👤 Jiahui Deng

Binding and uptake of triglyceride-rich lipoproteins (TRLs) in mice depend on heparan sulfate and the hepatic proteoglycan, syndecan-1 (SDC1). Alteration of glucosamine N-sulfation by deletion of glucosamine N-deacetylase-N-sulfotransferase 1 (Ndst1) and 2-O-sulfation of uronic acids by deletion of uronyl 2-O-sulfotransferase (Hs2st) led to diminished lipoprotein metabolism, whereas inactivation of glucosaminyl 6-O-sulfotransferase 1 (Hs6st1), which encodes one of the three 6-O-sulfotransferases, had little effect on lipoprotein binding. However, other studies have suggested that 6-O-sulfation may be important for TRL binding and uptake. In order to explain these discrepant findings, we used CRISPR/Cas9 gene editing to create a library of mutants in the human hepatoma cell line, Hep3B. Inactivation of EXT1 encoding the heparan sulfate copolymerase, NDST1 and HS2ST dramatically reduced binding of TRLs. Inactivation of HS6ST1 had no effect, but deletion of HS6ST2 reduced TRL binding. Compounding mutations in HS6ST1 and HS6ST2 did not exacerbate this effect indicating that HS6ST2 is the dominant 6-O-sulfotransferase and that binding of TRLs indeed depends on 6-O-sulfation of glucosamine residues. Uptake studies showed that TRL internalization was also affected in 6-O-sulfation deficient cells. Interestingly, genetic deletion of SDC1 only marginally impacted binding of TRLs but reduced TRL uptake to the same extent as treating the cells with heparin lyases. These findings confirm that SDC1 is the dominant endocytic proteoglycan receptor for TRLs in human Hep3B cells and that binding and uptake of TRLs depend on SDC1 and N- and 2-O-sulfation as well as 6-O-sulfation of heparan sulfate chains catalyzed by HS6ST2. Show less

Glucose-dependent insulinotropic polypeptide (GIP) receptor (GIPR) has been identified in multiple genome-wide association studies (GWAS) as a contributor to obesity, and GIPR knockout mice are protected against diet-induced obesity (DIO). On the basis of this genetic evidence, we developed anti-GIPR antagonistic antibodies as a potential therapeutic strategy for the treatment of obesity and observed that a mouse anti-murine GIPR antibody (muGIPR-Ab) protected against body weight gain, improved multiple metabolic parameters, and was associated with reduced food intake and resting respiratory exchange ratio (RER) in DIO mice. We replicated these results in obese nonhuman primates (NHPs) using an anti-human GIPR antibody (hGIPR-Ab) and found that weight loss was more pronounced than in mice. In addition, we observed enhanced weight loss in DIO mice and NHPs when anti-GIPR antibodies were codosed with glucagon-like peptide-1 receptor (GLP-1R) agonists. Mechanistic and crystallographic studies demonstrated that hGIPR-Ab displaced GIP and bound to GIPR using the same conserved hydrophobic residues as GIP. Further, using a conditional knockout mouse model, we excluded the role of GIPR in pancreatic β-cells in the regulation of body weight and response to GIPR antagonism. In conclusion, these data provide preclinical validation of a therapeutic approach to treat obesity with anti-GIPR antibodies. Show less

BACKGROUND Angiopoietin-like 4 (ANGPTL4) is neuroprotective when administered acutely for the treatment of cerebral ischemia. The aim of the present study was to evaluate the preventive effects of ANGPTL4 on the formation of brain edema and to determine whether it promotes the recovery of neurological function following intracerebral hemorrhage (ICH). MATERIAL AND METHODS Recombinant human ANGPTL4 (rhANGPTL4; 40 µg/kg) or a vehicle was administered intraperitoneally 5 min prior to bacterial collagenase-induced ICH in male C57/B6J mice. Behavioral tests were performed prior to ICH and at days 1, 3, 7, 14, 21, and 28 after ICH. Brain edema and hematoma volume were examined separately using the wet weight/dry weight method and hematoxylin-eosin staining. The integrity of the tight and adherens junctions was quantified via immunofluorescence. The ultrastructure of the blood-brain barrier (BBB) was examined using transmission electron microscopy. Vascular endothelial (VE)-cadherin, claudin-5, Src, and phospho-Src in the ipsilateral and contralateral striatum were detected by Western blot analysis. RESULTS RhANGPTL4 reduced brain edema and hematoma volume and improved neurological functional recovery over the subsequent 4 weeks when compared with the control group. rhANGPTL4 significantly increased VE-cadherin and claudin-5-positive areas and relative amounts in the peri‑hematoma region compared with the control group. In addition, ANGPTL4 significantly reduced the ratio of phospho-Src to Src. The significant reduction of Src kinase activity in the peri‑hematoma region of ANGPTL-treated mice was paralleled by a decrease in vascular permeability and edema formation. CONCLUSIONS These results suggest that ANGPTL4 is a relevant target for vasculoprotection and cerebral protection during stroke. Show less

