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Diabetes mellitus patients with pulmonary tuberculosis (DMTB) comorbidity has been recognized as a major obstacle towards achieving the World Health Organization goal of reducing the tuberculosis incidence rate by 90% in 2035. Host immune responses affected by diabetes can lead to increased susceptibility, severity and poor treatment outcomes in DMTB patients, and the underlying mechanisms have not yet been fully elucidated. This study aimed to identify key immunological and cellular components that contribute to increased morbidity and mortality in DMTB cases.We performed RNA-Seq of total RNA isolated from peripheral blood mononuclear cells from 3 TB, 3 diabetes mellitus, and 3 DMTB patients and healthy controls, and analyzed differential expression, pathway enrichment and clustering of differentially-expressed genes (DEGs) to identify biological pathways altered specifically in DMTB patients.Bioinformatic analysis of DEGs suggested that enhanced inflammatory responses, small GTPases, the protein kinase C signaling pathway, hemostasis and the cell cycle pathway are likely implicated in the pathogenesis of the DMTB comorbidity.The DMTB comorbidity is associated with an altered transcriptome and changes in various biological pathways. Our study provides new insights on the pathological mechanism that may aid the development of host-directed therapies for this increasingly prevalent disease in high TB burden countries.

Human natural killer (NK) cells are essential for controlling infection, cancer, and fetal development. NK cell functions are modulated by interactions between polymorphic inhibitory killer cell immunoglobulin-like receptors (KIR) and polymorphic HLA-A, -B, and -C ligands expressed on tissue cells. All HLA-C alleles encode a KIR ligand and contribute to reproduction and immunity. In contrast, only some HLA-A and -B alleles encode KIR ligands and they focus on immunity. By high-resolution analysis of KIR and HLA-A, -B, and -C genes, we show that the Chinese Southern Han (CHS) are significantly enriched for interactions between inhibitory KIR and HLA-A and -B. This enrichment has had substantial input through population admixture with neighboring populations, who contributed HLA class I haplotypes expressing the KIR ligands B*46:01 and B*58:01, which subsequently rose to high frequency by natural selection. Consequently, over 80% of Southern Han HLA haplotypes encode more than one KIR ligand. Complementing the high number of KIR ligands, the CHS KIR locus combines a high frequency of genes expressing potent inhibitory KIR, with a low frequency of those expressing activating KIR. The Southern Han centromeric KIR region encodes strong, conserved, inhibitory HLA-C-specific receptors, and the telomeric region provides a high number and diversity of inhibitory HLA-A and -B-specific receptors. In all these characteristics, the CHS represent other East Asians, whose NK cell repertoires are thus enhanced in quantity, diversity, and effector strength, likely augmenting resistance to endemic viral infections.

Diabetic retinopathy (DR) is a common and serious microvascular complication of diabetes mellitus (DM), but its pathological mechanism, especially the formation mechanism of new blood vessels remains unclear. Thrombospondin-1 (Patients diagnosed with DR and DM patients without retinal problems were included in the case-control study. DNA methylation detection of150 patients diagnosed with DR and 150 DM patients without retinal complications were enrolled in the first recruitment, seven DR patients and seven DM patients were enrolled in the second recruitment. The patients with DR showed promoter hypomethylation of

Background Discrepancies can occur with the use of clinical human immunodeficiency virus (HIV) diagnostic reagents for the HIV window period (WP; time from RNA to antibody detection by diagnostic or blood screening assays). Antiretroviral therapy (ART) during acute HIV infection can impact HIV-specific antibodies, antigens, and DNA/RNA detection. In this study, an HIV WP blood donor who initiated ART was monitored, evaluating the immunological and nucleic acid testing (NAT) results for early ART and discussing the potential effects on blood safety. Study design and methods This was a follow-up study of a HIV WP donor detected 36 hours after high-risk sexual behavior, who was subsequently treated with ART. Immunological and NAT methods were comparatively analyzed. Results The 4th generation HIV serologic assays were positive at Day 11, and the 3rd generation domestic anti-HIV assay was positive at Day 33. Individual donation (ID) NAT and minipool (MP) NAT of six samples were reactive, but 12-sample MP-NAT was nonreactive. ART resulted in a slow decline of HIV RNA, but HIV DNA was still detected on Day 757. Conclusion After ART, ID-NAT was more sensitive than MP-NAT or serologic detection; however, HIV DNA detection was more sensitive, with DNA but not RNA persistently detectable.

