsythia is considered a pathogen of periodontitis and forms a biofilm with multi-species bacteria in oral cavity. Lipopolysaccharide is a powerful immunostimulator and induces inflammation and shock. The purpose of this study was to investigate the characteristics of T. forsythia LPS in its co-cultivation with Fusobacterium nucleatum or Porphyromonas gingivalis. T. forsythia was co-cultured in the presence and absence of F. nucleatum and P. gingivalis and then T. forsythia LPS was extracted. The extracts were analyzed by SDS-PAGE and NF-κB reporter CHO cell lines. THP-1 cells were treated with the LPS and evaluated induction of cytokine expression by real-time RT-PCR and ELISA. For analysis of the bioactivity of T. forsythia LPS, the binding assay on LPS-binding protein (LBP) and CD14 was processed. The extracts did not contaminate other molecules except LPS and showed TLR4 agonists. Co-cultured T. forsythia LPS with P. gingivalis exhibited a lower level of induction of TNF-α, IL-1β, and IL-6 expression than singleor co-cultured T. forsythia LPS with F. nucleatum in the conditions of human serum. However, the three T. forsythia LPS did not show difference of cytokine induction in the serum free conditions. Co-cultured T. forsythia LPS with P. gingivalis exhibited a lower affinity to LBP and CD14 as binding site of O-antigen and attached at a lower level to THP-1 cells compared to single- or co-cultured T. forsythia LPS with F. nucleatum. The virulence of T. forsythia LPS was decreased by co-culturing with P. gingivalis and their affinity to LBP and CD14 was reduced, which may due to modification of O-antigen chain by P. gingivalis.     

The effects of galectin-1 on the gene expression of the transcription factors TBX21, GATA-3, FOXP3 and RORC

CD4⁺ T cells orchestrate the immune response by differentiating into T helper (Th) or regulatory (Treg) cell subsets that secrete distinct sets of cytokines. They also play a critical role in the pathogenesis of autoimmunity, asthma, allergy and, likely, cancer. The mechanisms involved in the regulation of CD4⁺ T cell homeostasis by galectin-1 remain poorly characterized. To investigate whether galectin-1 modulates the differentiation of CD4⁺ T cells, the effects of galectin-1 on the mRNA expression levels of TBX21, GATA-3, FOXP3 and RORC in activated peripheral blood mononuclear cells were examined. The expression levels of GATA-3 and FOXP3 mRNA were up-regulated after treatment with 1.0 μg/ml galectin-1 and were unchanged (for GATA-3) or slightly elevated (for FOXP3) compared with untreated cells when 2.0 μg/ml galectin-1 was added. At the same time, at both concentrations of galectin-1, we observed reduced TBX21 and RORC mRNA expression levels. These findings support the concept that galectin-1 skews the differentiation of CD4⁺ T cells towards Th2 and Treg cells.     

Differentiation of tortoises of the genera Testudo and Agrionemys (Testudinidae) based on the polymorphism of nuclear and mitochondrial markers

Based on polymorphism of the 12S rRNA gene and RAPD markers, differentiation of 122 tortoise individuals belonging to the three species of genus Testudo (T. kleinmanni, T. marginata, and T. graeca), six subspecies of T. graeca (T. g. nikolskii, T. g. pallasi, T. g. armeniaca, T. g. zarudnyi, T. g. terrestris, T. g. ibera), and two subspecies of the Central Asian tortoise Agrionenemys horsfieldii (A. h. horsfieldii, A. h. kazakhstanica) was performed. For comparison, 32 known sequences of 12S rRNA gene (392 bp) from tortoises of the two genera inhabiting the territories of Europe, Asia, and Africa were used. In the populations of A. horsfieldii, a total of six haplotypes, including three newly described variants, were identified. In the examined tortoises of the genus Testudo, eleven 12S rRNA haplotypes were identified. One new haplotype was detected in T. kleinmanni. Among the eight subspecies of T. graeca, eight haplotypes were identified, with four newly described ones. The reported RAPD markers generally supported the reconstructions obtained with the use of the mitochondrial marker. Similarly to the 12S rRNA-based reconstructions, two independent clusters included representatives of the two genera, Agrionemys and Testudio. Among the latter, representatives of T. marginata and T. kleinmanni, as well as T. graeca, with high statistical support values, formed two reciprocally monophyletic groups. Compared to the mitochondrial markers, RAPDs more statistically significantly discriminated the sample of T. g. terrestris and the four subspecies, T. g. ibera, T. g. armeniaca, T. g. pallasi, and T. g. nikolskii. In almost all cases except the representatives of T. g. ibera, the representatives of each of four subspecies formed individual subclusters. The geographical haplotype distribution patterns and possible evolutionary scenario of the origin and dispersal of tortoises of the two genera are discussed.     