The traditional Chinese medicine (TCM) Yuanshi Shengmai Chenggu Tablet is used for treating the common orthopedic disease, hormone‑induced avascular necrosis of the femoral head (ANFH) in China. However, its underlying mechanism and the changes induced in the treatment of ANFH remain to be fully elucidated. In the present study, through the use of isobaric Tag for Relative and Absolute Quantitation and multiple reaction monitoring quantifications, corticosteroid‑induced femoral head necrosis and the effects of treatment with Yuanshi Shengmai Chenggu Tablet were examined. The aim was to identify serum proteins, which may be potential serum markers for the early clinical diagnosis of ANFH, and maybe used to develop more rapid and convenient detection strategies. A total of five proteins were identified, comprising Ig mu chain C region, keratin, type I cytoskeletal 9, properdin, apolipoprotein A‑IV, and IQ and AAA domain‑containing protein 1. The expression levels of all five proteins were lower in ANFH and were higher following TCM treatment. These findings were confirmed using ELISA and western blot analysis. Show less

Wnt/β-catenin signaling activity is maintained in homeostasis by an expanding list of molecular determinants. However, the molecular components and the regulatory mechanisms involved in its fine-tuning remain to be determined. Here, we identified C9orf140, a tumor-specific protein, as a novel Axin1-interacting protein by tandem-affinity purification and mass spectrometry. We further showed that C9orf140 is a negative regulator of Wnt/β-catenin signaling in cultured cells as well as in zebrafish embryos. It functions upstream of β-catenin, outcompetes PP2A for binding to Axin1, influences the balance between phosphorylation and de-phosphorylation of β-catenin, and ultimately compromises Wnt3A-induced β-catenin accumulation. Interestingly, Wnt-induced C9orf140 expression via β-catenin. We propose that C9orf140 mediates a negative feedback loop of Wnt/β-catenin signaling by interacting with Axin1. Our results advance the current understanding of the exquisite control of Wnt/β-catenin signaling cascade, and provide evidence of the new role of C9orf140. Show less

Accumulating evidence has suggested a link between maternal di-(2-ethylhexyl)-phthalate (DEHP) exposure and various developmental abnormalities. However, the evidence regarding the effect of maternal DEHP exposure on fetal cardiac development is scarce. The present study aimed to determine the effect of maternal DEHP exposure on fetal cardiac development in mice and explore the possible involved mechanism preliminarily. The C57BL mice were randomly divided into four groups: the vehicle group (corn oil, n = 10), 250 mg kg Show less

Tyrosine kinase inhibitors (TKi) are effective against chronic myeloid leukemia (CML), but their inefficacy on leukemia stem cells (LSCs) may lead to relapse. To identify new druggable targets alternative to BCR/ABL, we investigated the role of the MEK5/ERK5 pathway in LSC maintenance in low oxygen, a feature of bone marrow stem cell niches. We found that MEK5/ERK5 pathway inhibition reduced the growth of CML patient-derived cells and cell lines in vitro and the number of leukemic cells in vivo. Treatment in vitro of primary CML cells with MEK5/ERK5 inhibitors, but not TKi, strikingly reduced culture repopulation ability (CRA), serial colony formation ability, long-term culture-initiating cells (LTC-ICs), and CD26-expressing cells. Importantly, MEK5/ERK5 inhibition was effective on CML cells regardless of the presence or absence of imatinib, and did not reduce CRA or LTC-ICs of normal CD34+ cells. Thus, targeting MEK/ERK5 may represent an innovative therapeutic approach to suppress CML progenitor/stem cells. Show less

Objective Given its effects on lipid metabolism, the apolipoprotein A1-C3-A5 ( APOA1-C3-A5) gene cluster is thought to play an important role in ischemic stroke pathogenesis. Here, we evaluated whether the APOA1-C3-A5 cluster is associated with ischemic stroke in the northern Chinese Han population. Methods This case-control study analyzed 812 patients with ischemic stroke and 844 healthy controls with regard to four APOA1-C3-A5 cluster promoter single nucleotide polymorphisms (SNPs), rs670, rs2854116, rs2854117, and rs662799, using the SNaPshot Multiplex sequencing assay. Potential associations among ischemic stroke, genotyping, and allele frequencies were assessed. Results APOA1 rs670 CT/TT genotypes, APOA5 rs662799 AG/GG genotypes, and the APOC3 rs2854116 CC genotype were associated with an increased risk of ischemic stroke according to multivariate logistic analysis after adjusting for confounding factors. A significantly increased risk for ischemic stroke was also identified among high-risk haplotypes (C-C-T-A and T-T-C-A) for rs670-rs2854116-rs2854117-rs662799. Conclusion This study showed that rs670, rs2854116, and rs662799 SNPs of the APOA1-C3-A5 cluster are associated with ischemic stroke in the northern Chinese Han population. Show less

