Aim: Type 2 Diabetes Mellitus (T2DM) is associated with both microvascular complications such as diabetic retinopathy (DR), and macrovascular complications like coronary artery disease (CAD). Genetic risk factors have a role in the development of these complications. In the present case-control study, we investigated genetic variations associated with DR and CAD in T2DM patients from the United Arab Emirates. Methods: A total of 407 Emirati patients with T2DM were recruited. Categorization of the study population was performed based on the presence or absence of DR and CAD. Seventeen Single Nucleotide Polymorphisms (SNPs), were selected for association analyses through search of publicly available databases, namely GWAS catalog, infinome genome interpretation platform and GWAS Central database. A multivariate logistic regression test was performed to evaluate the association between the 17 SNPs and DR, CAD, or both. To account for multiple testing, significance was set at p < 0.00294 using the Bonferroni correction. Results: The SNPs rs9362054 near the CEP162 gene and rs4462262 near the UBE2D1 gene were associated with DR (OR = 1.66, p = 0.001; OR = 1.37, p = 0.031; respectively), and rs12219125 near the PLXDC2 gene was associated (suggestive) with CAD (OR = 2.26, p = 0.034). Furthermore, rs9362054 near the CEP162 gene was significantly associated with both complications (OR = 2.27, p = 0.0021). The susceptibility genes for CAD (PLXDC2) and DR (UBE2D1) have a role in angiogenesis and neovascularization. Moreover, association between the ciliary gene CEP162 and DR was established in terms of retinal neural processing, confirming previous reports. Conclusions: The present study reports associations of different genetic loci with DR and CAD. We report new associations between CAD and PLXDC2, and DR with UBE2D1 using data from T2DM Emirati patients.

Dormant endospores of anaerobic, thermophilic bacteria found in cold marine sediments offer a useful model for studying microbial biogeography, dispersal, and survival. The dormant endospore phenotype confers resistance to unfavorable environmental conditions, allowing dispersal to be isolated and studied independently of other factors such as environmental selection. To study the resilience of thermospores to conditions relevant for survival in extreme cold conditions, their viability following different freezing treatments was tested. Marine sediment was frozen at either -80°C or -20°C for 10 days prior to pasteurization and incubation at +50°C for 21 days to assess thermospore viability. Sulfate reduction commenced at +50°C following both freezing pretreatments indicating persistence of thermophilic endospores of sulfate-reducing bacteria. The onset of sulfate reduction at +50°C was delayed in -80°C pretreated microcosms, which exhibited more variability between triplicates, compared to -20°C pretreated microcosms and parallel controls that were not frozen in advance. Microbial communities were evaluated by 16S rRNA gene amplicon sequencing, revealing an increase in the relative sequence abundance of thermophilic endospore-forming Firmicutes in all microcosms. Different freezing pretreatments (-80°C and -20°C) did not appreciably influence the shift in overall bacterial community composition that occurred during the +50°C incubations. Communities that had been frozen prior to +50°C incubation showed an increase in the relative sequence abundance of operational taxonomic units (OTUs) affiliated with the class Bacilli, relative to unfrozen controls. These results show that freezing impacts but does not obliterate thermospore populations and their ability to germinate and grow under appropriate conditions. Indeed the majority of the thermospore OTUs detected in this study (21 of 22) could be observed following one or both freezing treatments. These results are important for assessing thermospore viability in frozen samples and following cold exposure such as the very low temperatures that would be encountered during panspermia.

