Apolipoprotein C-III (APOC3) is a key regulator of plasma triglycerides levels. Increasing evidence has shown that loss-of-function mutations in APOC3 is associated with reduction in plasma triglyceri Show more
Apolipoprotein C-III (APOC3) is a key regulator of plasma triglycerides levels. Increasing evidence has shown that loss-of-function mutations in APOC3 is associated with reduction in plasma triglycerides levels and will confer a benefit in patients at high risk for cardiovascular disease. However, these favorable mutations were extremely distribution discrepant among different ethnics. In this study, the APOC3 gene was resequenced and we identified a common variant which located in the microRNA-binding site in APOC3 and would affect its expression and the risk of coronary heart disease (CHD). The molecular mechanism was explored. We found that the T allele of rs4225 suppressed APOC3 translation by facilitating miR-4271 binding, but not the G allele. Subjects carrying the GG genotype had higher plasma APOC3 levels (p for trend = 0.03) than those with the TT genotype. Furthermore, the T allele was significantly associated with decreased triglyceride levels [Beta (SE): -0.024 (0.020), P = 0.03]. Finally, the case-control study suggested that the TT genotype resulted in a significant reduction in overall CHD risk [OR, 0.89 (95% confidence interval, 0.77-0.98), P = 0.009]. In conclusion, our results provide evidence that the rs4225 in the 3'-UTR of APOC3 might contribute to the risk of CHD by interfering with miR-4271 binding. Show less
Wnt signaling components have been shown to control key events in embryogenesis and to maintain tissue homeostasis in the adult. Nkd1/2 and Axin1/2 protein families are required for feedback regulatio Show more
Wnt signaling components have been shown to control key events in embryogenesis and to maintain tissue homeostasis in the adult. Nkd1/2 and Axin1/2 protein families are required for feedback regulation of Wnt signaling. The mechanisms by which Nkd1 and Nkd2 exhibit significant differences in signal transduction remain incompletely understood. Here we report that Rnf25/AO7, a previously identified E3 ubiquitin ligase for Nkd2, physically interacts with Nkd1 and Axin in an E3 ligase-independent manner to strengthen Wnt signalling. To determine the biological role of Rnf25 in vivo, we found that the renal mesenchymal cell, in which rnf25 was knocked-down, also exhibited more epithelial characters than MOCK control. Meanwhile, the transcriptional level of rnf25 was elevated in three separate tumor tissues more than that in paracarcinomatous tissue. Depletion of Rnf25 in zebrafish embryos attenuated transcriptions of maternal and zygotic Wnt target genes. Our results indicated that Rnf25 might serve as a molecular device, controlling the different antagonizing functions against canonical Wnt signaling between Nkd1 and Nkd2 cooperated with Axin. Show less
A novel mutation, causing a phenotype we named frogleg because its most obvious characteristic is a severe splaying of the hind limbs, arose spontaneously in a colony of Sprague-Dawley rats. Frogleg i Show more
A novel mutation, causing a phenotype we named frogleg because its most obvious characteristic is a severe splaying of the hind limbs, arose spontaneously in a colony of Sprague-Dawley rats. Frogleg is a complex phenotype that includes abnormalities in hind limb function, reduced brain weight with dilated ventricles and infertility. Using micro-satellite markers spanning the entire rat genome, the mutation was mapped to a region of rat chromosome 1 between D1Rat131 and D1Rat287. Analysis of whole genome sequencing data within the linkage interval, identified a missense mutation in the branched-chain alpha-keto dehydrogenase kinase (Bckdk) gene. The protein encoded by Bckdk is an integral part of an enzyme complex located in the mitochondrial matrix of many tissues which regulates the levels of the branched-chain amino acids (BCAAs), leucine, isoleucine and valine. BCAAs are essential amino acids (not synthesized by the body), and circulating levels must be tightly regulated; levels that are too high or too low are both deleterious. BCKDK phosphorylates Ser293 of the E1α subunit of the BCKDH protein, which catalyzes the rate-limiting step in the catabolism of the BCAAs, inhibiting BCKDH and thereby, limiting breakdown of the BCAAs. In contrast, when Ser293 is not phosphorylated, BCKDH activity is unchecked and the levels of the BCAAs will decrease dramatically. The mutation is located within the kinase domain of Bckdk and is predicted to be damaging. Consistent with this, we show that in rats homozygous for the mutation, phosphorylation of BCKDH in the brain is markedly decreased relative to wild type or heterozygous littermates. Further, circulating levels of the BCAAs are reduced by 70-80% in animals homozygous for the mutation. The frogleg phenotype shares important characteristics with a previously described Bckdk knockout mouse and with human subjects with Bckdk mutations. In addition, we report novel data regarding peripheral neuropathy of the hind limbs. Show less
CTRP4 (C1q/TNF-related protein 4) is a secreted cytokine homologous to adiponectin, which plays an important role in immunity and metabolism. This study was designed to generate CTRP4 monoclonal antib Show more
