👤 Zhe Wan

Epithelial-mesenchymal transition (EMT) plays a critical role in cancer progression and is primarily regulated by several EMT-inducing transcription factors (EMT-TFs), including TWIST1, TWIST2, SNAI1, SNAI2, ZEB1, and ZEB2. However, the prognostic value of EMT-TFs remains controversial in head and neck squamous cell carcinoma (HNSCC). Studies on the prognostic role of EMT-TFs in HNSCC were searched for in the Web of Science, Science Direct, Proquest, EMBASE, PubMed, and Cochrane Library. Meta-analysis was performed by using Revman 5.2 software. The pooled analysis showed that overexpression of EMT-TFs indicated a poor overall survival (OS) (HR = 1.93, 95% CI = 1.67-2.23) of HNSCC. Subgroup analysis for individual EMT-TFs revealed that overexpression of TWIST1 (HR = 1.61, 95% CI = 1.29-2.02), SNAI1 (HR = 2.17, 95% CI = 1.63-2.88), SNAI2 (HR = 1.90, 95% CI = 1.38-2.62), and ZEB1 (HR = 2.70, 95% CI = 1.61-4.53) were significantly associated with poor OS of HNSCC. These findings support the hypothesis that overexpression of EMT-TFs indicates a poor prognosis for HNSCC patients. Show less

Existing evidence has shown that circulating Epstein-Barr virus (EBV)-miR-BART13-3p is highly expressed in plasma of nasopharyngeal carcinoma (NPC) patients, especially among patients with advanced diseases. However, the exact role that EBV-miR-BART13-3p plays in the development of NPC remains poorly understood. Here we show that up-regulated expression of EBV-miR-BART13-3p leads to increased capacity in migration and invasion of NPC cells Show less

Procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) was induced in hypoxia and participated in cancer development. However, the role of PLOD2 in endometrial carcinoma remains unclear. To explore the influences and regulation mechanism of PLOD2 in endometrial carcinoma under hypoxic condition. The small interfering RNA (siRNA) targeting to PLOD2 and pcDNA3.1-PLPD2 were transfected to endometrial carcinoma cells to alter PLOD2 expression. Cell proliferation ability was determined by colony formation assay. Wound healing assay used to detect cell migration ability. Transwell invasion assay was used to detect cell invasion ability. PLOD2 and Hypoxia-inducible factor-1α (HIF-1α) were induced by hypoxia. Down-regulation of PLOD2 did not affect endometrial carcinoma cell proliferation ability, while inhibited cell migration, invasion under hypoxic condition. Besides, down-regulation of PLOD2 increased the levels of γ-catenin and E-cadherin and decreased levels of Fibronectin and Snail under hypoxic condition. Down-regulation of PLOD2 also inactivated Src and phosphoinositide 3-kinase (PI3K)/ protein kinase B (Akt) signaling under hypoxic condition. The promoting effects of PLOD2 overexpression on migration, invasion and epithelial-mesenchymal transition (EMT) of endometrial carcinoma cells were reversed by Akt inhibitor (MK2206) under hypoxic condition. PLOD2 expression was increased in endometrial carcinoma cells under hypoxic condition. PLOD2 modulated migration, invasion, and EMT of endometrial carcinoma cells via PI3K/Akt signaling. PLOD2 may be a potential therapeutic target for endometrial carcinoma. Show less