A combination of phenotype microarrays, targeted stress resistance and virulence assays and comparative genome analysis was used to compare a set of Listeria monocytogenes strains including those involved in previous Swiss foodborne listeriosis outbreaks. Despite being highly syntenic in gene content these strains showed significant phenotypic variation in utilization of different carbon (C)-sources as well as in resistance of osmotic and pH stress conditions that are relevant to host and food associated environments. An outbreak strain from the 2005 Swiss Tomme cheese listeriosis outbreak (Lm3163) showed the highest versatility in C-sources utilized whereas the strain responsible for the 1983 to 1987 Vacherin Montd'or cheese listeriosis outbreak (LL195) showed the highest tolerance to both osmotic and pH stress conditions among the examined strains. Inclusion of L-norvaline led to enhanced resistance of acidic stress in all the examined strains and there were strain-strain-specific differences observed in the ability of other amino acids and urea to enhance acid stress resistance in L. monocytogenes. A strain dependent inhibition pattern was also observed upon inclusion of β-phenylethylamine under alkaline stress conditions. In targeted phenotypic analysis the strain-specific differences in salt stress tolerance uncovered in phenotypic microarrays were corroborated and variations in host cell invasion and virulence among the examined strains were also revealed. Outbreak associated strains representing lineage I serotype 4b showed superior pathogenicity in a zebrafish infection model whilst Lm3163 a lineage II serotype 1/2a outbreak strain demonstrated the highest cellular invasion capacity amongst the tested strains. A genome wide sequence comparison of the strains only revealed few genetic differences between the strains suggesting that variations in gene regulation and expression are largely responsible for the phenotypic differences revealed among the examined strains. Our results have generated data that provides a potential basis for the future design of improved Listeria specific media to enhance routine detection and isolation of this pathogen as well as provide knowledge for developing novel methods for its control in food.

The protease high temperature requirement A from the gastric pathogen Helicobacter pylori (HtrA Hp ) belongs to the well conserved family of serine proteases. HtrA Hp is an important secreted virulence factor involved in the disruption of tight and adherens junctions during infection. Very little is known about the function of HtrA Hp in the H. pylori cell physiology due to the lack of htrA knockout strains. Here, using a newly constructed ΔhtrA mutant strain, we found that bacteria deprived of HtrA Hp showed increased sensitivity to certain types of stress, including elevated temperature, pH and osmotic shock, as well as treatment with puromycin. These data indicate that HtrA Hp plays a protective role in the H. pylori cell, presumably associated with maintenance of important periplasmic and outer membrane proteins. Purified HtrA Hp was shown to be very tolerant to a wide range of temperature and pH values. Remarkably, the protein exhibited a very high thermal stability with the melting point (Tm) values of above 85°C. Moreover, HtrA Hp showed the capability to regain its active structure following treatment under denaturing conditions. Taken together, our work demonstrates that HtrA Hp is well adapted to operate under harsh conditions as an exported virulence factor, but also inside the bacterial cell as an important component of the protein quality control system in the stressed cellular envelope.

Most microbes invading through mucosal surfaces cause disease and therefore strategies to induce mucosal immune responses are strongly needed. Vitamin A metabolites, such as retinoic acid (RA), play crucial roles in programming T and B cells to home to mucosal compartments, therefore we evaluated the capacity of RA to elicit mucosal immune responses against tuberculosis (TB) after parenteral vaccination. We found that mice immunized through subcutaneous injections with the TB subunit vaccine (CAF01+H56) in presence of RA show enhanced mucosal H56-specific IgA responses and enhanced Ag-specific CD4+ T lymphocytes homing to the lung as compared with control mice. Immunization with CAF01+H56 in presence of RA resulted in lower bacterial loads in the lungs of mice 14 days after challenge with virulent Mycobacterium tuberculosis (Mtb) as compared to mice immunized in the absence of RA or vaccinated with BCG. Higher amounts of IFNγ and IL-17 pro-inflammatory cytokines were found in lung homogenates of mice immunized with CAF01+H56 and RA 24 h after Mtb infection. However, 6 weeks after infection the protection was comparable in vaccinated mice with or without RA even though treatment with RA during immunization is able to better contain the inflammatory response by the host. Furthermore, at later stage of the infection a higher percentage of Mtb specific CD4+PD1+ T lymphocytes were found in the lungs of mice immunized with CAF01+H56 and RA. These data show that an enhanced mucosal immune response is generated during parenteral vaccination in presence of RA. Furthermore, RA treatment contained the bacterial growth at an early stage of the infection and limited the inflammatory response in the lung at later time points.