Observational studies suggest associations of triglyceride levels with longevity and frailty. This study aimed to test whether the associations are causal. We used data from the Rugao Longevity and Ageing Study, a population-based cohort study performed in Rugao, China. A variant in the APOA5 gene region (rs662799) was used as the genetic instrument. Mendelian randomization (MR) analyses were performed to examine the associations of genetically predicted triglycerides with two ageing phenotypes - longevity ( ≥95 years) and frailty (modified Fried frailty phenotype and Rockwood frailty index). C allele of rs662799 was robustly associated with higher triglyceride levels in the comparison group (β = 0.301 mmol/L per allele, p < 0.001), with an F statistic of 95.3 and R Show less

Genome-wide association studies (GWAS) on Parkinson's disease (PD) have mostly been done in Europeans and Japanese. No study has been done in Han Chinese, which make up nearly a fifth of the world population. We conducted the first Han Chinese GWAS analysing a total of 22,729 subjects (5,125 PD cases and 17,604 controls) from Singapore, Hong Kong, Malaysia, Korea, mainland China and Taiwan. We performed imputation, merging and logistic regression analyses of 2,402,394 SNPs passing quality control filters in 779 PD cases, 13,227 controls, adjusted for the first three principal components. 90 SNPs with association P < 10-4 were validated in 9 additional sample collections and the results were combined using fixed-effects inverse-variance meta-analysis. We observed strong associations reaching genome-wide significance at SNCA, LRRK2 and MCCC1, confirming their important roles in both European and Asian PD. We also identified significant (P < 0.05) associations at 5 loci (DLG2, SIPA1L2, STK39, VPS13C and RIT2), and observed the same direction of associations at 9 other loci including BST1 and PARK16. Allelic heterogeneity was observed at LRRK2 while European risk SNPs at 6 other loci including MAPT and GBA-SYT11 were non-polymorphic or very rare in our cohort. Overall, we replicate associations at SNCA, LRRK2, MCCC1 and 14 other European PD loci but did not identify Asian-specific loci with large effects (OR > 1.45) on PD risk. Our results also demonstrate some differences in the genetic contribution to PD between Europeans and Asians. Further pan-ethnic meta-analysis with European GWAS cohorts may unravel new PD loci. Show less

17β-Hydroxysteroid dehydrogenase type 3 (17β-HSD3) is expressed almost exclusively in the testes and specifically converts the weak androgenic androstenedione to active testosterone (T) in the presence of NADPH. Additionally, studies have demonstrated that 17β-HSD3 is over-expressed in hormone-dependent prostate cancer. T, which interacts with the androgen receptor (AR), eventually stimulates the growth of prostate cancer cells. Defects in T synthesis or action impair the development of the male phenotype during embryogenesis and cause the autosomal recessive disorder male pseudohermaphroditism. Affected individuals are often born with female-appearing external genitalia and are reared as females. Since 17β-HSD3 plays a central role in T production, it has been recognized as a promising therapeutic target to reduce the circulating level of androgens and to suppress androgen-sensitive tumor proliferation. In recent decades, improvements have been made in the development of 17β-HSD3 inhibitors. Herein, we give an overview of the main structure and function of human 17β-HSD3 and summarize steroidal and non-steroidal inhibitors of 17β-HSD3, which can be a potential target for prostate cancer. Show less

Human colorectal cancer stem cells (CSCs) are tumour initiating cells that can self-renew and are highly tumorigenic and chemoresistant. While genetic mutations associated with human colorectal cancer development are well-known, little is known about how and whether epigenetic factors specifically contribute to the functional properties of human colorectal CSCs. Here we report that the KDM3 family of histone demethylases plays an important role in tumorigenic potential and survival of human colorectal CSCs by epigenetically activating Wnt target gene transcription. The depletion of KDM3 inhibits tumorigenic growth and chemoresistance of human colorectal CSCs. Mechanistically, KDM3 not only directly erases repressive H3K9me2 marks, but also helps to recruit histone methyltransferase MLL1 to promote H3K4 methylation, thereby promoting Wnt target gene transcription. Our results suggest that KDM3 is a critical epigenetic factor in Wnt signalling that orchestrates chromatin changes and transcription in human colorectal CSCs, identifying potential therapeutic targets for effective elimination of CSCs. Show less