CTRP4 (C1q/TNF-related protein 4) is a secreted cytokine homologous to adiponectin, which plays an important role in immunity and metabolism. This study was designed to generate CTRP4 monoclonal antibodies (MAbs). Splenocytes extracted from mice immunized with prokaryotic protein were fused with myeloma cells Sp2/0 to generate hybridoma cells. Three hybridoma strains (16, 33, and 35) were chosen and their MAbs were purified. The specificity and affinity were identified by Western blot and enzyme-linked immunosorbent assay, whereas CTRP4 endogenous expression was identified by immunofluorescence staining. These MAbs could be useful tools for basic research and potential clinical application. Show less
Ovarian cancer is one of the most common malignancies encountered in the world. In ovarian cancer tissues of patients, NEU1 was expressed in a higher level than that in adjacent normal tissues. In thi Show more
Ovarian cancer is one of the most common malignancies encountered in the world. In ovarian cancer tissues of patients, NEU1 was expressed in a higher level than that in adjacent normal tissues. In this research, we aimed to elucidate the role of NEU1 siRNA on proliferation, apoptosis, and invasion of OVCAR3 and SKOV3 cells which expressed NEU1 notably. By cell viability assay and flow cytometry method, we found that NEU1 siRNA effectively inhibited the cancer proliferation, arrested cells cycle at G0/G1 phase, and induced apoptosis when compared to the Mock group. Result of transwell assay showed that invasion of cells in OVCAR3 and SKOV3 treated with NEU1 siRNA were suppressed significantly. Gene set enrichment analysis showed that lysosome and oxidative phosphorylation related signal pathway were associated with the NEU1 expression. In addition, Western blot revealed that expressions of Cln3 and Cln5 were depressed, and ATP5B and ATP5J expressions were also reduced. In conclusion, NEU1 siRNA can effectively inhibit proliferation, apoptosis, and invasion of human ovarian cancer cells by targeting lysosome and oxidative phosphorylation signaling, which can serve as a new target ovarian cancer treatment. Show less
Circulating blood cell counts and indices are important indicators of hematopoietic function and a number of clinical parameters, such as blood oxygen-carrying capacity, inflammation, and hemostasis. Show more
Circulating blood cell counts and indices are important indicators of hematopoietic function and a number of clinical parameters, such as blood oxygen-carrying capacity, inflammation, and hemostasis. By performing whole-exome sequence association analyses of hematologic quantitative traits in 15,459 community-dwelling individuals, followed by in silico replication in up to 52,024 independent samples, we identified two previously undescribed coding variants associated with lower platelet count: a common missense variant in CPS1 (rs1047891, MAF = 0.33, discovery + replication p = 6.38 × 10(-10)) and a rare synonymous variant in GFI1B (rs150813342, MAF = 0.009, discovery + replication p = 1.79 × 10(-27)). By performing CRISPR/Cas9 genome editing in hematopoietic cell lines and follow-up targeted knockdown experiments in primary human hematopoietic stem and progenitor cells, we demonstrate an alternative splicing mechanism by which the GFI1B rs150813342 variant suppresses formation of a GFI1B isoform that preferentially promotes megakaryocyte differentiation and platelet production. These results demonstrate how unbiased studies of natural variation in blood cell traits can provide insight into the regulation of human hematopoiesis. Show less
Excess ammonia is produced during fasting when amino acids are used for glucogenesis. Together with ureagenesis, glucogenesis occurs in periportal hepatocytes mediated mainly through the peroxisome pr Show more
Excess ammonia is produced during fasting when amino acids are used for glucogenesis. Together with ureagenesis, glucogenesis occurs in periportal hepatocytes mediated mainly through the peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). In vivo experiments showed that fasting strongly stimulated mice glucagon secretion, hepatic PGC-1α, sirtuin 3 (SIRT3) and sirtuin 5 (SIRT5) expression and ureagenesis enzymatic activity such as carbamoyl phosphate synthetase 1 (CPS1) and ornithine transcarbamoylase (OTC). Interestingly, (15)N-labeled urea and (13)C-labeled glucose production in wild-type mice were significantly increased compared with PGC-1α null mice by [(15)N,(13)C]alanine perfused liver. Glucagon significantly stimulated ureagenesis, expression of SIRT3, SIRT5 and the activities of CPS1 and OCT but did not stimulate PGC-1α silencing hepatocytes in mice periportal hepatocytes. Contrarily, PGC-1α overexpression significantly increased the expression of SIRT3, SIRT5 and the activities of CPS1 and OTC, but induced no significant changes in CPS1 and OTC expression. Morever, SIRT3 directly deacetylates and upregulates the activity of OTC, while SIRT5 deacetylates and stimulates the activity of CPS1. During fasting, PGC-1α facilitates ureagenesis in mouse periportal hepatocytes by deacetylating CPS1 and OTC modulated by mitochondrial deacetylase, SIRT3 and SIRT5. This mechanism may be relevant to ammonia detoxification and metabolic homeostasis in liver during fasting. Show less
Recent studies have shown that variants in FAT atypical cadherin 3 (FAT3), kinectin 1 (KTN1), discs large homolog2 (DLG2) and deleted in colorectal cancer (DCC) genes influence the structure of the hu Show more