To analyze the effect of vitamin D supplementation on the improvement of diabetes mellitus based on plasma proteomics.Five-week-old SPF spontaneously obese rats with type 2 diabetes were randomly divided into a diabetic group and a diabetic vitamin D intervention group, and the control group was Zucker lean rats. The fasting blood glucose of the rats in each group was compared with that of the diabetic vitamin D group, and the plasma proteins of the rats in each group were compared by quantitative analysis of the high-resolution mass spectrometry system iTRAQ, and KEGG signaling pathway analysis was performed.The fasting blood glucose of rats in the diabetic vitamin D intervention group was significantly lower than that of the diabetic group, and the proteins that were differentially expressed in the diabetic vitamin D intervention group were significantly improved. KEGG signaling pathway analysis revealed that the differential proteins in the diabetic group were mainly distributed among enzymes, exosomal proteins, and peptidases and inhibitors, and that the number of differences in these three classes of proteins was significantly reduced in the diabetic intervention group.Vitamin D supplementation can improve the differential expression of fasting glucose and plasma proteins in the diabetic rats.

The molecular basis of MNS blood group variants is not fully clear yet. In this study, we have characterized mRNA variants of GYPA and GYPB genes to reveal whether alternative RNA splicing may cause antigenic diversity of the MNS system.Total RNA was extracted from peripheral blood of Chinese blood donors and full-length cDNA products were generated. A nested polymerase chain reaction (PCR)-based method was established for fragment amplification and Sanger sequencing. Resulted full-length mRNA sequences were aligned with GYPA or GYPB genomic sequences respectively for exon identification. Amino acid (AA) sequences of GPA and GPB proteins were extrapolated and GYPA-EGFP, GYPB-EGFP fusion genes were generated to monitor subcellular distribution of the encoded glycophorin (GP) proteins.Totally 10 blood samples were analysed. GYPB mRNAs of all the subjects demonstrated frequent exon insertion or deletion whereas this kind of variation was only observed in 3 of 10 GYPA mRNA samples. None of the reported Miltenberger hybrids was detected in any of the mRNA samples. The alternative splicing resulted in changes of AA sequences in N-terminal domains where the MNS antigenic motifs resided; however, subcellular localizations of GP-EGFP fusion proteins showed that the above-mentioned AA changes did not affect cell surface distribution of the encoded GP proteins.Alternative RNA splicing may influence the antigenic features of GP proteins but not their cell surface distribution. Therefore, GYPA and GYPB mRNA characterization might be an invaluable supplement to serological phenotyping and DNA-based genotyping in MNS blood grouping.

Background Appropriate iodine intake for adults is essential to reduce the prevalence of thyroid diseases, but there is little research data on iodine requirement of Chinese population. This study aimed to explore the iodine requirement of young adults to maintain a healthy status based on ‘overflow theory’. Methods Iodine-balance experiment has been performed in this project. We conducted an 18-day study consisted of a 6-day acclimation period and 3 consecutive experimental stages in 37 Chinese healthy young adults (23 female and 14 male). Each stage was consumed for 4 days. Strictly-controlled low-iodine intake diets were provided for adults in the first period, an egg or 125mL milk was added in the second and third period, respectively. The dietary samples, 24-h urine specimens and faeces of volunteers were collected daily for assessment of iodine intake and excretion in volunteers. Results Mean values of iodine intake (22.7±3.6, 35.1±3.7, and 52.2±3.8μg/d), excretion (64.7±13.9, 62.3±12.6, and 94.3±14.5μg/d) and iodine balance (-35.2±19.5, -21.0±19.8, and -33.5±26.9μg/d) were significantly different among three periods for male (PP < 0.001 for all). No significant difference was observed among the 3 periods for female in the iodine balance (-30.5±9.3, -25.9±7.3, and -27.6±12.1μg/d). The linear regression equation of iodine excretion on iodine intake was Y=0.979X+37.04 (male) and Y=0.895X+31.48 (female). Compared with stage 2, iodine excretion increments in stage 3 had exceeded the iodine intake increment for men. The ratio of increment was 1.675 for male when the average iodine intake was 52.2μg/d in stage 3. When the iodine excretion increment equaled to the iodine intake increment, the daily iodine intake of men was 47.0μg. Conclusion We have evaluated the iodine requirement of young adults in southern China based on overflow theory. Our results indicate the lower limit of iodine requirement for Chinese young men is 47.0μg/d. The trial was registered at www.chictr.org.cn as ChiCTR1800014877.