Esophageal cancer (EC) is the most lethal cancer, and it is of significant concern worldwide, particularly in China. However, there are no effective treatments to cure it, such as chemotherapy, surgery, or radiotherapy. This is attributed to the lack of understanding of the molecular mechanisms of EC. Recently, the superfamily of Jmj-containing KDMs has been shown to play an important role in tumorigenesis in various cancers, including EC. In this study, we demonstrated that JMJD1C was upregulated in patient EC tissues and different EC cell lines. Furthermore, JMJD1C levels were positively correlated with the TNM stage. Moreover, the colony formation assay, CCK8, and cell number count assay showed that the knockdown of JMJD1C inhibited EC cell proliferation. Western blot analysis and the quantitative real-time polymerase chain reaction assay showed that the knockdown of JMJD1C repressed the protein and mRNA levels of YAP1 via regulating the H3K9me2 activity, but not the H3K9me1 activity. The colony formation assay, CCK8 analysis, and cell number count assay revealed that inhibition of EC cell proliferation by the knockdown of JMJD1C was rescued by overexpression of YAP1. Taken together, our results demonstrated that JMJD1C controls the proliferation of EC via modulation of H3K9me2 activity, targeting the YAP1 gene expression and functions as a tumor suppressor in EC. This novel pathway may serve as a therapeutic target for EC patients. Show less

A lack of sufficient oligodendrocyte myelination contributes to remyelination failure in demyelinating disorders. miRNAs have been implicated in oligodendrogenesis; however, their functions in myelin regeneration remained elusive. Through developmentally regulated targeted mutagenesis, we demonstrate that miR-219 alleles are critical for CNS myelination and remyelination after injury. Further deletion of miR-338 exacerbates the miR-219 mutant hypomyelination phenotype. Conversely, miR-219 overexpression promotes precocious oligodendrocyte maturation and regeneration processes in transgenic mice. Integrated transcriptome profiling and biotin-affinity miRNA pull-down approaches reveal stage-specific miR-219 targets in oligodendrocytes and further uncover a novel network for miR-219 targeting of differentiation inhibitors including Lingo1 and Etv5. Inhibition of Lingo1 and Etv5 partially rescues differentiation defects of miR-219-deficient oligodendrocyte precursors. Furthermore, miR-219 mimics enhance myelin restoration following lysolecithin-induced demyelination as well as experimental autoimmune encephalomyelitis, principal animal models of multiple sclerosis. Together, our findings identify context-specific miRNA-regulated checkpoints that control myelinogenesis and a therapeutic role for miR-219 in CNS myelin repair. Show less

Chronic treatment with second-generation antipsychotic drugs (SGAs) has been associated with an increased risk of metabolic syndrome. To evaluate the longitudinal changes in glucose-lipid homeostasis after SGA use, we studied the time-dependent effects of olanzapine (OLZ) (3 mg/kg, b.i.d.) or clozapine (CLZ) (20 mg/kg, b.i.d.) treatment on metabolic profiles for 9 weeks in rats. Although only OLZ significantly increased body weight in rats, both OLZ and CLZ elevated blood lipid levels. Chronic OLZ treatment induced significant weight gain leading to a higher fasting insulin level and impaired glucose tolerance, whereas CLZ lowered fasting insulin levels and impaired glucose tolerance independent of weight gain. Treatment with both drugs deranged AKT/GSK phosphorylation and up-regulated muscarinic M3 receptors in the rats' livers. Consistent with an elevation in lipid levels, both OLZ and CLZ significantly increased the protein levels of nuclear sterol regulatory element-binding proteins (SREBPs) in the liver, which was associated with improvement in hepatic histamine H1R. However, enhanced carbohydrate response element binding protein (ChREBP) signalling was observed in only CLZ-treated rats. These results suggest that SGA-induced glucose-lipid metabolic disturbances could be independent of weight gain, possibly through activation of SREBP/ChREBP in the liver. Show less

Although lots of genes have been revealed to relate to sporadic amyotrophic lateral sclerosis (sALS), its genetic mechanisms still need to be further explored. We aimed to search the novel genetic factors of sALS and assess their contribution. We constructed an integrative dataset based on the 3227 subsignificant genes (P value < 0.01) from two sALS-related genome-wide association studies (GWAS) (the US and Irish studies). A significant replication between both studies was confirmed by the gene set enrichment analysis in the integral level (P value < 10 Show less