Recent studies have shown that variants in FAT atypical cadherin 3 (FAT3), kinectin 1 (KTN1), discs large homolog2 (DLG2) and deleted in colorectal cancer (DCC) genes influence the structure of the human mesolimbic reward system. We conducted a systematic analysis of the potential functional single nucleotide polymorphisms (SNPs) in these genes associated with heroin addiction. We scanned the functional regions of these genes and identified 20 SNPs for genotyping by using the SNaPshot method. A total of 1080 samples, comprising 523 cases and 557 controls, were analyzed. We observed that DCC rs16956878, rs12607853, and rs2292043 were associated with heroin addiction. The T alleles of rs16956878 (p = 0.0004) and rs12607853 (p = 0.002) were significantly enriched in the case group compared with the controls. A lower incidence of the C allele of rs2292043 (p = 0.002) was observed in the case group. In block 2 of DCC (rs2292043-rs12607853-rs16956878), the frequency of the T-T-T haplotype was significantly higher in the case group than in the control group (p = 0.024), and fewer C-C-C haplotypes (p = 0.006) were detected in the case group. DCC may be an important candidate gene in heroin addiction, and rs16956878, rs12607853, and rs2292043 may be risk factors, thereby providing a basis for further genetic and biological research. Show less
This study was to determine the association between several single nucleotide polymorphisms (SNPs) in the dedicator of cytokinesis 7 (DOCK7), proprotein convertase subtilisin/kexin type 9 (PCSK9) and Show more
This study was to determine the association between several single nucleotide polymorphisms (SNPs) in the dedicator of cytokinesis 7 (DOCK7), proprotein convertase subtilisin/kexin type 9 (PCSK9) and polypeptide N-acetylgalactosaminyltransferase 2 (GALNT2) and serum lipid levels. Genotyping of 9 SNPs was performed in 881 Jing subjects and 988 Han participants. Allele and genotype frequencies of the detected SNPs were different between the two populations. Several SNPs were associated with triglyceride (TG, rs10889332, rs615563, rs7552841, rs1997947, rs2760537, rs4846913 and rs11122316), high-density lipoprotein (HDL) cholesterol (rs1997947), low-density lipoprotein (LDL) cholesterol (rs1168013 and rs7552841), apolipoprotein (Apo) A1 (rs1997947), ApoB (rs10889332 and rs7552841), and ApoA1/ApoB ratio (rs7552841) in Jing minority; and with TG (rs10889332, rs615563, rs7552841, rs11206517, rs1997947, rs4846913 and rs11122316), HDL cholesterol (rs11206517 and rs4846913), LDL cholesterol (rs1168013), ApoA1 (rs11206517 and rs4846913), ApoB (rs7552841), and ApoA1/ApoB ratio (rs4846913) in Han nationality. Strong linkage disequilibria were noted among the SNPs. The commonest haplotype was G-C-G-C-T-G-C-C-G (>10%). The frequencies of C-C-G-C-T-G-T-C-G, G-C-A-C-T-G-C-C-G, G-C-G-C-T-A-C-C-A, G-C-G-C-T-G-C-C-A, G-C-G-C-T-G-T-C-A haplotypes were different between the two populations. Haplotypes could explain much more serum lipid variation than any single SNP alone especially for TG. Differences in lipid profiles between the two populations might partially attribute to these SNPs and their haplotypes. Show less
Little is known about the association between the single nucleotide polymorphisms (SNPs) and haplotypes of the dedicator of cytokinesis 7 (DOCK7), pro-protein convertase subtilisin/kexin type 9 (PCSK9 Show more
Little is known about the association between the single nucleotide polymorphisms (SNPs) and haplotypes of the dedicator of cytokinesis 7 (DOCK7), pro-protein convertase subtilisin/kexin type 9 (PCSK9) and polypeptide N-acetylgalactosaminyltransferase 2 (GALNT2) and serum lipid traits in the Chinese populations. This study was to determine the association between nine SNPs in the three genes and their haplotypes and hypercholesterolaemia (HCH)/hypertriglyceridaemia (HTG), and to identify the possible gene-gene interactions among these SNPs. Genotyping was performed in 733 HCH and 540 HTG participants. The haplotype of C-C-G-C-T-G-C-C-G [in the order of DOCK7 rs1168013 (G>C), rs10889332 (C>T); PCSK9 rs615563 (G>A), rs7552841 (C>T), rs11206517 (T>G); and GALNT2 rs1997947 (G>A), rs2760537 (C>T), rs4846913 (C>A) and rs11122316 (G>A) SNPs] was associated with increased risk of HCH and HTG. The haplotypes of C-C-G-C-T-G-C-C-A and G-C-G-T-T-G-T-C-G were associated with a reduced risk of HCH and HTG. The haplotypes of G-C-G-C-T-G-C-C-A and G-C-G-C-T-G-T-C-G were associated with increased risk of HCH. The haplotypes of C-T-G-C-T-G-C-C-G, G-C-A-C-T-G-C-C-G and G-C-G-C-T-G-C-C-A were associated with an increased risk of HTG. The haplotypes of G-C-G-C-T-G-T-C-A and G-C-G-T-T-G-T-C-G were associated with a reduced risk of HTG. In addition, possible inter-locus interactions among the DOCK7, PCSK9 and GALNT2 SNPs were also noted. However, further functional studies of these genes are still required to clarify which SNPs are functional and how these genes actually affect the serum lipid levels. Show less
The gut microbiota plays profound roles in host metabolism and the inflammatory response associated with the development of obesity. Dusp6-deficient mice have been shown to be resistant to diet-induce Show more
The gut microbiota plays profound roles in host metabolism and the inflammatory response associated with the development of obesity. Dusp6-deficient mice have been shown to be resistant to diet-induced obesity, but the mechanism behind this remains unclear. 16S ribosomal RNA gene analysis demonstrated that dusp6-deficient mice harbour unique gut microbiota with resistance to diet-induced-obesity-mediated alteration of the gut microbiome. Using a germ-free mouse model, we found that faecal/gut microbiota derived from dusp6-deficient mice significantly increased energy expenditure and reduced weight gain in recipient wild-type mice fed on a high-fat diet. On analysis of the intestinal transcriptome of dusp6-deficient mice, we found that dusp6 deficiency mainly induced biological processes involved in metabolism and the extracellular matrix, particularly the peroxisome proliferator-activated receptor gamma (Pparγ) pathway and tight-junction genes. Furthermore, dusp6-deficient mice have a high-fat-diet-specific transcriptomic response to reverse the expression of genes associated with intestinal barrier functions and mucosal immunity involved in microbiome homeostasis. This study demonstrates that dusp6 deficiency is a strong genetic factor shaping gut microbiota, and that it confers obesity protection by ameliorating the gut microbiota response to diet-mediated stress. Show less