The role of metabolomics in autoimmune diseases has been a rapidly expanding area in researches over the last decade, while its pathophysiologic impact on systemic lupus erythematosus (SLE) remains poorly elucidated. In this study, we analyzed the metabolic profiling of fecal samples from SLE patients and healthy controls based on ultra-high-performance liquid chromatography equipped with mass spectrometry for exploring the potential biomarkers of SLE. The results showed that 23 differential metabolites and 5 perturbed pathways were identified between the two groups, including aminoacyl-tRNA biosynthesis, thiamine metabolism, nitrogen metabolism, tryptophan metabolism, and cyanoamino acid metabolism. In addition, logistic regression and ROC analysis were used to establish a diagnostic model for distinguishing SLE patients from healthy controls. The combined model of fecal PG 27:2 and proline achieved an area under the ROC curve of 0.846, and had a good diagnostic efficacy. In the present study, we analyzed the correlations between fecal metabolic perturbations and SLE pathogenesis. In summary, we firstly illustrate the comprehensive metabolic profiles of feces in SLE patients, suggesting that the fecal metabolites could be used as the potential non-invasive biomarkers for SLE.

Pseudomonas plecoglossicida is a temperature-dependent opportunistic pathogen which is associated with a variety of diseases in fish. During the development of "white nodules" disease, the expression of htpG in P. plecoglossicida was found to be significantly up-regulated at its virulent temperature of 18°C. The infection of htpG-RNAi strain resulted in the onset time delay, reduction in mortality and infection symptoms in spleen of Epinephelus coioides, and affected the bacterial tissue colonization. In order to reveal the effect of htpG silencing of P. plecoglossicida on the virulence regulation in P. plecoglossicida and immune response in E. coioides, dual RNA-seq was performed and a pathogen-host integration network was constructed. Our results showed that infection induced the expression of host genes related to immune response, but attenuated the expression of bacterial virulence genes. Novel integration was found between host immune genes and bacterial virulence genes, while IL6, IL1R2, IL1B, and TLR5 played key roles in the network. Further analysis with GeneMANIA indicated that flgD and rplF might play key roles during the htpG-dependent virulence regulation, which was in accordance with the reduced biofilm production, motility and virulence in htpG-RNAi strain. Meanwhile, IL6 and IL1B were found to play key roles during the defense against P. plecoglossicida, while CELA2, TRY, CPA1, CPA2, and CPB1 were important targets for P. plecoglossicida attacking to the host.

Filet quality traits determine consumer satisfaction and affect profitability of the aquaculture industry. Soft flesh is a criterion for fish filet downgrades, resulting in loss of value. Filet firmness is influenced by many factors, including rate of protein turnover. A 50K transcribed gene SNP chip was used to genotype 789 rainbow trout, from two consecutive generations, produced in the USDA/NCCCWA selective breeding program. Weighted single-step GBLUP (WssGBLUP) was used to perform genome-wide association (GWA) analyses to identify quantitative trait loci affecting filet firmness and protein content. Applying genomic sliding windows of 50 adjacent SNPs, 212 and 225 SNPs were associated with genetic variation in filet shear force and protein content, respectively. Four common SNPs in the ryanodine receptor 3 gene (RYR3) affected the aforementioned filet traits; this association suggests common mechanisms underlying filet shear force and protein content. Genes harboring SNPs were mostly involved in calcium homeostasis, proteolytic activities, transcriptional regulation, chromatin remodeling, and apoptotic processes. RYR3 harbored the highest number of SNPs (n = 32) affecting genetic variation in shear force (2.29%) and protein content (4.97%). Additionally, based on single-marker analysis, a SNP in RYR3 ranked at the top of all SNPs associated with variation in shear force. Our data suggest a role for RYR3 in muscle firmness that may be considered for genomic- and marker-assisted selection in breeding programs of rainbow trout.