Moringa oleifera is a promising plant species for oil and forage, but its genetic improvement is limited. Our current breeding program in this species focuses on exploiting the functional genes associated with important agronomical traits. Here, we screened reliable reference genes for accurately quantifying the expression of target genes using the technique of real-time quantitative polymerase chain reaction (RT-qPCR) in M. oleifera. Eighteen candidate reference genes were selected from a transcriptome database, and their expression stabilities were examined in 90 samples collected from the pods in different developmental stages, various tissues, and the roots and leaves under different conditions (low or high temperature, sodium chloride (NaCl)- or polyethyleneglycol (PEG)- simulated water stress). Analyses with geNorm, NormFinder and BestKeeper algorithms revealed that the reliable reference genes differed across sample designs and that ribosomal protein L1 (RPL1) and acyl carrier protein 2 (ACP2) were the most suitable reference genes in all tested samples. The experiment results demonstrated the significance of using the properly validated reference genes and suggested the use of more than one reference gene to achieve reliable expression profiles. In addition, we applied three isotypes of the superoxide dismutase (SOD) gene that are associated with plant adaptation to abiotic stress to confirm the efficacy of the validated reference genes under NaCl and PEG water stresses. Our results provide a valuable reference for future studies on identifying important functional genes from their transcriptional expressions via RT-qPCR technique in M. oleifera. Show less

Hepatic apolipoprotein A-IV (apoA-IV) expression is correlated with hepatic triglyceride (TG) content in mouse models of chronic hepatosteatosis, and steatosis-induced hepatic apoA-IV gene expression is regulated by nuclear transcription factor cAMP-responsive element-binding protein H (CREBH) processing. To define what aspects of TG homeostasis regulate hepatic CREBH processing and apoA-IV gene expression, several mouse models of attenuated VLDL particle assembly were subjected to acute hepatosteatosis induced by an overnight fast or short term ketogenic diet feeding. Compared with chow-fed C57BL/6 mice, fasted or ketogenic diet-fed mice displayed increased hepatic TG content, which was highly correlated (r Show less

The apolipoprotein C3 (APOC3) gene, which is a member of the APOA1/C3/A4/A5 gene cluster, plays a crucial role in lipid metabolism. Dyslipidemia is an important risk factor for ischemic stroke. In the present study, we performed a hospital-based case-control study of 895 ischemic stroke patients and 883 control subjects to examine the effects of four APOC3 single nucleotide polymorphisms (SNPs) (rs2854116, rs2854117, rs4520 and rs5128) on the risk of ischemic stroke in a northern Chinese Han population. The SNaPshot Multiplex sequencing assay was used for SNP genotyping, and the potential association of genotype distributions and allele frequencies with ischemic stroke was analyzed statistically. Compared with the GG genotype, the CC+GC genotype of rs5128 was significantly associated with an increased risk in females (adjusted OR = 3.38, 95% CI = 1.82-6.28, P <0.01) after all of the risk factors were adjusted for with logistic regression analyses. A similar relationship was found between the rs4520 polymorphism and ischemic stroke risk in Han Chinese women. Under a recessive genetic model, the TT+TC genotypes of this variant increased ischemic stroke risk (adjusted OR = 2.05; 95% CI = 1.28-3.29; P <0.01). Haplotype analysis revealed that in males, the T-C-T-C haplotype of rs2854116-rs2854117-rs4520-rs5128 was significantly more frequent in the ischemic stroke group than in the control group (OR = 1.49, 95% CI = 1.18-1.87, P<0.01). The results of our study indicate that the APOC3 polymorphisms contribute to ischemic stroke susceptibility in females in the northern Chinese Han population. Show less

Previous work identified RMEL3 as a lncRNA with enriched expression in melanoma. Analysis of The Cancer Genome Atlas (TCGA) data confirmed RMEL3 enriched expression in melanoma and demonstrated its association with the presence of BRAFV600E. RMEL3 siRNA-mediated silencing markedly reduced (95%) colony formation in different BRAFV600E melanoma cell lines. Multiple genes of the MAPK and PI3K pathways found to be correlated with RMEL3 in TCGA samples were experimentally confirmed. RMEL3 knockdown led to downregulation of activators or effectors of these pathways, including FGF2, FGF3, DUSP6, ITGB3 and GNG2. RMEL3 knockdown induces gain of protein levels of tumor suppressor PTEN and the G1/S cyclin-Cdk inhibitors p21 and p27, as well as a decrease of pAKT (T308), BRAF, pRB (S807, S811) and cyclin B1. Consistently, knockdown resulted in an accumulation of cells in G1 phase and subG0/G1 in an asynchronously growing population. Thus, TCGA data and functional experiments demonstrate that RMEL3 is required for MAPK and PI3K signaling, and its knockdown decrease BRAFV600E melanoma cell survival and proliferation. Show less