T cell dysfunction has a crucial role in establishing and maintaining viral persistence. We have previously shown a decline in miR-181a, which regulates CD4
Dyslipidemia is common in polycystic ovary syndrome (PCOS). This study was aimed to investigate whether fatty acid desaturase genes (FADS), a dyslipidemia-related gene cluster, are associated with PCO Show more
Dyslipidemia is common in polycystic ovary syndrome (PCOS). This study was aimed to investigate whether fatty acid desaturase genes (FADS), a dyslipidemia-related gene cluster, are associated with PCOS. We scanned variations of FADS genes using our previous data of genome-wide association study (GWAS) for PCOS and selected rs174570 for further study. The case-control study was conducted in an independent cohort of 1918 PCOS cases and 1889 age-matched controls and family-based study was conducted in a set of 243 core family trios with PCOS probands. Minor allele frequency (allele T) of rs174570 was significantly lower in PCOS cases than that in age-matched controls (P = 2.17E-03, OR = 0.85), even after adjustment of BMI and age. PCOS subjects carrying CC genotype had higher testosterone level and similar lipid/glucose level compared with those carrying TT or TC genotype. In trios, transmission disequilibrium test (TDT) analysis revealed risk allele C of rs174570 was significantly over-transmitted (P = 2.00E-04). Decreased expression of FADS2 was detected in PCOS cases and expression quantitative trait loci (eQTL) analysis revealed the risk allele C dosage was correlated with the decline of FADS2 expression (P = 0.002). Our results demonstrate that FADS1-FADS2 are susceptibility genes for PCOS. Show less
Fatty acid desaturase genes (FADS1-FADS2) encode desaturases participating in the biosynthesis of long-chain polyunsaturated fatty acids. As long-chain polyunsaturated fatty acids are implicated in ma Show more
Fatty acid desaturase genes (FADS1-FADS2) encode desaturases participating in the biosynthesis of long-chain polyunsaturated fatty acids. As long-chain polyunsaturated fatty acids are implicated in major depressive disorder (MDD) and suicide risk, and as both are partly heritable, we studied the association of FADS1-FADS2 polymorphisms with MDD (635 cases, 480 controls) and suicide attempt status (291 attempters, 344 MDD nonattempters). Eighteen FADS-related single-nucleotide polymorphisms were genotyped from Caucasians enrolled in Madrid (n=791) or New York City (n=324) and entered as predictors into logistic regression analyses with diagnostic group or suicide attempt history as outcomes and location and sex as covariates. No associations were observed between any single-nucleotide polymorphisms and diagnosis or attempt status. As statistical power was adequate, we conclude that FADS1-FADS2 genetic variants may not be a common determinant of MDD. Show less
The n-3 fatty acid desaturase gene fat1 codes for the n-3 desaturase enzyme, which can convert n-6 polyunsaturated fatty acids (PUFAs) to n-3 PUFAs. The n-3 PUFAs are essential components required for Show more
The n-3 fatty acid desaturase gene fat1 codes for the n-3 desaturase enzyme, which can convert n-6 polyunsaturated fatty acids (PUFAs) to n-3 PUFAs. The n-3 PUFAs are essential components required for normal cellular function and have preventive and therapeutic effects on many diseases. Goat is an important domestic animal for human consumption of meat and milk. To elevate the concentrations of n-3 PUFAs and examine the regulatory mechanism of fat1 in PUFA metabolism in goat cells, we successfully constructed a humanized fat1 expression vector and confirmed the efficient expression of fat1 in goat ear skin-derived fibroblast cells (GEFCs) by qRT-PCR and Western blot analysis. Fatty acid analysis showed that fat1 overexpression significantly increased the levels of total n-3 PUFAs and decreased the levels of total n-6 PUFAs in GEFCs. In addition, qRT-PCR results indicate that the FADS1 and FADS2 desaturase genes, ELOV2 and ELOV5 elongase genes, ACO and CPT1 oxidation genes, and PPARa and PPARγ transcription factors are up-regulated, and transcription factors of SREBP-1c gene are down-regulated in the fat1 transgenic goat cells. Overall, fat1-overexpression resulted in an increase in the n-3 fatty acids and altered expression of PUFA synthesis related genes in GEFCs. This work lays a foundation for both the production of fat1 transgenic goats and further study of the mechanism of fat1 function in the PUFAs metabolism. Show less
Many studies have explored whether the Notch signaling pathway has a tumor-suppressive or an oncogenic role in various tumors; however, the role of the Notch signaling pathway in salivary adenoid cyst Show more