Extreme phenotype sampling (EPS) is a popular study design used to reduce genotyping or sequencing costs. Assuming continuous phenotype data are available on a large cohort, EPS involves genotyping or sequencing only those individuals with extreme phenotypic values. Although this design has been shown to have high power to detect genetic effects even at smaller sample sizes, little attention has been paid to the effects of confounding variables, and in particular population stratification. Using extensive simulations, we demonstrate that the false positive rate under the EPS design is greatly inflated relative to a random sample of equal size or a "case-control"-like design where the cases are from one phenotypic extreme and the controls randomly sampled. The inflated false positive rate is observed even with allele frequency and phenotype mean differences taken from European population data. We show that the effects of confounding are not reduced by increasing the sample size. We also show that including the top principal components in a logistic regression model is sufficient for controlling the type 1 error rate using data simulated with a population genetics model and using 1,000 Genomes genotype data. Our results suggest that when an EPS study is conducted, it is crucial to adjust for all confounding variables. For genetic association studies this requires genotyping a sufficient number of markers to allow for ancestry estimation. Unfortunately, this could increase the costs of a study if sequencing or genotyping was only planned for candidate genes or pathways; the available genetic data would not be suitable for ancestry correction as many of the variants could have a true association with the trait.

Motivation: The number of ion channels is increasing rapidly. As many of them are associated with diseases, they are the targets of more than 700 drugs. The discovery of new ion channels is facilitated by computational methods that predict ion channels and their types from protein sequences. Methods: We used the SVMProt and the k-skip-n-gram methods to extract the feature vectors of ion channels, and obtained 188- and 400-dimensional features, respectively. The 188- and 400-dimensional features were combined to obtain 588-dimensional features. We then employed the maximum-relevance-maximum-distance method to reduce the dimensions of the 588-dimensional features. Finally, the support vector machine and random forest methods were used to build the prediction models to evaluate the classification effect. Results: Different methods were employed to extract various feature vectors, and after effective dimensionality reduction, different classifiers were used to classify the ion channels. We extracted the ion channel data from the Universal Protein Resource (UniProt, http://www.uniprot.org/) and Ligand-Gated Ion Channel databases (http://www.ebi.ac.uk/compneur-srv/LGICdb/LGICdb.php), and then verified the performance of the classifiers after screening. The findings of this study could inform the research and development of drugs.

DNA damage adaptation (DDA) allows the division of cells with unrepaired DNA damage. DNA repair deficient cells might take advantage of DDA to survive. The Fanconi anemia (FA) pathway repairs DNA interstrand crosslinks (ICLs), and deficiencies in this pathway cause a fraction of breast and ovarian cancers as well as FA, a chromosome instability syndrome characterized by bone marrow failure and cancer predisposition. FA cells are hypersensitive to ICLs; however, DDA might promote their survival. We present the FA-CHKREC Boolean Network Model, which explores how FA cells might use DDA. The model integrates the FA pathway with the G2 checkpoint and the checkpoint recovery (CHKREC) processes. The G2 checkpoint mediates cell-cycle arrest (CCA) and the CHKREC activates cell-cycle progression (CCP) after resolution of DNA damage. Analysis of the FA-CHKREC network indicates that CHKREC drives DDA in FA cells, ignoring the presence of unrepaired DNA damage and allowing their division. Experimental inhibition of WIP1, a CHKREC component, in FA lymphoblast and cancer cell lines prevented division of FA cells, in agreement with the prediction of the model.

The prevalent class of snake venom serine proteases (SVSP) in Viperidae venoms is the thrombin-like enzymes, which, similarly to human thrombin, convert fibrinogen into insoluble fibrin monomers. However, thrombin-like serine proteases differ from thrombin by being unable to activate factor XIII, thus leading to the formation of loose clots and fibrinogen consumption. We report the functional and biological characterization of a recombinant thrombin-like serine protease from Crotalus durissus collilineatus, named rCollinein-1.