Determination of the exogenous factors that regulate differentiation of neural stem/progenitor cells into neurons, oligodendrocytes and astrocytes is an important step in the clinical therapy of spinal cord injury (SCI). The Notch pathway inhibits the differentiation of neural stem/progenitor cells and Lingo-1 is a strong negative regulator for myelination and axon growth. While Lingo-1 shRNA and N-[N-(3, 5-difluorophenacetyl)-1-alanyl]-S-Phenylglycinet-butylester (DAPT), a Notch pathway inhibitor, have been used separately to help repair SCI, the results have been unsatisfactory. Here we investigated and elucidated the preliminary mechanism for the effect of Lingo-1 shRNA and DAPT on neural stem/progenitor cells differentiation. We found that neural stem/progenitor cells from E14 rat embryos expressed Nestin, Sox-2 and Lingo-1, and we optimized the transduction of neural stem/progenitor cells using lentiviral vectors encoding Lingo-1 shRNA. The addition of DAPT decreased the expression of Notch intracellular domain (NICD) as well as the downstream genes Hes1 and Hes5. Expression of NeuN, CNPase and GFAP in DAPT treated cells and expression of NeuN in Lingo-1 shRNA treated cells confirmed differentiation of neural stem/progenitor cells into neurons, oligodendrocytes and astrocytes. These results revealed that while Lingo-1 shRNA and Notch signaling inhibitor DAPT both promoted differentiation of neural stem cells into neurons, only DAPT was capable of driving neural stem/progenitor cells differentiation into oligodendrocytes and astrocytes. Since we were able to show that both Lingo-1 shRNA and DAPT could drive neural stem/progenitor cells differentiation, our data might aid the development of more effective SCI therapies using Lingo-1 shRNA and DAPT. Show less

Leucine-rich repeat and immunoglobulin domain-containing protein (LINGO)-1 is expressed in neural stem cells, and its neutralization results in sustained neuronal immaturity. Thus, targeted inhibition of LINGO-1 via RNA interference may enhance transplanted neural stem cell survival and neuronal differentiation in vivo. Furthermore, LINGO-1 RNA interference in neural stem cells represents a potential therapeutic strategy for spinal cord injury. Department of Spine Surgery, First Affiliated Hospital of Sun Yat-sen University. Translational Medicine Center Research Laboratory, First Affiliated Hospital of Sun Yat-sen University. Female Sprague-Dawley rats. The animals were divided into three groups that underwent laminectomy and complete spinal cord transection accompanied by transplantation of control-RNA interference-treated or LINGO-1-RNA interference-treated neural stem cells at the injured site in vivo. In vitro, neural stem cells were divided into four groups for the following treatments: control, control RNA interference lentivirus, LINGO-1 RNA interference lentivirus and LINGO-1 complementary DNA lentivirusand the Key Projects of the Natural Science Foundation of Guangdong Province (No. S2013020012818). Neural stem cells in each treatment group were examined for cell survival and neuronal differentiation in vitro and in vivo via immunofluorescence and Western blot analysis. Axonal regeneration and tissue repair were assessed via retrograde tracing using Fluorogold, electron microscopy, hematoxylin-eosin staining and MRI. Rats were also examined for functional recovery based on the measurement of spinal cord-evoked potentials and the Basso-Beattie-Bresnahan score. LINGO-1-RNA interference-treated neural stem cell transplantation increased tissue repair and functional recovery of the injured spinal cord in rats. Similarly, LINGO-1 RNA interference increased neural stem cell survival and neuronal differentiation in vitro. The mechanism underlying the effect of LINGO-1 RNA interference on the injured rat spinal cord may be that the significant inhibition of LINGO-1 expression in neural stem cells inactivated the RhoA and Notch signaling pathways, which act downstream of LINGO-1. Our findings indicate that transplantation of LINGO-1-RNA interference-treated neural stem cells facilitates functional recovery after spinal cord injury and represents a promising potential strategy for the repair of spinal cord injury. Show less