Electrochemical sensors are essential for point-of-care testing (POCT) and wearable sensing devices. Establishing an efficient electron transfer route between redox enzymes and electrodes is key for converting enzyme-catalyzed reactions into electrochemical signals, and for the development of robust, sensitive, and selective biosensors. We demonstrate that the site-specific incorporation of a novel synthetic amino acid (2-amino-3-(4-mercaptophenyl)propanoic acid) into redox enzymes, followed by an S-click reaction to wire the enzyme to the electrode, facilitates electron transfer. The fabricated biosensor demonstrated real-time and selective monitoring of tryptophan (Trp) in blood and sweat samples, with a linear range of 0.02-0.8 mm. Further developments along this route may result in dramatic expansion of portable electrochemical sensors for diverse health-determination molecules.

Human natural killer (NK) cells are essential for controlling infection, cancer and fetal development. NK cell functions are modulated by interactions between polymorphic inhibitory killer cell immunoglobulin-like receptors (KIR) and polymorphic HLA-A, -B and -C ligands expressed on tissue cells. AllHLA-Calleles encode a KIR ligand and contribute to reproduction and immunity. In contrast, only someHLA-Aand-Balleles encode KIR ligands and they focus on immunity. By high-resolution analysis ofKIRandHLA-A,-Band-Cgenes, we show that the Chinese Southern Han are significantly enriched for interactions between inhibitory KIR and HLA-A and -B. This enrichment has had substantial input through population admixture with neighboring populations, who contributedHLA class Ihaplotypes expressing the KIR ligands B*46:01 and B*58:01, which subsequently rose to high frequency by natural selection. Consequently, over 80% of Southern HanHLAhaplotypes encode more than one KIR ligand. Complementing the high number of KIR ligands, the Chinese Southern HanKIRlocus combines a high frequency of genes expressing potent inhibitory KIR, with a low frequency of those expressing activating KIR. The Southern Han centromericKIRregion encodes strong, conserved, inhibitory HLA-C specific receptors, and the telomeric region provides a high number and diversity of inhibitory HLA-A and -B specific receptors. In all these characteristics, the Southern Han represent other East Asians, whose NK cell repertoires are thus enhanced in quantity, diversity and effector strength, likely through natural selection for resistance to endemic viral infections.

BACKGROUND: Due to the low incidence of concurrent human immunodeficiency virus (HIV) and syphilis infection identified during the window period, little is known about the clinical manifestations, diagnosis, and treatment efficacy at very early stages.METHODS: One longitudinal study was conducted in a 42-year-old blood donor who was concurrently infected with syphilis and HIV. This blood donor was treated with a penicillin-based regimen and early antiretroviral therapy (ART). Immunological and NAT methods were comparatively analyzed.RESULTS: A regular male donor who had two occasions of high-risk sexual behaviors 41 and 35 days before donation(DBD) donated whole blood at the Shenzhen Blood Center. Antiretroviral therapy was initiated at the 28th day after donation (DAD), and syphilis treatment was received at the 49th DAD. Microbiological analysis using a 4th generation anti-HIV ELISA and CLIA indicated a positive signal at the 6th DAD. All NAT tests for HIV RNA were reactive except the minipool (MP) NAT of six pooled samples at 117th DAD. The HIV viral load declined more than 4-log in copies per milliliter over 3 months, until reaching non-detectable levels at 246th DAD. Nevertheless, HIV-1 DNA was still detectable at 403rd DAD. Among all methods utilized, anti-TP ELISA detected syphilis infection at the earliest time. Toluidine red unheated serum test (TRUST) was negative around the 80th DAD.CONCLUSION: Concurrent infection with Syphilis and HIV during window period did not significantly change the sensitivity of reagents in detection nor alter the therapeutic efficacy for the treatment of both pathogens.

To verify a HLA-DQB1*03:90N allele and method to improve the accuracy of HLA typing.A total of 2265 hematopoietic stem cell donors from Shenzhen Branch of China Marrow Donor Program in 2018 were initially detected by a PCR sequence-specific oligonucleotide probe (SSOP) method. Among these, a rare HLA-DQB1 allele was identified by sequence-based tying (SBT) and Ion Torrent S5 next generation sequencing (NGS).The SSOP typing result suggested the HLA-DQB1 to be a rare allele, while an insertion and a deletion was suspected in its exon 2 by SBT, which were confirmed by NGS as DQB1*03:90N and DQB1*06:01, respectively.Rare alleles suspected by the SSOP method should be verified by other methods to ensure the accuracy of HLA genotyping. Rare alleles formed by deletions can be detected by NGS with accuracy.

