Whole-genome sequencing identifies novel genes for autism in Chinese trios.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder with high genetic heritability but heterogeneity. Fully understanding its genetics requires whole-genome sequencing (WGS), but the ASD studies utilizing WGS data in Chinese population are limited. In this study, we present a WGS study for 334 individuals, including 112 ASD patients and their non-ASD parents. We identified 146 de novo variants in coding regions in 85 cases and 60 inherited variants in coding regions. By integrating these variants with an association model, we identified 33 potential risk genes (P<0.001) enriched in neuron and regulation related biological process. Besides the well-known ASD genes (SCN2A, NF1, SHANK3, CHD8 etc.), several high confidence genes were highlighted by a series of functional analyses, including CTNND1, DGKZ, LRP1, DDN, ZNF483, NR4A2, SMAD6, INTS1, and MRPL12, with more supported evidence from GO enrichment, expression and network analysis. We also integrated RNA-seq data to analyze the effect of the variants on the gene expression and found 12 genes in the individuals with the related variants had relatively biased expression. We further presented the clinical phenotypes of the proband carrying the risk genes in both our samples and Caucasian samples to show the effect of the risk genes on phenotype. Regarding variants in non-coding regions, a total of 74 de novo variants and 30 inherited variants were predicted as pathogenic with high confidence, which were mapped to specific genes or regulatory features. The number of de novo variants found in patient was significantly associated with the parents' ages at the birth of the child, and gender with trend. We also identified small de novo structural variants in ASD trios. The results in this study provided important evidence for understanding the genetic mechanism of ASD.

Tacrolimus inhibits insulin release and promotes apoptosis of Min6 cells through the inhibition of the PI3K/Akt/mTOR pathway.

As a calcineurin inhibitor, tacrolimus is commonly used as a first­line immunosuppressant in organ transplant recipients. Post­transplantation diabetes mellitus (PTDM) is a common complication following kidney transplantation and is associated with immunosuppressant drugs, such as tacrolimus. PTDM caused by tacrolimus may be related to its influence on insulin secretion and insulin resistance. However, the specific mechanism has not been fully elucidated. The aim of the present study was to investigate whether the PI3K/Akt/mTOR signaling pathway served an important role in the pathogenesis of PTDM induced by tacrolimus. In the present study, the Cell Counting Kit­8 assay was used to measure the effect of tacrolimus on the viability of Min6 mouse insulinoma cells. The effects of tacrolimus on the insulin secretion and the activity of caspase­3 of Min6 cells stimulated by glucose exposure were measured by ELISA. Superoxide dismutase (SOD) and malondialdehyde (MDA) levels were measured using WST­8 and thiobarbituric acid assays, respectively. The effects of tacrolimus on the mRNA expression levels of PI3K, Akt and mTOR were detected by reverse transcription­quantitative PCR (RT­qPCR), whereas the protein expression levels of PI3K, Akt, mTOR, phosphorylated (p)­AKT and p­mTOR in Min6 cells were assessed using western blotting. The present data indicated that, compared with the control group, 5, 25 and 50 ng/ml tacrolimus treatment could inhibit the insulin secretion of Min6 cells stimulated by glucose solution, and 50 ng/ml tacrolimus could notably decrease the stimulation index (P<0.05). Moreover, 50 ng/ml tacrolimus markedly increased the activity of caspase­3 by 175.1% (P<0.05), it also decreased the SOD activity (P<0.01) and increased MDA levels (P<0.05). The RT­qPCR results demonstrated that the mRNA expression levels of PI3K, Akt and mTOR were downregulated by 25 and 50 ng/ml tacrolimus (P<0.01). Furthermore, the western blotting results suggested that tacrolimus had no significant effects on the expression levels of total PI3K, Akt and mTOR proteins (P>0.05), but 25 and 50 ng/ml tacrolimus could significantly inhibit the expression levels of p­Akt and p­mTOR (P<0.01). In conclusion, tacrolimus decreased the activity and insulin secretion of pancreatic ß cells and induced the apoptosis of islet ß cells by inhibiting the mRNA expression levels of PI3K, Akt and mTOR and reducing the phosphorylation of Akt and mTOR proteins in the PI3K/Akt/mTOR signaling pathway, which may ultimately lead to the occurrence of diabetes mellitus, and may be considered as one of the specific mechanisms of PTDM caused by tacrolimus.

Dyadobacter luticola sp. nov., isolated from a sewage sediment sample.

A Gram-stain-negative, non-motile, aerobic bacterial strain, designated T17T was isolated from a sample of sewage sediment from a Busan park (Republic of Korea). Phylogenetic analysis based on 16S rRNA gene sequences showed that strain T17T had the highest 16S rRNA gene sequence similarity to Dyadobacter soli KCTC 22481T (97.3 %), D.yadobacter fermentans DSM 18053T (97.1 %) and D.yadobacter sediminis CGMCC 1.12895T (97.1 %). The isolate exhibited relatively low levels of DNA-DNA relatedness with respect to D. soli KCTC 22481T (28.2±3.6 %). The DNA G+C content was 49.1 mol%. The unique respiratory quinone was MK-7 and the major polar lipids were phosphatidylethanolamine, five unidentified lipids, four aminolipids, two unidentified phospholipids and one glycophospholipid. The predominant cellular fatty acids (>5 % of total) were summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH; 44.3 %), iso-C15 : 0 (15.7 %), C16 : 1ω5c (9.6 %), iso-C17 : 0 3-OH (9.3 %) and C16 : 0 (5.6 %). Moreover, physiological and biochemical characteristics distinguished strain T17T from its related species, including temperature and pH ranges for growth, being positive for acetate hydrolysis, and being negative for acid produced from melibiose and rhamnose. The genotypic, chemotaxonomic and phenotypic data revealed that strain T17T represents a novel species of the genus Dyadobacter, for which the name Dyadobacter luticola sp. nov. is proposed. The type strain is T17T (=KCTC 52981T=CCTCC AB 2017091T).

Quantification of lactate in synovia by microchip with contactless conductivity detection.

This article describes the determination of lactate in synovia by microchip capillary electrophoresis (MCE) integrated with contactless conductivity detection (CCD). The optimal running buffer consists of 10mM tris(hydroxymethyl)aminomethane, 1 mM HCl, and 0.1 mM hexadecyltrimethylammonium bromide (pH 9.1). The quantitative measurement of lactate in dilute synovia samples can be finished in less than 40s. The results indicated that the peak area had a good linear relationship with lactate concentration in the range of 20 to 1000 µM, and the correlation coefficient was 0.9984. The average recovery was from 96.6% to 106.1%, and the interday relative standard deviation was less than 4.0% (n=6). The limit of detection (signal/noise=3) reached 6.5 µM. To validate the assay results, we compared the current method with the high-performance liquid chromatography method by measuring lactate in synovia samples. The data analysis verified that there was no significant difference between the two methods. Due to significant features such as low cost, integration, and miniaturization, the MCE-CCD method may have great potential in clinical diagnosis.