PI3Kδ has been found to be over-expressed in B-Cell-related malignancies. Despite the clinical success of the first selective PI3Kδ inhibitor, CAL-101, inhibition of PI3Kδ itself did not show too much cytotoxic efficacy against cancer cells. One possible reason is that PI3Kδ inhibition induced autophagy that protects the cells from death. Since class III PI3K isoform PIK3C3/Vps34 participates in autophagy initiation and progression, we predicted that a PI3Kδ and Vps34 dual inhibitor might improve the anti-proliferative activity observed for PI3Kδ-targeted inhibitors. We discovered a highly potent ATP-competitive PI3Kδ/Vps34 dual inhibitor, PI3KD/V-IN-01, which displayed 10-1500 fold selectivity over other PI3K isoforms and did not inhibit any other kinases in the kinome. In cells, PI3KD/V-IN-01 showed 30-300 fold selectivity between PI3Kδ and other class I PI3K isoforms. PI3KD/V-IN-01 exhibited better anti-proliferative activity against AML, CLL and Burkitt lymphoma cell lines than known selective PI3Kδ and Vps34 inhibitors. Interestingly, we observed FLT3-ITD AML cells are more sensitive to PI3KD/V-IN-01 than the FLT3 wt expressing cells. In AML cell inoculated xenograft mouse model, PI3KD/V-IN-01 exhibited dose-dependent anti-tumor growth efficacies. These results suggest that dual inhibition of PI3Kδ and Vps34 might be a useful approach to improve the PI3Kδ inhibitor's anti-tumor efficacy. Show less

Deciding appropriate therapy for multiple myeloma (MM) is challenging because of the occurrence of multiple chromosomal changes and the fatal nature of the disease. In the current study, gamabufotalin (GBT) was isolated from toad venom, and its tumor-specific cytotoxicity was investigated in human MM cells. We found GBT inhibited cell growth and induced apoptosis with the IC50 values <50 nM. Mechanistic studies using functional approaches identified GBT as an inhibitor of c-Myc. Further analysis showed that GBT especially evoked the ubiquitination and degradation of c-Myc protein, thereby globally repressing the expression of c-Myc target genes. GBT treatment inhibited ERK and AKT signals, while stimulating the activation of JNK cascade. An E3 ubiquitin-protein ligase, WWP2, was upregulated following JNK activation and played an important role in c-Myc ubiquitination and degradation through direct protein-protein interaction. The antitumor effect of GBT was validated in a xenograft mouse model and the suppression of MM-induced osteolysis was verified in a SCID-hu model in vivo. Taken together, our study identified the potential of GBT as a promising therapeutic agent in the treatment of MM. Show less

Apolipoprotein (apo)A-IV is a lipid emulsifying protein linked to a range of protective roles in obesity, diabetes, and cardiovascular disease. It exists in several states in plasma including lipid-bound in HDL and chylomicrons and as monomeric and dimeric lipid-free/poor forms. Our recent x-ray crystal structure of the central domain of apoA-IV shows that it adopts an elongated helical structure that dimerizes via two long reciprocating helices. A striking feature is the alignment of conserved proline residues across the dimer interface. We speculated that this plays important roles in the structure of the lipid-free protein and its ability to bind lipid. Here we show that the systematic conversion of these prolines to alanine increased the thermodynamic stability of apoA-IV and its propensity to oligomerize. Despite the structural stabilization, we noted an increase in the ability to bind and reorganize lipids and to promote cholesterol efflux from cells. The novel properties of these mutants allowed us to isolate the first trimeric form of an exchangeable apolipoprotein and characterize it by small-angle x-ray scattering and chemical cross-linking. The results suggest that the reciprocating helix interaction is a common feature of all apoA-IV oligomers. We propose a model of how self-association of apoA-IV can result in spherical lipoprotein particles, a model that may have broader applications to other exchangeable apolipoprotein family members. Show less

In the present study, we demonstrate that prolonged treatment by trastuzumab induced resistance of NCI-N87 gastric cancer cells to trastuzumab. The resistant cells possessed typical characteristics of epithelial to mesenchymal transition (EMT)/cancer stem cells and acquired more invasive and metastatic potentials both in vitro and in vivo. Long term treatment with trastuzumab dramatically inhibited the phosphorylation of Akt, but triggered the activation of STAT3. The level of IL-6 was remarkably increased, implicating that the release of IL-6 that drives the STAT3 activation initiates the survival signaling transition. Furthermore, the Notch activities were significantly enhanced in the resistant cells, companied by upregulation of the Notch ligand Jagged-1 and the Notch responsive genes Hey1 and Hey2. Inhibiting the endogenous Notch pathway reduced the IL-6 expression and restored the sensitivities of the resistant cells to trastuzumab. Blocking of the STAT3 signaling abrogated IL-6-induced Jagged-1 expression, effectively inhibited the growth of the trastuzumab resistant cells, and enhanced the anti-tumor activities of trastuzumab in the resistant cells. These findings implicate that the IL-6/STAT3/Jagged-1/Notch axis may be a useful target and that combination of the Notch or STAT3 inhibitors with trastuzumab may prevent or delay clinical resistance and improve the efficacy of trastuzumab in gastric cancer. Show less