The interaction of lipase with Ligupurpuroside B was studied by multiple spectroscopic techniques, enzyme activity and molecular modeling under simulative physiological condition. According to Stern-Volmer equation, fluorescence of lipase was quenched by Ligupurpuroside B via a static quenching mechanism because of formation of Ligupurpuroside B-lipase complex. Binding constants, number of binding sites & thermodynamic parameters were evaluated. The values of ΔGo (−25.085 kJ mol−1), ΔHo (−12.14 kJ mol−1) and ΔSo (+43.45 J mol−1 K−1) at 298 K indicated that Ligupurpuroside B-lipase interaction is spontaneous and hydrophobic interaction is the main force stabilizing the Ligupurpuroside B-lipase complex. The enzyme activity assay showed that Ligupurpuroside B inhibited lipase activity efficiently. Synchronous fluorescence spectra (SFS) suggested that Ligupurpuroside B is closer to Trp residues than to Tyr residues. All above experimental results were confirmed by molecular docking studies, which further indicated the binding site of Ligupurpuroside B on the surface of lipase, and the amino acid residues of lipase interacting with Ligupurpuroside B. Our present research work gives valuable information on the design of drugs with lipase as a carrier and should be useful for food industries.

HLA-DQB1*06:01:22 differs from HLA-DQB1*06:01:01:01 by one nucleotide substitution in codon 189 in exon 4.

Ubiquitin-specific protease 18 (USP18) as a negative regulator of the Jak/STAT signaling pathway plays an important role in the host innate immune response. USP18 has been shown to bind to the type I interferon receptor subunit 2 (IFNAR2) to down-regulate the Jak/STAT signaling. In this study, we showed that insulin receptor substrate (IRS)-4 functioned as a novel USP18-binding protein. Co-precipitation assays revealed that two regions (amino acids 335-400 and 1094-1257) of IRS4 were related to bind to the C- terminal region of USP18. IRS4 binding to USP18 diminished the inhibitory effect of USP18 on Jak/STAT signaling. IRS4 over-expression enhanced while IRS4 knock-down suppressed the Jak/STAT signaling in the presence of IFN-a stimulation. As such, IRS4 increased IFN-a-mediated anti-HCV activity. Mechanistically, IRS4 promoted the IFN-a-induced Jak/STAT signaling by interact with USP18. These results suggested that IRS4 binds to USP18 to diminish the blunting effect of USP18 on IFN-a-induced Jak/STAT signaling. Our findings indicated that IRS4 is a novel USP18-binding protein that can be used to boost the host innate immunity to control HCV, and potentially other viruses that are sensitive to IFN-a.

HLA-A*24:02:78 differs from HLA-A*24:02:01:01 in exon 3 by a single nucleotide.

Background Hepatitis B virus (HBV) infection is one of the major concerns for the safety of blood transfusion in high-prevalent countries such as in China. Prior studies outside of China have shown hepatitis B surface antigen (HBsAg) false-reactive rate of 0.02% to 0.04%. Similarly, false-negative HBsAg and HBV DNA results may occur in infected donors. Our study analyzed HBsAg enzyme-linked immunosorbent assay (ELISA)-reactive but NAT-negative donations in Shenzhen Blood Center, China. Study design and methods HBsAg ELISA-positive/NAT-negative plasma samples identified from screening 101,025 donations during 2017-2018 were analyzed by molecular and serologic tests including neutralization, chemiluminescence immunoassays, and various HBV DNA amplification assays. Molecular characterizations of HBsAg-positive/NAT-negative samples were determined by quantitative polymerase chain reaction (qPCR) and nested PCR amplification of the basic core and precore promotor regions (295 base pairs) and HBsAg (S) region (496 base pairs). Results Screening of 101,025 eligible blood donations identified 157 (0.16%, 95% confidence interval, 0.13%-0.18%) HBsAg ELISA-positive/NAT-negative plasma samples; of those, 71 (45.2%) were HBsAg confirmed positive by further HBsAg testing and DNA positive by molecular tests with increased sensitivity. Of the 71, all but one was antibody to hepatitis B core antigen reactive without antibody to hepatitis B surface antigen, yielding one recent (window-period) HBV infection. Of the remaining donations, 80 (51%) were not considered as HBV-infected donors, and 6 (3.8%) were interpreted as indeterminate since HBsAg results were discordant with unconfirmed HBV DNA results. In the 71 confirmed positives, HBsAg levels ranged from 0.05 to 400 IU/mL and HBV DNA from 6 to 2654 IU/mL; however, the correlation between the two was weak (R2 = 0.24). Conclusion Fewer than half of HBsAg ELISA-positive/NAT-negative samples were confirmed as HBsAg positive. Our study demonstrates that in highly HBV-endemic countries, assays with high sensitivity and specificity may be required.