Many studies have explored whether the Notch signaling pathway has a tumor-suppressive or an oncogenic role in various tumors; however, the role of the Notch signaling pathway in salivary adenoid cystic carcinoma (SACC) is still unknown. In this study, we attempt to define the role of Notch2 signaling in cell growth, invasion, and migration in SACC. We compared Notch2 expression in clinical SACC samples with that of normal samples by using immunohistochemical staining. Then, we down-regulated Notch2 expression to observe the effect of Notch2 on proliferation, invasion, migration, and the expression of known target genes of Notch signal pathway. According to our results, Notch2 expression was higher in SACC tissues compared with normal tissues. Knockdown of Notch2 inhibited cell proliferation, invasion, and migration in vitro and down-regulated the expression of HEY2 and CCND1. The results of this study suggest that Notch2 has an essential role in the cell growth, invasion, and migration of SACC. Notch2 may therefore be a potential target gene for the treatment of SACC by interfering with cell growth and metastasis. Show less
Two ongoing phase II clinical trials (RENEW and SYNERGY) have been developed to test the efficacy of anti-LINGO-1 antibodies in acute optic neuritis and relapsing forms of multiple sclerosis, respecti Show more
Two ongoing phase II clinical trials (RENEW and SYNERGY) have been developed to test the efficacy of anti-LINGO-1 antibodies in acute optic neuritis and relapsing forms of multiple sclerosis, respectively. Across a range of experimental models, LINGO-1 has been found to inhibit neuron and oligodendrocyte survival, axon regeneration, and (re)myelination. The therapeutic effects of anti-LINGO-1 antibodies on optic nerve axonal loss and regeneration have not yet been investigated. In this series of studies we investigate if LINGO-1 antibodies can prevent acute inflammatory axonal loss, and promote axonal regeneration after injury in rodent optic nerves. The effects of anti-LINGO-1 antibody on optic nerve axonal damage were assessed using rodent myelin oligodendrocyte glycoprotein experimental autoimmune encephalomyelitis (EAE), and its effects on axonal regeneration were assessed in optic nerve crush injury models. In the optic nerve, anti-LINGO-1 antibody therapy was associated with improved optic nerve parallel diffusivity measures on MRI in mice with EAE and reduced axonal loss in rat EAE. Both anti-LINGO-1 antibody therapy and the genetic deletion of LINGO-1 reduced nerve crush-induced axonal degeneration and enhanced axonal regeneration. These data demonstrate that LINGO-1 blockade is associated with axonal protection and regeneration in the injured optic nerve. Show less
The precise molecular alterations driving castration-resistant prostate cancer (CRPC) are not clearly understood. Using a novel network-based integrative approach, here, we show distinct alterations i Show more
The precise molecular alterations driving castration-resistant prostate cancer (CRPC) are not clearly understood. Using a novel network-based integrative approach, here, we show distinct alterations in the hexosamine biosynthetic pathway (HBP) to be critical for CRPC. Expression of HBP enzyme glucosamine-phosphate N-acetyltransferase 1 (GNPNAT1) is found to be significantly decreased in CRPC compared with localized prostate cancer (PCa). Genetic loss-of-function of GNPNAT1 in CRPC-like cells increases proliferation and aggressiveness, in vitro and in vivo. This is mediated by either activation of the PI3K-AKT pathway in cells expressing full-length androgen receptor (AR) or by specific protein 1 (SP1)-regulated expression of carbohydrate response element-binding protein (ChREBP) in cells containing AR-V7 variant. Strikingly, addition of the HBP metabolite UDP-N-acetylglucosamine (UDP-GlcNAc) to CRPC-like cells significantly decreases cell proliferation, both in-vitro and in animal studies, while also demonstrates additive efficacy when combined with enzalutamide in-vitro. These observations demonstrate the therapeutic value of targeting HBP in CRPC. Show less
Diagnostic next-generation sequencing (NGS)-based gene panels are increasingly used for prevalent disorders with genetic and clinical heterogeneity. Clinical development, validation, and quality manag Show more
Diagnostic next-generation sequencing (NGS)-based gene panels are increasingly used for prevalent disorders with genetic and clinical heterogeneity. Clinical development, validation, and quality management of these panels ideally includes reference samples containing prevalent pathogenic variants; however, clinical domain expertise to select appropriate variants may not be present, samples are often not publicly available, and their inclusion is associated with added cost. Expert-designed, multiplexed controls can remedy some of these challenges. One approach relies on spiking biosynthetic fragments carrying desired variants into human genomic DNA. We piloted the utility of this approach for hypertrophic cardiomyopathy. Data from >3000 previously sequenced probands were used to select 10 common pathogenic and/or technically challenging variants in the top hypertrophic cardiomyopathy genes. Multiplexed controls were constructed across a range of ideal and realistic allelic fractions for heterozygous germline variants. NGS was performed in quadruplicate, and results were compared with diagnostic NGS data for the source patient samples. Overall, results were indistinguishable from patient-derived data with variants being detected at or reasonably close to the targeted allelic fraction ratios. The exception was a common 25-bp deletion in MYBPC3, underscoring the importance of including such variants in test development. These controls may be an attractive addition to the repertoire of materials for development, validation, and quality monitoring of clinical NGS assays. Show less