Hypertriglyceridemia severity is linked to acute pancreatitis prognosis, but it remains unknown why a portion of severe hypertriglyceridemia patients do not develop severe acute pancreatitis. To investigate whether hypertriglyceridemia subtypes affect acute pancreatitis progression, we analyzed two genetically modified hypertriglyceridemia mouse models-namely, glycosylphosphatidylinositol high-density lipoprotein binding protein 1 knockout (Gpihbp1-/-) and apolipoprotein C3 transgenic (ApoC3-tg) mice. Acute pancreatitis was induced by 10 intraperitoneal caerulein injections. Biochemical assays and pathological analysis were performed for the severity evaluation of acute pancreatitis. Plasma triglyceride-rich lipoproteins (TRLs), including chylomicrons and very low-density lipoprotein (VLDL), were collected via ultracentrifugation to evaluate their cytotoxic effects on primary pancreatic acinar cells (PACs). We found that the particle sizes of Gpihbp1-/- TRLs were larger than ApoC3-tg TRLs. Severe pancreatic injury with large areas of pancreatic necrosis in the entire lobule was induced in Gpihbp1-/- mice when plasma triglyceride levels were greater than 2000 mg/dL. However, ApoC3-tg mice with the same triglyceride levels did not develop large areas of pancreatic necrosis, even upon the administration of poloxamer 407 to further increase triglyceride levels. Meanwhile, in the acute pancreatitis model, free fatty acids (FFAs) in the pancreas of Gpihbp1-/- mice were greater than in ApoC3-tg mice. TRLs from Gpihbp1-/- mice released more FFAs and were more toxic to PACs than those from ApoC3-tg mice. Chylomicrons from patients showed the same effects on PACs as TRLs from Gpihbp1-/- mice. Gpihbp1-/- mice with triglyceride levels below 2000 mg/dL had milder pancreatic injury and less incidence of pancreatic necrosis than those with triglyceride levels above 2000 mg/dL, similar to Gpihbp1-/-mice with triglyceride levels above 2000 mg/dL but with fenofibrate administration. These findings demonstrated that hypertriglyceridemia subtypes with large TRL particles could affect acute pancreatitis progression and that chylomicrons showed more cytotoxicity than VLDL by releasing more FFAs. Show less

The function of ten-eleven translocation methylcytosine dioxygenase 1 (TET1) in cancer is background dependent and may be involved in the initial step of active DNA demethylation, while there is little research to decipher the role of TET1 in DNA methylation-sensitive colon cancer. Downregulated TET1 expression assayed by quantitative real-time PCR (qRT-PCR) was observed in both colon cancer samples and cancer cell lines of HT29, HCT116, and SW48. Such downregulation could promote colon cancer cells proliferation as indicated by the fact that shTET1 could increase the viability of HT29 and HCT116 cells determined by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide and cell count assay accompanied with upregulation of β-catenin (CTNNB1) and WNT luciferase activity, which was further confirmed as shTET1 could increase the tumor volume and tumor weight, and decrease the body weight in HT29 cells inoculated BALB/C nude mice. The CTNNB1 transfection could rescue the cell growth diminished by normal expression of TET1. shTET1 could promote axis inhibition protein1 (AXIN1) expression and the cell proliferation effect induced by TET1 short hairpin RNA was attenuated by co-inhibition of AXIN1. All of these indicate that TET1 can suppress colon cancer proliferation and the inhibition of the β-catenin pathway is AXIN1 dependent. Show less

This study was designed to identify the potential key protein interaction networks, genes, and correlated pathways in early-onset colorectal cancer (CRC) via bioinformatics methods. We selected microarray data GSE4107 consisting 12 patient's colonic mucosa and 10 healthy control mucosa; initially, the GSE4107 were downloaded and analyzed using limma package to identify differentially expressed genes (DEGs). A total of 131 DEGs consisting of 108 upregulated genes and 23 downregulated genes of patients in early-onset CRC were selected by the criteria of adjusted P values <.01 and |log2 fold change (FC)| ≥ 2. The gene ontology functional enrichment analysis and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were accomplished to view the biological process, cellular components, molecular function, and the KEGG pathways of DEGs. Finally, protein-protein interactions (PPIs) were constructed, and the hub protein module was identified. Genes such as ACTA2, ACTG2, MYH11, CALD1, MYL9, TPM2, and LMOD1 were strongly implicated in CRC. In summary, in this study, we indicated that molecular mechanisms were involved in muscle contraction and vascular smooth muscle contraction signaling pathway, which improve our understanding of CRC and could be used as new therapeutic targets for CRC. Show less

Growing evidence has highlighted the immune response as an important feature of carcinogenesis and therapeutic efficacy in clear cell renal cell carcinoma (ccRCC). This study categorized ccRCC cases into high and low score groups based on their immune/stromal scores generated by the ESTIMATE algorithm, and identified an association between these scores and prognosis. Differentially expressed tumor environment (TME)-related genes extracted from common upregulated components in immune and stromal scores were described using functional annotations and protein-protein interaction (PPI) networks. Most PPIs were selected for further prognostic investigation. Many additional previously neglected signatures, including Show less