The use of animal venoms and their toxins as material sources for biotechnological applications has received much attention from the pharmaceutical industry. L-amino acid oxidases from snake venoms (SV-LAAOs) have demonstrated innumerous biological effects and pharmacological potential against different cancer types. Hepatocellular carcinoma has increased worldwide, and the aberrant DNA methylation of liver cells is a common mechanism to promote hepatic tumorigenesis. Moreover, tumor microenvironment plays a major role in neoplastic transformation. To elucidate the molecular mechanisms responsible for the cytotoxic effects of SV-LAAO in human cancer cells, this study aimed to evaluate the cytotoxicity and the alterations in DNA methylation profiler in the promoter regions of cell-cycle genes induced by BjussuLAAO-II, an LAAO from Bothrops jaracussu venom, in human hepatocellular carcinoma (HepG2) cells in monoculture and co-culture with endothelial (HUVEC) cells.

L-Glutamate (L-Glu), the major excitatory neurotransmitter in the mammalian Central Nervous System (CNS), is essential to cognitive functions. However, when L-Glu is accumulated in large concentrations at the synaptic cleft, it can induce excitotoxicity that results in secondary damage implicated in many neurological disorders. Current therapies for the treatment of neurological disorders are ineffective and have side effects associated with their use; therefore, there is a need to develop novel treatments. In this regard, previous studies have shown that neuroactive compounds obtained from the venom of the spider Parawixia bistriata have neuroprotective effects in vitro and in vivo. In this sense, this work aimed to evaluate potential neuroprotective effects of fraction RT10, obtained from this spider venom, on primary cultures of neuron and glial cells subjected to glutamate excitotoxicity insults.

While microRNAs (miRNAs) are known to play a critical role in the progression of colorectal cancer, the role of miR-107 remains unknown. We evaluated its role and explored the underlying mechanism.

Environmental risk factors have been shown to alter DNA copy number variations (CNVs). Recently, CNVs have been described to arise after low-dose ionizing radiation in vitro and in vivo. Development of cost- and size-effective laser-driven electron accelerators (LDEAs), capable to deliver high energy beams in pico- or femtosecond durations requires examination of their biological effects. Here we studied in vitro impact of LDEAs radiation on known CNV hotspots in human peripheral blood lymphocytes on single cell level.

Rationale: Targeted delivery of therapeutic drugs or imaging agents to injured blood vessels via nanocarriers is likely to be dependent on the particle shape, yet cubic nanoparticle carriers have not been reported for vascular targeting. Here, we demonstrate that cuboidal cyclodextrin frameworks possess superior hemostasis effect and injured vessels targeting compared with spherical counterpart. Methods: Cuboidal and biocompatible γ-cyclodextrin metal-organic frameworks (CD-MOFs) are synthesized, tethered via crosslinking and surface modification with GRGDS peptide (GS5-MOFs). The specific interactions of cubic GS5-MOF nanoparticles with activated platelets were investigated by in vitro platelet aggregation assay and atomic force microscopy measurements (AFM). The hemostatic capacity and injured vessel targeting efficacy were evaluated in vivo. Results: Cuboidal GS5-MOF nanoparticles exhibit enhanced adhesion and aggregation with activated platelets in vitro under static condition and a physiologically relevant flow environment. The cubic GS5-MOF nanoparticles show efficient hemostatic effects with bleeding time and blood loss decrease of 90% and strong injured vessel targeting in vivo, markedly superior to spherical γ-CD nanosponges with the same chemical composition. Conclusions: These results clearly highlight the contribution of the cuboidal shape of GS5-MOFs to the enhanced aggregation of activated platelets and high targeting to damaged vessels. The cuboidal nanoparticle system provides an innovative delivery platform for the treatment and diagnosis of vascular diseases.