Yu-Bing Dai, Yi-Fei Miao, Wan-Fu Wu+8 more · 2016 · Proceedings of the National Academy of Sciences of the United States of America · National Academy of Sciences · added 2026-04-24
The etiology of peripheral squamous cell lung cancer (PSCCa) remains unknown. Here, we show that this condition spontaneously develops in mice in which the genes for two oxysterol receptors, Liver X R Show more
The etiology of peripheral squamous cell lung cancer (PSCCa) remains unknown. Here, we show that this condition spontaneously develops in mice in which the genes for two oxysterol receptors, Liver X Receptor (LXR) α (Nr1h3) and β (Nr1h2), are inactivated. By 1 y of age, most of these mice have to be euthanized because of severe dyspnea. Starting at 3 mo, the lungs of LXRα,β(Dko) mice, but not of LXRα or LXRβ single knockout mice, progressively accumulate foam cells, so that by 1 y, the lungs are covered by a "golden coat." There is infiltration of inflammatory cells and progressive accumulation of lipid in the alveolar wall, type 2 pneumocytes, and macrophages. By 14 mo, there are three histological lesions: one resembling adenomatous hyperplasia, one squamous metaplasia, and one squamous cell carcinoma characterized by expression of transformation-related protein (p63), sex determining region Y-box 2 (Sox2), cytokeratin 14 (CK14), and cytokeratin 13 (CK13) and absence of thyroid transcription factor 1 (TTF1), and prosurfactant protein C (pro-SPC). RNA sequencing analysis at 12 mo confirmed a massive increase in markers of M1 macrophages and lymphocytes. The data suggest a previously unidentified etiology of PSCCa: cholesterol dysregulation and M1 macrophage-predominant lung inflammation combined with damage to, and aberrant repair of, lung tissue, particularly the peripheral parenchyma. The results raise the possibility that components of the LXR signaling may be useful targets in the treatment of PSCCa. Show less
Perfluorooctane sulfonate (PFOS), one persistent organic pollutant, has been widely detected in the environment, wildlife and human. Currently few studies have documented the effects of chronic PFOS e Show more
Perfluorooctane sulfonate (PFOS), one persistent organic pollutant, has been widely detected in the environment, wildlife and human. Currently few studies have documented the effects of chronic PFOS exposure on lipid metabolism, especially in aquatic organisms. The underlying mechanisms of hepatotoxicity induced by chronic PFOS exposure are still largely unknown. The present study defined the effects of chronic exposure to low level of PFOS on lipid metabolism using zebrafish as a model system. Our findings revealed a severe hepatic steatosis in the liver of males treated with 0.5μM PFOS as evidenced by hepatosomatic index, histological assessment and liver lipid profiles. Quantitative PCR assay further indicated that PFOS significantly increase the transcriptional expression of nuclear receptors (nr1h3, rara, rxrgb, nr1l2) and the genes associated with fatty acid oxidation (acox1, acadm, cpt1a). In addition, chronic PFOS exposure significantly decreased liver ATP content and serum level of VLDL/LDL lipoprotein in males. Taken together, these findings suggest that chronic PFOS exposure induces hepatic steatosis in zebrafish via disturbing lipid biosynthesis, fatty acid β-oxidation and excretion of VLDL/LDL lipoprotein, and also demonstrate the validity of using zebrafish as an alternative model for PFOS chronic toxicity screening. Show less
Recent studies have shown that activation of liver X receptors (LXRs) attenuates the development of atherosclerosis, not only by regulating lipid metabolism but also by suppressing inflammatory signal Show more
Recent studies have shown that activation of liver X receptors (LXRs) attenuates the development of atherosclerosis, not only by regulating lipid metabolism but also by suppressing inflammatory signaling. Sphingosine 1-phosphate receptor 2 (S1PR2), an important inflammatory gene product, plays a role in the development of various inflammatory diseases. It was proposed that S1PR2 might be regulated by LXR-α. In the present study, the effect of LXR-α on tumor necrosis factor-α (TNF-α)-induced S1PR2 expression in human umbilical vein endothelial cells (HUVECs) was investigated and the underlying mechanism was explored. The results demonstrated that TNF-α led to an increase in S1PR2 expression and triggered a downregulation of LXR-α expression in HUVECs as well. Downregulation of LXR-α with specific small interfering RNA (siRNA) remarkably enhanced the primary as well as TNF-α-induced expression of S1PR2 in HUVECs. Activation of LXR-α by agonist GW3965 inhibited both primary and TNF-α-induced S1PR2 expression. GW3965 also attenuated S1PR2-induced endothelial barrier dysfunction. The data further showed that TNF-α induced a significant decrease in miR-130a-3p expression. Overexpression of miR-130a-3p with mimic product reduced S1PR2 protein expression, and inhibition of miR-130a-3p by specific inhibitor resulted in an increase in S1PR2 protein expression. Furthermore, activation of LXRs with agonist enhanced the expression of miR-130a-3p, and knockdown of LXR-α by siRNA suppressed miR-130a-3p expression. These results suggest that LXR-α might downregulate S1PR2 expression via miR-130a-3p in quiescent HUVECs. Stimulation of TNF-α attenuates the activity of LXR-α and results in enhanced S1PR2 expression. Show less
Adaptation to hypoxia is an important process physiologically and pathologically. Hypoxia-inducible factor-1α (HIF-1α) participates in the cancer biology of numerous endocrine tumors, including their Show more