As an oncogene, long noncoding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) can promote tumor metastasis. Hyperexpression of MALAT1 has been observed in many malignant tumors, including hepatocellular carcinoma (HCC). However, the role and mechanism of MALAT1 in HCC remain unclear. Thirty human HCC and paracancerous tissue specimens were collected, and the human hepatoma cell lines Huh7 and HepG2 were cultured according to standard methods. MALAT1 and Snail family zinc finger (Slug) expression were measured by real-time PCR, immunohistochemistry, and western blotting. Luciferase reporter assay and RNA immunoprecipitation (RIP) assay verified the direct interaction between miR-124-3p and Slug(SNAI2) or MALAT1. Wound healing and transwell assays were performed to examine invasion and migration, and a subcutaneous tumor model was established to measure tumor progression in vivo. MALAT1 expression was upregulated in HCC tissues and positively correlated with Slug expression. MALAT1 and miR-124-3p bind directly and reversibly to each other. MALAT1 silencing inhibited cell migration and invasion. miR-124-3p inhibited HCC metastasis by targeting Slug. MALAT1 regulates Slug through miR-124-3p, affecting HCC cell metastasis. Thus, the MALAT1/miR-124-3p/Slug axis plays an important role in HCC. Show less

Peanut allergy (PA) is a complex disease with both environmental and genetic risk factors. Previously, PA loci were identified in filaggrin (FLG) and HLA in candidate gene studies, and loci in HLA were identified in a genome-wide association study and meta-analysis. We sought to investigate genetic susceptibility to PA. Eight hundred fifty cases and 926 hyper-control subjects and more than 7.8 million genotyped and imputed single nucleotide polymorphisms (SNPs) were analyzed in a genome-wide association study to identify susceptibility variants for PA in the Canadian population. A meta-analysis of 2 phenotypes (PA and food allergy) was conducted by using 7 studies from the Canadian, American (n = 2), Australian, German, and Dutch (n = 2) populations. An SNP near integrin α6 (ITGA6) reached genome-wide significance with PA (P = 1.80 × 10 This study identifies multiple novel loci as risk factors for PA and food allergy and establishes C11orf30 as a risk locus for both PA and food allergy. Multiple genes (C11orf30/EMSY, SKAP1, and CTNNA3) identified by this study are involved in epigenetic regulation of gene expression. Show less

Diminished cardiac contractile function is a characteristic feature of dilated cardiomyopathy (DCM) and many other heart failure (HF) causing etiologies. We tested the hypothesis that targeting the sarcomere to increase cardiac contractility can effectively prevent the DCM phenotype in muscle-LIM protein knockout (MLP Show less

To develop a cost-effective molecular regulator to improve growth metabolism and immunity of animals, a recombinant plasmid co-expressing fatty acid desaturase (mFat-1) and pig insulin growth like factor 1 (IGF-1) genes was constructed by the 2 A self-cleavage technique. After entrapment within modified chitosan nanoparticles (chitosan modified with polyethyleneglycol-polyethylenimine, CPP), the recombinant plasmid was injected intramuscularly into mice. Compared with controls, co-expression of mFat-1 and IGF-1 significantly raised the level of serum IGF-1, and increased the liver and muscle docosa hexaenoic acid (DHA) content. Th and Tc cell levels were also elevated, as were expression levels of serum IL-4 and IL-6 genes. These results demonstrate that the immunity and metabolism of an animal can be effectively improved by co-expression of mFat-1 and IGF-1 genes in vivo, which may contribute to further development of novel immunomodulators with beneficial effects on growth metabolism and immunity. Show less