Rationale: Early and accurate detection of disease is crucial for its prevention, identification, and treatment. However, most of disease diagnostics is still limited in clinical laboratories due to the need of complicated instruments and professional personnel. Herein, we reported a smartphone-based synergistically enhanced colorimetric method for molecular diagnostics in our point of care (POC) smart cup platform. Methods: A disposable microfluidic chip was developed for colorimetric loop-mediated isothermal amplification (LAMP) detection of multiple HPV DNA in our POC smart cup platform. The colorimetric detection takes advantage of synergistic effect of PPi4- and H+ ions, two byproducts of LAMP reaction. Color signal of LAMP assay was recorded and analyzed by our custom Android app (dubbed "Hue Analyzer"). Results: Our method not only significantly improves colorimetric readout, but also provides a 10-fold increase in detection sensitivity. It has been successfully applied for HPV-associated cancer screening with spiked saliva and clinical swab samples. Conclusion: The proposed POC diagnostic platform is completely compatible with other nucleic acid biomarkers and has great potential for personalized health monitoring and disease prevention.

A stem cell-mediated bioengineered tooth root (bio-root) has proven to be a prospective tool for the treatment of tooth loss. As shown in our previous studies, dental follicle cells (DFCs) are suitable seeding cells for the construction of bio-roots. However, the DFCs which can only be obtained from unerupted tooth germ are restricted. Stem cells from human exfoliated deciduous teeth (SHEDs), which are harvested much more easily through a minimally invasive procedure, may be used as an alternative seeding cell. In this case, we compared the odontogenic characteristics of DFCs and SHEDs in bio-root regeneration. Methods: The biological characteristics of SHEDs and DFCs were determined in vitro. The cells were then induced to secrete abundant extracellular matrix (ECM) and form macroscopic cell sheets. We combined the cell sheets with treated dentin matrix (TDM) for subcutaneous transplantation into nude mice and orthotopic jaw bone implantation in Sprague-Dawley rats to further verify their regenerative potential. Results: DFCs exhibited a higher proliferation rate and stronger osteogenesis and adipogenesis capacities, while SHEDs displayed increased migration ability and excellent neurogenic potential. Both dental follicle cell sheets (DFCSs) and sheets of stem cells from human exfoliated deciduous teeth (SHEDSs) expressed not only ECM proteins but also osteogenic and odontogenic proteins. Importantly, similar to DFCSs/TDM, SHEDSs/TDM also successfully achieved the in vivo regeneration of the periodontal tissues, which consist of periodontal ligament fibers, blood vessels and new born alveolar bone. Conclusions: Both SHEDs and DFCs possessed a similar odontogenic differentiation capacity in vivo, and SHEDs were regarded as a prospective seeding cell for use in bio-root regeneration in the future.

Background: Adiponectin (ADIPOQ) is a hormone primarily synthesized by adipocytes and encoded by the ADIPOQ gene, which exerts anti-inflammatory, antiatheratogenic and insulin sensitizing functions. It has been shown that its plasma concentrations are decreased in individuals with metabolic syndrome (MS) and type 2 diabetes mellitus (DM2), which could be due to variations in the gene coding for this protein. The aim of this study was to detect the +45 T>G polymorphism of the ADIPOQ gene in subjects with DM2 and MS in Maracaibo municipality, Zulia state, Venezuela. Methods: A total of 90 subjects who attended the Center for Metabolic Endocrine Research "Dr. Félix Gómez" were enrolled for this study, 46 of which had MS-DM2 and 44 of which were healthy control individuals. Genomic DNA was extracted from blood samples and PCR-restriction fragment length polymorphism analysis was carried out for the promoter region of the ADIPOQ gene. Likewise, the +45 T> G polymorphism was identified and correlated with MS and DM2 in the studied population. Results: The most frequent allele in both groups was the T allele, and the predominant genotype was homozygous T/T (79%). Genotypes with heterozygous T/G and G/G homozygous polymorphism were more frequent in the control group than in the MS-DM2 group. Regarding the individuals with T/G and G/G genotypes, statistically significant lower mean values ​​were found for fasting glucose, total cholesterol, triacylglycerides, abdominal circumference, and for the medians of systolic and diastolic blood pressure. Odds ratio were calculated for the presence or absence of MS and DM2. Conclusions: The results suggested that the presence of the G allele exerts a protective effect on the carrier individuals, thus avoiding the appearance of the aforementioned metabolic alterations.