Up-regulation of MDP and tuftsin gene expression in Th1 and Th17 cells as an adjuvant for an oral Lactobacillus casei vaccine against anti-transmissible gastroenteritis virus

The role of muramyl dipeptide (MDP) and tuftsin in oral immune adjustment remains unclear, particularly in a Lactobacillus casei (L. casei) vaccine. To address this, we investigated the effects of different repetitive peptides expressed by L. casei, specifically the MDP and tuftsin fusion protein (MT) repeated 20 and 40 times (20MT and 40MT), in mice also expressing the D antigenic site of the spike (S) protein of transmissible gastroenteritis virus (TGEV) on intestinal and systemic immune responses and confirmed the immunoregulation of these peptides. Treatment of mice with a different vaccine consisting of L. casei expressing MDP and tuftsin stimulated humoral and cellular immune responses. Both 20MT and 40MT induced an increase in IgG and IgA levels against TGEV, as determined using enzyme-linked immunosorbent assay. Increased IgG and IgA resulted in the activation of TGEV-neutralising antibody activity in vitro. In addition, 20MT and 40MT stimulated the differentiation of innate immune cells, including T helper cell subclasses and regulatory T (Treg) cells, which induced robust T helper type 1 and T helper type 17 (Th17) responses and reduced Treg T cell immune responses in the 20MT and 40MT groups, respectively. Notably, treatment of mice with L. casei expressing 20MT and 40MT enhanced the anti-TGEV antibody immune responses of both the humoral and mucosal immune systems. These findings suggest that L. casei expressing MDP and tuftsin possesses substantial immunopotentiating properties, as it can induce humoral and T cell-mediated immune responses upon oral administration, and it may be useful in oral vaccines against TGEV challenge.     

Knockdown of LYRM1 Rescues Insulin Resistance and Mitochondrial Dysfunction Induced by FCCP in 3T3-L1 Adipocytes

LYR motif-containing 1 (LYRM1) was recently discovered to be involved in adipose tissue homeostasis and obesity-associated insulin resistance. We previously demonstrated that LYRM1 overexpression might contribute to insulin resistance and mitochondrial dysfunction. Additionally, knockdown of LYRM1 enhanced insulin sensitivity and mitochondrial function in 3T3-L1 adipocytes. We investigated whether knockdown of LYRM1 in 3T3-L1 adipocytes could rescue insulin resistance and mitochondrial dysfunction induced by the cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP), a mitochondrion uncoupler, to further ascertain the mechanism by which LYRM1 is involved in obesity-associated insulin resistance. Incubation of 3T3-L1 adipocytes with 1 µM FCCP for 12 h decreased insulin-stimulated glucose uptake, reduced intracellular ATP synthesis, increased intracellular reactive oxygen species (ROS) production, impaired insulin-stimulated Glucose transporter type 4 (GLUT4) translocation, and diminished insulin-stimulated tyrosine phosphorylation of Insulin receptor substrate-1 (IRS-1) and serine phosphorylation of Protein Kinase B (Akt). Knockdown of LYRM1 restored insulin-stimulated glucose uptake, rescued intracellular ATP synthesis, reduced intracellular ROS production, restored insulin-stimulated GLUT4 translocation, and rescued insulin-stimulated tyrosine phosphorylation of IRS-1 and serine phosphorylation of Akt in FCCP-treated 3T3-L1 adipocytes. This study indicates that FCCP-induced mitochondrial dysfunction and insulin resistance are ameliorated by knockdown of LYRM1.     

Association between Fok I vitamin D receptor gene (VDR) polymorphism and impulsivity in alcohol-dependent patients

Vitamin D appears to have an important role in the modulation of the central nervous system. Vitamin D exerts its biological effects through its interaction with the vitamin D receptor (VDR). Located on chromosome 12 (12q13.1), the VDR gene has many different polymorphisms. Some of them are known to affect the VDR function, such as FokI (rs2228570, T/C) single nucleotide polymorphism. We aimed to explore a potential relationship between FokI VDR polymorphism and impulsiveness in alcohol-dependent (AD) patients. The study population consisted of 148 patients diagnosed with alcohol dependence (DSM-IV criteria) and 212 healthy controls. DNA was extracted from whole blood samples using the standard procedure. Genotypes were analyzed using a real-time PCR method. We found that FokI VDR gene polymorphism was associated with impulsivity [Barratt Impulsiveness Scale (BIS)-11 total score; P = 0.014], and with attentional impulsivity (BIS-11 subscale; P = 0.002) in the male AD patients. Our results suggest that CC FokI genotype of the VDR gene is associated with a higher level of impulsivity in these patients. This finding supports the hypothesis that impulsiveness, which significantly contributes to development of alcohol dependence, has a genetic background.     