Adaptation to hypoxia is an important process physiologically and pathologically. Hypoxia-inducible factor-1α (HIF-1α) participates in the cancer biology of numerous endocrine tumors, including their proliferation and differentiation. In the present study, the hypothesis that HIF-1α promotes tumorigenesis in thyroid cancer via upregulating angiogenesis-associated markers is investigated. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis were used to examine the expression of HIF-1α in thyroid cancer cell lines, and to detect the expression of WW domain containing E3 ubiquitin protein ligase (WWP)2, WWP9, vascular endothelial growth factor (VEGF) and VEGF receptor 2 (VEGFR2) in MZ-CRC-1 and TT thyroid cancer cells. Cell proliferation was measured using a Cell Count Kit-8. Cell apoptosis and cell cycle was assessed by flow cytometry. Cell invasive ability was examined by Matrigel transwell analysis. RT-qPCR and western blot analyses demonstrated that the mRNA and protein expression levels of HIF-1α were significant higher in MZ-CRC-1 and TT thyroid cancer cells than in another three thyroid cancer cells (P<0.01). HIF-1α knockdown cells demonstrated inhibition of cell proliferation and invasion, arrested cell cycle at the G1 phase, and induction of cell apoptosis. The protein expression levels of WWP2, WWP9, VEGF and VEGFR2 were decreased in HIF-1α knockdown MZ-CRC-1 and TT cells. In conclusion, HIF-1α may be important in cell apoptosis and invasion of thyroid cancer cells, likely through regulating WWP2, WWP9, VEGF and VEGFR2 expression. Show less
Lung cancer has been a hot area of research because of its high incidence and mortality. In this study, WWP2, an E3 ubiquitin ligase, is proposed to be an oncoprotein contributing to lung tumorigenesi Show more
Lung cancer has been a hot area of research because of its high incidence and mortality. In this study, WWP2, an E3 ubiquitin ligase, is proposed to be an oncoprotein contributing to lung tumorigenesis. We attempted to determine if WWP2 gene expression is correlated with the development of human lung adenocarcinoma. Real-time PCR and western blotting were used to detect the expression of WWP2 in 65 paired lung adenocarcinoma and adjacent normal lung tissues. We found that WWP2 expression was elevated in lung adenocarcinoma tissues and was correlated with the tumor differentiation stage, TNM stage and presence of lymph node metastasis. We performed CCK-8 and colony formation assays and found that down-regulation of WWP2 inhibited proliferation in A549 and SPC-A-1 cells. A wound healing assay and trans-well invasion assays showed that down-regulation of WWP2 inhibited the migration and invasion of lung adenocarcinoma cells. It could be predicted from these data that elevated expression of WWP2 may play a role in facilitating the development of lung adenocarcinoma. Show less
We determined the alleles of ten single nucleotide poly-morphisms (SNPs) in the APOA5/A4/C3/A1 gene cluster and in APOB in Han Chinese from Xinjiang Shihezi, China using MALDI-TOF mass spectrometry, a Show more
We determined the alleles of ten single nucleotide poly-morphisms (SNPs) in the APOA5/A4/C3/A1 gene cluster and in APOB in Han Chinese from Xinjiang Shihezi, China using MALDI-TOF mass spectrometry, and explored the correlation between these SNPs and dyslipidemia through a case-control study design with 250 pa-tients and 250 normal controls. All SNPs except for APOA5 rs2072560 conformed to Hardy-Weinberg equilibrium (all P > 0.05). APOA5 rs651821, APOA4 rs5104, APOC3 rs734104, and APOC3 rs5128 geno-type and allele frequencies were significantly different between groups (all P < 0.01). For rs651821, the risks of dyslipidemia for the CC or CC+CT genotypes were 9.917 or 1.859 times that of TT, and the risk of the C vs T allele was 2.027. For rs5104, the AG, GG, or AG+GG risks were 1.797, 1.861, and 1.809 times AA, and the G vs A risk was 1.427. For rs734104, the CT, CC, or CC+CT risks were 1.851, 2.570, and 1.958 times TT, and the C vs T risk was 1.610. For rs5128, the GC or CC+GC risks were 1.738 or 1.749 times GG, and the C vs G risk was 1.477. Compared with the wild-type haplotype TATG, the risks of dyslipidemia with CGCC, TGCC, or CATG haplotypes (odds ratios = 2.434, 1.503, and 2.740, respectively) were significantly higher. Our results suggested that these four SNPs were significantly associated with dyslipidemia in Xinjiang Shihezi Han Chinese, and might serve as risk factors for dyslipidemia. Individuals carrying the CGCC, TGCC, or CATG haplotypes were prone to dyslipidemia. Show less
Severe hypertriglyceridemia is a well-known cause of pancreatitis. Usually, there is a moderate increase in plasma triglyceride level during pregnancy. Additionally, certain pre-existing genetic trait Show more