Background Blood donor plasma samples were detected by the Ultrio Plus NAT system for HBV, HCV and HIV-1 in Shenzhen blood center, China. Reactive samples underwent further discriminatory testing of a single virus by the same methodology. A large number of cases of non-discriminated reactive (NDR) donors were found, leaving potential risk of transmitting HBV if not deferrals. This study identified those non-discriminated samples. Methods The NDR plasma samples from blood donation screening were detected and classified by additional molecular and serological tests. Molecular characterizations of DNA+ NDR were determined by sequencing analysis. Results A number of 259 (0.21%) NDR plasma samples from screening of 123,280 eligible blood donors were detected, which presented a higher rate (91.1%) of anti-HBc reactivity and nearly half (46.7%) of HBV DNA+ that classified as occult HBV infection (OBI). Most OBI strains were wild-type HBV, but some substitutions V168A, S174 N, V177A, Q129R/L/H, G145A/R in S region of genotype B (OBIB) and T47K/V/A, P49H/L, Q101R/H/K, S174 N, L175S, V177A, T118 M/R/K, G145R/A/K/E, R160K/N in S region of genotype C (OBIC) strains were identified in high frequency. Conclusion Nearly half of NDR blood samples were identified as OBI, in which a number of important mutations were detected. NDR donation might have potential risk for HBV transmission, but need to be further investigated.

To assess the association of KIR/HLA alleles with hepatocellular carcinoma (HCC) and hepatitis B virus (HBV) infection among ethnic Han Chinese patients from southern China.For 95 patients with HCC and 171 healthy controls, the genotype of HLA-C alleles was determined with a PCR sequence-specific oligonucleotides typing method on an Illumina GenDx NGSgo platform. Genotypes comprised of HLA-C and KIR gene alleles were also subjected to statistical analysis.In total 16 KIR genes (2DL2, 2DS2, 2DS3, 2DS5, 3DS1, 2DS1, 2DL5, 2DS4, 3DL1, 3DP1, 2DL3, 2DP1, 3DL3, 2DL1, 3DL2 and 2DL4) were discovered in the two groups. The frequencies of KIR2DL3 alleles and combinational genotypes of KIR2DL3/HLA-C1C2 were significantly lower in the patient group compared with the controls (0.9368 vs. 0.9883, χ²3.84; P0.05, OR = 0.1; 0.0112 vs. 0.2663, χ²3.84; P0.05, RR = 0.03). The frequency of HLA-C2C2 genotype of the patient group was significantly lower than that of the controls (0.0316 vs. 0.2690, P0.05, RR = 0.09), while the frequency of HLA-C1C2 genotype was significantly higher than that of the controls (0.2316 vs. 0.0058, P0.05, RR = 51.23).Above results suggested that the KIR2DL3 allele is associated with lower risk for HCC. There may be individual difference in patients with HCC and HBV infection but various combinations of KIR/HLA alleles.

Human natural killer (NK) cells are essential for controlling infection, cancer and fetal development. NK cell functions are modulated by interactions between polymorphic inhibitory killer cell immunoglobulin-like receptors (KIR) and polymorphic HLA-A, -B and -C ligands expressed on tissue cells. All HLA-C alleles encode a KIR ligand and contribute to reproduction and immunity. In contrast, only some HLA-A and -B alleles encode KIR ligands and they focus on immunity. By high-resolution analysis of KIR and HLA-A, -B and -C genes, we show that the Chinese Southern Han are significantly enriched for interactions between inhibitory KIR and HLA-A and -B. This enrichment has had substantial input through population admixture with neighboring populations, who contributed HLA class I haplotypes expressing the KIR ligands B*46:01 and B*58:01, which subsequently rose to high frequency by natural selection. Consequently, over 80% of Southern Han HLA haplotypes encode more than one KIR ligand. Complementing the high number of KIR ligands, the Chinese Southern Han KIR locus combines a high frequency of genes expressing potent inhibitory KIR, with a low frequency of those expressing activating KIR. The Southern Han centromeric KIR region encodes strong, conserved, inhibitory HLA-C specific receptors, and the telomeric region provides a high number and diversity of inhibitory HLA-A and -B specific receptors. In all these characteristics, the Southern Han represent other East Asians, whose NK cell repertoires are thus enhanced in quantity, diversity and effector strength, likely through natural selection for resistance to endemic viral infections.