Protoplast fusion enhances lignocellulolytic enzyme activities in Trichoderma reesei

Protoplast fusion was used to obtain a higher production of lignocellulolytic enzymes with protoplast fusion in Trichoderma reesei. The fusant strain T. reesei JL6 was obtained from protoplast fusion from T. reesei strains QM9414, MCG77, and Rut C-30. Filter paper activity of T. reesei JL6 increased by 18 % compared with that of Rut C-30. β-Glucosidase, hemicellulase and pectinase activities of T. reesei JL6 were also higher. The former activity was 0.39 Uml^?1, while those of QM9414, MCG77, and Rut C-30 were 0.13, 0.11, and 0.16 Uml^?1, respectively. Pectinase and hemicellulase activities of JL6 were 5.4 and 15.6 Uml^?1, respectively, which were slightly higher than those of the parents. The effects of corn stover and wheat bran carbon sources on the cellulase production and growth curve of T. reesei JL6 were also investigated.     

Environmental Heterogeneity and Microbial Inheritance Influence Sponge-Associated Bacterial Composition of Spongia lamella

Sponges are important components of marine benthic communities. High microbial abundance sponges host a large diversity of associated microbial assemblages. However, the dynamics of such assemblages are still poorly known. In this study, we investigated whether bacterial assemblages present in Spongia lamella remained constant or changed as a function of the environment and life cycle. Sponges were collected in multiple locations and at different times of the year in the western Mediterranean Sea and in nearby Atlantic Ocean to cover heterogeneous environmental variability. Co-occurring adult sponges and offsprings were compared at two of the sites. To explore the composition and abundance of the main bacteria present in the sponge mesohyl, embryos, and larvae, we applied both 16S rRNA gene-denaturing gradient gel electrophoresis (DGGE) and sequencing of excised DGGE bands and quantitative polymerase chain reactions (qPCR). On average, the overall core bacterial assemblage showed over 60 % similarity. The associated bacterial assemblage fingerprints varied both within and between sponge populations, and the abundance of specific bacterial taxa assessed by qPCR significantly differed among sponge populations and between adult sponge and offsprings (higher proportions of Actinobacteria in the latter). Sequences showed between 92 and 100 % identity to sequences previously reported in GenBank, and all were affiliated with uncultured invertebrate bacterial symbionts (mainly sponges). Sequences were mainly related to Chloroflexi and Acidobacteria and a few to Actinobacteria and Bacteroidetes. Additional populations may have been present under detection limits. Overall, these results support that both ecological and biological sponge features may shape the composition of endobiont bacterial communities in S. lamella.     

Association Between Matrix Metalloproteinase Family Gene Polymorphisms and Ischemic Stroke: a Meta-analysis

Numerous studies have studied the relationships between matrix metalloproteinase (MMP) family gene polymorphisms and ischemic stroke. However, findings remain controversial. The objective of this study was to evaluate the relationships between MMP gene polymorphisms and ischemic stroke by using a meta-analysis. The PubMed, Embase, and Cochrane library databases were systemically searched. Data were extracted by two independent reviewers, and pooled odds ratio (OR) with 95 % confidence interval (CI) were calculated. Eleven studies were enrolled, including a total of 589 cases and 494 controls of MMP-1 -1607 1G/2G; 1,817 cases and 1,731 controls of MMP-3 -1612 5A/6A; and 540 cases and 547 controls of MMP-9 -1562C/T. Under the dominant and recessive models, respectively, the overall ORs and 95 % CIs of -1607 2G were 1.54, 1.16–2.04 (P?=?0.005) and 1.25, 0.95–1.65 (P?=?0.457); the overall ORs and 95 % CIs of -1612 6A were 1.01, 0.84–1.21 (P?=?0.003) and 0.88, 0.75–1.03 (P?=?0.057); and the overall ORs and 95 % CIs of -1562T were 0.78, 0.59–1.02 (P?=?0.460) and 1.65, 0.73–3.75 (P?=?0.340). No publication bias was found in this meta-analysis. This meta-analysis showed that MMP-1 -1607 1G/2G and MMP-3 -1612 5A/6A were risk factors for ischemic stroke, while MMP-9 -1562C/T was not associated with ischemic stroke.     

Past hybridization between Quercus macrocarpa and Quercus gambelii

Introgression between two allopatric species,Quercus macrocarpa andQ. gambelii, has been observed in two separate locations: northeastern New Mexico and the Black Hills of western South Dakota and adjacent Wyoming. The probability that this introgression is the result of long-range pollination appears remote. Presumably these two species hybridized during a period of past sympatric association. Further indication of past sympatry in the Black Hills is the presence of a common species of obligate parasite, i.e., wasp(Cynips insulensis) onQ. gambelii in the Rocky Mountains and onQ. macrocarpa of the Black Hills. The hybrid oaks in New Mexico probably reflect a westward migration ofQ. macrocarpa during pluvial periods of the Pleistocene.Quercus gambelii most likely reached the Black Hills during the warmer postglacial hypsithermal era. The hybridization reported here may reflect secondary sympatry, i.e., sympatric occurrence after the species, or their ancestors, became geographically separated.