Severe hypertriglyceridemia is a well-known cause of pancreatitis. Usually, there is a moderate increase in plasma triglyceride level during pregnancy. Additionally, certain pre-existing genetic traits may render a pregnant woman susceptible to development of severe hypertriglyceridemia and pancreatitis, especially in the third trimester. To elucidate the underlying mechanism of gestational hypertriglyceridemic pancreatitis, we undertook DNA mutation analysis of the lipoprotein lipase (LPL), apolipoprotein C2 (APOC2), apolipoprotein A5 (APOA5), lipase maturation factor 1 (LMF1), and glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 (GPIHBP1) genes in five unrelated pregnant Chinese women with severe hypertriglyceridemia and pancreatitis. DNA sequencing showed that three out of five patients had the same homozygous variation, p.G185C, in APOA5 gene. One patient had a compound heterozygous mutation, p.A98T and p.L279V, in LPL gene. Another patient had a compound heterozygous mutation, p.A98T & p.C14F in LPL and GPIHBP1 gene, respectively. No mutations were seen in APOC2 or LMF1 genes. All patients were diagnosed with partial LPL deficiency in non-pregnant state. As revealed in our study, genetic variants appear to play an important role in the development of severe gestational hypertriglyceridemia, and, p.G185C mutation in APOA5 gene appears to be the most common variant implicated in the Chinese population. Antenatal screening for mutations in susceptible women, combined with subsequent interventions may be invaluable in the prevention of potentially life threatening gestational hypertriglyceridemia-induced pancreatitis. Show less
Myocardial infarction (MI), a leading cause of death around the world, displays a complex pattern of inheritance. When MI occurs early in life, genetic inheritance is a major component to risk. Previo Show more
Myocardial infarction (MI), a leading cause of death around the world, displays a complex pattern of inheritance. When MI occurs early in life, genetic inheritance is a major component to risk. Previously, rare mutations in low-density lipoprotein (LDL) genes have been shown to contribute to MI risk in individual families, whereas common variants at more than 45 loci have been associated with MI risk in the population. Here we evaluate how rare mutations contribute to early-onset MI risk in the population. We sequenced the protein-coding regions of 9,793 genomes from patients with MI at an early age (≤50 years in males and ≤60 years in females) along with MI-free controls. We identified two genes in which rare coding-sequence mutations were more frequent in MI cases versus controls at exome-wide significance. At low-density lipoprotein receptor (LDLR), carriers of rare non-synonymous mutations were at 4.2-fold increased risk for MI; carriers of null alleles at LDLR were at even higher risk (13-fold difference). Approximately 2% of early MI cases harbour a rare, damaging mutation in LDLR; this estimate is similar to one made more than 40 years ago using an analysis of total cholesterol. Among controls, about 1 in 217 carried an LDLR coding-sequence mutation and had plasma LDL cholesterol > 190 mg dl(-1). At apolipoprotein A-V (APOA5), carriers of rare non-synonymous mutations were at 2.2-fold increased risk for MI. When compared with non-carriers, LDLR mutation carriers had higher plasma LDL cholesterol, whereas APOA5 mutation carriers had higher plasma triglycerides. Recent evidence has connected MI risk with coding-sequence mutations at two genes functionally related to APOA5, namely lipoprotein lipase and apolipoprotein C-III (refs 18, 19). Combined, these observations suggest that, as well as LDL cholesterol, disordered metabolism of triglyceride-rich lipoproteins contributes to MI risk. Show less
Cilia harbor sensory receptors for various signaling cascades critical for vertebrate development. However, the mechanisms underlying the ciliary homeostasis of sensory receptors remain elusive. Here, Show more
Cilia harbor sensory receptors for various signaling cascades critical for vertebrate development. However, the mechanisms underlying the ciliary homeostasis of sensory receptors remain elusive. Here, we demonstrate that BBS-4 and BBS-5, two distinct BBSome components, show unexpected functional redundancy in the context of cilia in C. elegans. BBS-4 directly interacts with BBS-5 and the interaction can be disrupted by a conserved mutation identified in human BBS4. Surprisingly, we found that BBS-4 and BBS-5 act redundantly in the BBSome to regulate the ciliary removal, rather than the ciliary entry or retrograde IFT transport, of various sensory receptors. Further analyses indicate that co-depletion of BBS-4 and BBS-5 disrupts the lysosome-targeted degradative sorting of ciliary sensory receptors. Moreover, mammalian BBS4 and BBS5 also interact directly and coordinate the ciliary removal of polycystin 2. Hence, we reveal a novel and highly conserved role for the BBSome in fine-tuning ciliary signaling by regulating the ciliary removal of sensory receptors for lysosomal degradation. Show less
Non-obstructive azoospermia (NOA), a severe form of male infertility, is often suspected to be linked to currently undefined genetic abnormalities. To explore the genetic basis of this condition, we s Show more
Non-obstructive azoospermia (NOA), a severe form of male infertility, is often suspected to be linked to currently undefined genetic abnormalities. To explore the genetic basis of this condition, we successfully sequenced ~650 infertility-related genes in 757 NOA patients and 709 fertile males. We evaluated the contributions of rare variants to the etiology of NOA by identifying individual genes showing nominal associations and testing the genetic burden of a given biological process as a whole. We found a significant excess of rare, non-silent variants in genes that are key epigenetic regulators of spermatogenesis, such as BRWD1, DNMT1, DNMT3B, RNF17, UBR2, USP1 and USP26, in NOA patients (P = 5.5 × 10(-7)), corresponding to a carrier frequency of 22.5% of patients and 13.7% of controls (P = 1.4 × 10(-5)). An accumulation of low-frequency variants was also identified in additional epigenetic genes (BRDT and MTHFR). Our study suggested the potential associations of genetic defects in genes that are epigenetic regulators with spermatogenic failure in human. Show less