HLA-B*15:435 has 5 nt changes from HLA-B*15:09:01 in exon 3 and 4.

To study the distribution of MICA alleles among ethnic Han Chinese blood donors from Shenzhen and their linkage disequilibrium with HLA-B gene.For 143 randomly selected blood donors, the MICA and HLA-B alleles were determined with a PCR-sequence based typing (SBT) method. Allelic frequency, haplotypic diversity and linkage disequilibrium were analyzed with a Pypop software.Thirteen MICA and 35 HLA-B alleles were identified among the 143 blood donors, among which MICA*008:01 had the highest frequency (76/286), whilst MICA*008:01-HLA-B*40:01 and MICA*010-HLA-B*46:01 were the most common haplotypes. No novel allele was identified.The allele frequencies, haplotype diversities and linkage disequilibrium parameters under a high resolution can facilitate further studies and applications of the MICA and HLA-B genes.

To list the key points for quality control during HLA-A, B, C, DRB1 and DQB1 allele typing by taking consideration of hardware, software and experimental procedures.A total of 10 167 samples from randomly selected healthy blood donors and donor-recipient pairs from Shenzhen were typed for exons 2-4 of HLA-A, B, C, exon 2 of HLA-DRB1, and exons 2 and 3 of HLA-DQB1 by PCR- sequence-based typing. For 56 cases whose forward and reverse sequences were inconsistent, the samples were re-checked by a PCR-sequence specific oligonucleotide probe method. Novel alleles not included in the IMGT/HLA database were cloned and sequenced using in-house primers.Eight novel HLA alleles were identified. A table for key positions of single nucleotide polymorphisms (SNPs) were generated, which summarized the key points for quality control during HLA-A, B, C, DRB1 and DQB1 allele typing. Among the listed SNPs, 3 were located at the HLA-A locus, 8 were at the HLA-B locus, 6 were at the C locus, 6 were at the DQB1 locus, and 4 were at the DRB1 locus. To ensure the quality control, an unique sample number for DNA transferring tubes in the process of experiment should be considered.A protocol for quality control should be enforced by checking all of the key points. The SNPs and critical control points of the alleles should be examined to ensure the accuracy of HLA typing results.

The aim of our study was to explore the role of nuclear factor erythroid 2 (NF-E2)-related factor 2 (Nrf2) on the exposure of SiOTo understand the mechanism of NP-induced oxidative stress, the involvement of oxidative-stress-responding transcription factors and the Nrf2/antioxidant reactive element (ARE) signaling pathway in the toxicity of SiOA549 cells showed a significant cytotoxic effect while A549-shNrf2 cells showed decreased cell viability after nm-SiOOur findings suggested that Nrf2 could protect against oxidative stress induced by SiO

Purpose Pathologic myopia described as myopia accompanied by severe deformation of the eye besides excessive elongation of eye, is usually a genetic heterogeneous disorder characterized by extreme, familial, early-onset vision loss. However, the exact pathogenesis of pathologic myopia remains unclear. In this study, we screened a Han Chinese family with pathologic myopia to identify the causative mutation and explore the possible pathogenic mechanism based on evaluation of the biological functions of the mutation. Methods We identified the mutations in a family with pathologic myopia by single nucleotide polymorphism array combined with short tandem repeat microsatellite marker analysis and exome sequencing. Mutations were validated among family members by direct Sanger sequencing. The subcellular localization of the protein variant was investigated by immunofluorescence, and the stability of the mutant protein was determined by immunoblotting. Intracellular levels of adenosine triphosphate and reactive oxygen species and complex I activity were measured by traditional biochemical methods to determine the functional role of the disease-associated mutation. Results The novel missense mutation: c.798C>G (p.Asp266Glu) in NDUFAF7, cosegregated with the disease and the resulting amino acid substitution affected a highly conserved residue in its protein. The mutation D266E in NDUFAF7 impaired complex I activity, which resulted in decreased ATP levels in cultured cells. Conclusions We propose that the heterozygous mutation (c.798C>G) in NDUFAF7 may contribute to the pathogenesis of pathologic myopia, possibly by interfering with the phototransduction cascade. Mitochondrial dysfunction during eye development may lead to pathologic myopia.