Schizophrenia is a heritable, heterogeneous common psychiatric disorder. In this study, we evaluated the hypothesis that de novo variants (DNVs) contribute to the pathogenesis of schizophrenia. We performed exome sequencing in Chinese patients (N = 45) with schizophrenia and their unaffected parents (N = 90). Forty genes were found to contain DNVs. These genes had enriched transcriptional co-expression profile in prenatal frontal cortex (Bonferroni corrected p < 9.1 × 10(-3)), and in prenatal temporal and parietal regions (Bonferroni corrected p < 0.03). Also, four prenatal anatomical subregions (VCF, MFC, OFC and ITC) have shown significant enrichment of connectedness in co-expression networks. Moreover, four genes (LRP1, MACF1, DICER1 and ABCA2) harboring the damaging de novo mutations are strongly prioritized as susceptibility genes by multiple evidences. Our findings in Chinese schizophrenic patients indicate the pathogenic role of DNVs, supporting the hypothesis that schizophrenia is a neurodevelopmental disease. Show less

Phytochrome-interacting factor 3 (PIF3) activates light-responsive transcriptional network genes in coordination with the circadian clock and plant hormones to modulate plant growth and development. However, little is known of the roles PIF3 plays in the responses to abiotic stresses. In this study, the cloning and functional characterization of the ZmPIF3 gene encoding a maize PIF3 protein is reported. Subcellular localization revealed the presence of ZmPIF3 in the cell nucleus. Expression patterns revealed that ZmPIF3 is expressed strongly in leaves. This expression responds to polyethylene glycol, NaCl stress, and abscisic acid application, but not to cold stress. ZmPIF3 under the control of the ubiquitin promoter was introduced into rice. No difference in growth and development between ZmPIF3 transgenic and wild-type plants was observed under normal growth conditions. However, ZmPIF3 transgenic plants were more tolerant to dehydration and salt stresses. ZmPIF3 transgenic plants had increased relative water content, chlorophyll content, and chlorophyll fluorescence, as well as significantly enhanced cell membrane stability under stress conditions. The over-expression of ZmPIF3 increased the expression of stress-responsive genes, such as Rab16D, DREB2A, OSE2, PP2C, Rab21, BZ8 and P5CS, as detected by real-time PCR analysis. Taken together, these results improve our understanding of the role ZmPIF3 plays in abiotic stresses signaling pathways; our findings also indicate that ZmPIF3 regulates the plant response to drought and salt stresses. Show less

Apolipoprotein (apo)A-IV plays important roles in dietary lipid and glucose metabolism, and knowledge of its structure is required to fully understand the molecular basis of these functions. However, typical of the entire class of exchangeable apolipoproteins, its dynamic nature and affinity for lipid has posed challenges to traditional high resolution structural approaches. We previously reported an x-ray crystal structure of a dimeric truncation mutant of apoA-IV, which showed a unique helix-swapping molecular interface. Unfortunately, the structures of the N and C termini that are important for lipid binding were not visualized. To build a more complete model, we used chemical cross-linking to derive distance constraints across the full-length protein. The approach was enhanced with stable isotope labeling to overcome ambiguities in determining molecular span of the cross-links given the remarkable similarities in the monomeric and dimeric apoA-IV structures. Using 51 distance constraints, we created a starting model for full-length monomeric apoA-IV and then subjected it to two modeling approaches: (i) molecular dynamics simulations and (ii) fitting to small angle x-ray scattering data. This resulted in the most detailed models yet for lipid-free monomeric or dimeric apoA-IV. Importantly, these models were of sufficient detail to direct the experimental identification of new functional residues that participate in a "clasp" mechanism to modulate apoA-IV lipid affinity. The isotope-assisted cross-linking approach should prove useful for further study of this family of apolipoproteins in both the lipid-free and -bound states. Show less

Thyroid cancer is a malignant neoplasm originated from thyroid cells. It can be classified into papillary carcinomas (PTCs) and anaplastic carcinomas (ATCs). Although ATCs are in an very aggressive status and cause more death than PTCs, their difference is poorly understood at molecular level. In this study, we focus on the transcriptome difference among PTCs, ATCs and normal tissue from a published dataset including 45 normal tissues, 49 PTCs and 11 ATCs, by applying a machine learning method, maximum relevance minimum redundancy, and identified 9 genes (BCL2, MRPS31, ID4, RASAL2, DLG2, MY01B, ZBTB5, PRKCQ and PPP6C) and 1 miscRNA (miscellaneous RNA, LOC646736) as important candidates involved in the progression of thyroid cancer. We further identified the protein-protein interaction (PPI) sub network from the shortest paths among the 9 genes in a PPI network constructed based on STRING database. Our results may provide insights to the molecular mechanism of the progression of thyroid cancer. Show less