A pair of ruthenium(II) complex enantiomers, Δ- and Λ-[Ru(bpy)2PBIP]2+ {bpy=2,2'-bipyridine, PBIP=2-(4-bromophenyl)imidazo[4,5-f]1,10-phenanthroline} have been synthesized and characterized. The systematic comparative studies between two enantiomers on their DNA binding-behaviors with calf thymus DNA (CT DNA) were carried out by viscosity measurements, spectrophotometric methods and molecular simulation technology. Additional assays were performed to explore the cytotoxicity of the ruthenium(II) enantiomers against tumor cell lines. DNA-binding studies show that both the enantiomers can bind to CT DNA via intercalative mode, and the Δ form binds to CT DNA more strongly than the Λ form does. Molecular simulation further shows that both the two enantiomers intercalate between base pairs of DNA in minor groove, and that the Δ form intercalates into DNA more deeply than the Λ form does. In addition, the cell proliferation assays show that the Δ form induces a greater cytotoxicity than the Λ form on human cervical cancer HeLa cells, which is positive correlated with the results in DNA binding studies and molecular docking, and implies that the DNA binding affinities of ruthenium(II) polypyridyl complexes might be constitute to the part of their anticancer mechanisms.

X-linked intellectual disability (XLID) has been associated with various genes. Diagnosis of XLID, especially for non-syndromic ones (NS-XLID), is often hampered by the heterogeneity of this disease. Here we report the case of a Chinese family in which three males suffer from intellectual disability (ID). The three patients shared the same phenotype: no typical clinical manifestation other than IQ score ≤ 70. For a genetic diagnosis for this family we carried out whole exome sequencing on the proband, and validated 16 variants of interest in the genomic DNA of all the family members. A missense mutation (c.710G > T), which mapped to exon 6 of the Rab GDP-Dissociation Inhibitor 1 (GDI1) gene, was found segregating with the ID phenotype, and this mutation changes the 237th position in the guanosine diphosphate dissociation inhibitor (GDI) protein from glycine to valine (p. Gly237Val). Through molecular dynamics simulations we found that this substitution results in a conformational change of GDI, possibly affecting the Rab-binding capacity of this protein. In conclusion, our study identified a novel GDI1 mutation that is possibly NS-XLID causative, and showed that whole exome sequencing provides advantages for detecting novel ID-associated variants and can greatly facilitate the genetic diagnosis of the disease.

To identify the differential expression of membrane proteins after HaCaT cell was treated with 15 nm silica nanomaterials (SiO2).The HaCaT cells were cultured for 24 h under 15 nm SiO2 in various concentrations (2. 5, 5. 0, 10. 0 mg/L), and ddH2O were used as control. The cell viability were measured with CCK-8 assay. The membrane proteins of SiO2-treated group (10. 0 mg/L) and controls were extracted by ProteoExtract subcellular proteome extraction kit. The differentially expressed membrane proteins were analyzed by a two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-TOF-MS). Bioinformatics analysis was used to reveal the biological functions and predict transmembrane domains of differential expressed proteins. The expression of differential membrane proteins were measured by Western blot analysis.The cell viability was significantly decreased with the increases of 15 nm SiO2 exposure levels. After treatment with 2. 5, 5. 0, 10. 0 mg/L of 15nm SiO2, the cell viability was assessed to (91. 3% ± 6. 1%), (81. 7% ± 7. 0%) and (74. 0% ± 2. 6%) of control level (P0. 05), respectively. In the proteomic analysis, a total of 10 proteins were identified as differential expression in the SiO2-treated simples compared with controls. Among these, 7 of these proteins were predicted as membrane proteins with at least one significant transmembrane domain. The most dominant function that the identified proteins involved in was binding and structural molecule activity. The differential expression of G protein-coupled receptor 179 (GPR 179) and L-plastin (LCP1) were verified by Western-blot analysis under 15nm SiO2 exposure in various concentrations.The exposure of 15 nm SiO2 can significantly reduce the cell proliferation and induce a down-regulation of membrane protein expression in HaCaT cells.

To put the insight into the trichloroethylene (TCE)-induced effect on the differential expression of subcellular proteins in human normal liver cell line (L-02).The membrane proteins and nuclear proteins of TCE-treated (8.0 mmol/L) group and controls were extracted by subcellular proteome extraction kit, respectively. The TCE-induced differentially expressions were analyzed by a two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization tandem time-of-flight spectrometry (MALDI-TOF-MS). Bioinformatics analysis was used to reveal the biological processes and predict transmembrane domains of differential expressed proteins. The expression of ATP synthase subunit beta (ATP5B), heterogeneous nuclear ribonucleoprotein H2 (hnRNP H2) and far up steam element-binding protein 1 (FUBP1) were measured under TCE treatment by Western blot.After TCE treatment for 24 h in L-02 cells, 14 membrane proteins and 18 nuclear proteins were identified as differential expression. After treated with TCE in concentrations of 0, 2.0, 4.0 and 8.0 mmol/L for 24 h, the relative levels of ATP5B expression were 1.00±0.03, 1.21±0.14, 1.25±0.12 and 1.48±0.17 (F = 8.51, P = 0.007), the relative levels of hnRNP H2 expression were 1.00±0.09, 1.22±0.15, 1.43±0.21, 1.53±0.17 (F = 6.57, P = 0.015), respectively; the relative levels of FUBP1 expression were 1.00±0.11, 0.91±0.07, 0.73±0.04 and 0.67±0.03 (F = 15.81, P = 0.001), respectively, which were consistent with the results in proteomics. The bioinformatics analysis showed that the most dominant biological process were involved in RNA processing (10 proteins, P = 2.46×10(-6)), especially in RNA splicing (9 proteins, P = 1.77×10(-7)).The exposure of TCE could alter the expression of membrane proteins and nuclear proteins in L-02 cells. These abnormal expressed proteins involved in RNA splicing would provide novel clues for further understanding of TCE-induced hepatotoxicity.

To compare the DNA methylation-related alteration induced by trichloroethylene (TCE) in human hepatic L-02 cells (L-02 cells) and SET deficient cells, and reveal the role of SET on the mechanisms in TCE-induced epigenetic pathway.The L-02 cells and pre-established SET deficient cells were treated with different TCE concentrations, and the changes of total cell viability, DNA methylation level and DNA methyltransferases (DNMTs) activity were measured, respectively. In addition, the TCE-induced alteration in the protein expression of DNMT1, DNMT3a and DNMT3b were analyzed by Western blotting.After treatment with TCE for 24 h, the cell proliferation level was significantly decreased in both cell lines. When concentrations of TCE were 0, 1.0, 2.0, 4.0 and 8.0 mmol/L, the proliferation levels of L-02 cells were 100.00±2.70, 83.34±2.38, 75.56±4.51, 71.67±2.77 and 66.67±1.63, respectively (F = 58.29, P0.001); the cell proliferation levels of SET deficient cells were 101.12±1.67, 85.01±2.33, 79.44±1.67, 78.337±3.89 and 76.11±3.33, respectively (F = 42.41, P0.001). When concentration of TCE reached 4.0 mmol/L, the difference of cell proliferation level between two groups was statistically significant (t = -3.51; P = 0.013). After treated by TCE for 24 h, the global DNA methylation significantly decreased in both cell lines (F value was 212.87 and 79.32, respectively, P0.001). The difference between two groups was not statistically significant. After treated by TCE for 24 h, the methyltransferases activities were significantly decreased in both cell cells (F values were 77.92 and 113.80, respectively, P-0.001). The SET deficiency could inhibit the decrease of methyltransferases activity under TCE treatment. When the concentration of TCE reached 8.0 mmol/L, the enzymatic activity of L-02 cells and SET deficient cells decreased to 67.61%±2.85% and 72.97%± 1.94%, respectively. The difference between two groups was statistically significant (t = -3.94, P = 0.008). After treated with TCE for 24 h, concentrations of TCE were 0, 1.0, 2.0, 4.0 and 8.0 mmol/L, and the relative protein levels of DNMT1 in normal L-02 cells increased significantly to 1.00±0.03, 1.28±0.04, 1.20±0.04, 1.62±0.05, 1.43±0.04 (F = 103.00, P0.001); In SET deficient cells, the expressions of DNMT1 were 1.00±0.04, 0.96±0.02, 1.19±0.05, 0.85±0.03, 0.83±0.03, which was significantly down-regulated under TCE treatment (F = 44.18, P0.001).SET deficiency can significantly attenuate the TCE-induced decreases of cell viability and DNMTs activity, as well as alteration of protein expression of DNMT1 in L-02 cells, which indicated that SET was involved in the mechanism of TCE-induced cytotoxicity and epigenetic pathway in L-02 cells.