Nuclear receptors (NRs) are a class of transcription factors that regulate many cellular functions through manipulation of gene expression and also play important roles in tumorigenesis, proliferation, progression and prognosis in various kinds of cancers according to recent studies. This work aimed to determine the predictive ability of NRs in muscle-invasive bladder cancer (MIBC). A total of 308 MIBC patients with complete clinicopathological and RNASeq data from The Cancer Genome Atlas (TCGA) cohort were collected for filtration. Genes showed clear correlations with overall survival (OS) and recurrence free survival (RFS) were further validated in 123 MIBC patients recruited consecutively from 2008 to 2012 in Fudan University Shanghai Cancer Center (FUSCC) cohort. Cox proportional hazards regression model and Kaplan-Meier plot were used to assess the relative factors. In TCGA cohort, we found that high NR1H3 (HR=0.779, 95% CI: 0.634 - 0.957), NR2C1 (HR=0.673, 95% CI: 0.458 - 0.989) and NR2F6 (HR=0.750, 95% CI: 0.574 - 0.980) expressions were independent factors of favorable OS, while only low NR1H3 (log-rank test, P=0.0076) and NR2F6 (log-rank test, P=0.0395) expressions had the ability to predict poor prognosis for RFS. Further, in FUSCC validating cohort, we confirmed that low NR1H3 expression level was independent factor of poor OS (HR=1.295, 95% CI: 1.064 - 1.576) and it had the ability to predict poor RFS (log-rank test, P=0.0059). Low NR1H3 expression level is an independent prognostic factor of poor OS, and can also predict worse RFS in MIBC patients. Our "TCGA filtrating and local database validating" model can help reveal more prognostic biomarkers and cast a new light in understanding certain gene function in MIBC. Show less

The class III phosphoinositide 3-kinase VPS34 plays a key role in the regulation of vesicular trafficking and macroautophagy. So far, we know little about the molecular mechanism of VPS34 activation besides its interaction with regulatory proteins to form complexes. Here, we report that VPS34 is specifically acetylated by the acetyltransferase p300, and p300-mediated acetylation represses VPS34 activity. Acetylation at K771 directly diminishes the affinity of VPS34 for its substrate PI, while acetylation at K29 hinders the VPS34-Beclin 1 core complex formation. Inactivation of p300 induces VPS34 deacetylation, PI3P production, and autophagy, even in AMPK 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 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

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

Mitogen/extracellular signal-regulated kinase kinase-5 (MEK5) has been confirmed to play a pivotal role in tumor carcinogenesis and progression. However, few studies have investigated the role of MEK5 in colorectal cancer (CRC). MEK5 expression was determined by immunohistochemistry (IHC) in tissue microarrays (TMAs) containing 2 groups of tissues, and western blotting was used to confirm MEK5 expression in 8 cases of primary CRC tissues and paired normal mucosa. RNA interference was used to verify the biological function of MEK5 gene in the development of CRC. IHC revealed the expression of MEK5 was higher in tumor tissues (38.1 %), compared with adjacent normal tissue (8.3 %). Western blot showed that, MEK5 expression was upregulated in CRC tumor tissues compared with normal tissue. Analysis of clinical pathology parameters indicated MEK5 overexpression was significantly correlated with the depth of invasion, lymph node metastasis, distant metastasis and histological grade. Survival analysis revealed that MEK5 overexpression negatively correlated with cancer-free survival (hazard ratio 1.64, P = 0.017). RNA interference-mediated knockdown of MEK5 in SW480 colon cancer cells decreased their proliferation, division, migration and invasiveness in vitro and slowed down tumors growth in mice engrafted with the cells. MEK5 plays an important role in CRC progression and may be a potential molecular target for the treatment of CRC. Show less

Anaphase-promoting complex/cyclosome/Cdh1 is a multi-subunit ubiquitin E3 ligase that drives M to G1 cell cycle progression through primarily earmarking various substrates for ubiquitination and subsequent degradation by the 26S proteasome. Notably, emerging evidence suggested that Cdh1 could also function in various cellular processes independent of anaphase-promoting complex/cyclosome. To this end, we recently identified an anaphase-promoting complex/cyclosome-independent function of Cdh1 in modulating osteoblast differentiation through activating Smurf1, one of the NEDD4 family of HECT domain-containing E3 ligases. However, it remains largely unknown whether Cdh1 could exert its tumor suppressor role through similarly modulating the E3 ligase activities of other NEDD4 family members, most of which have characterized important roles in tumorigenesis. Here we report that in various tumor cells, Cdh1, conversely, suppresses the E3 ligase activity of WWP2, another NEDD4 family protein, in an anaphase-promoting complex/cyclosome-independent manner. As such, loss of Cdh1 activates WWP2, leading to reduced abundance of WWP2 substrates including PTEN, which subsequently activates PI3K/Akt oncogenic signaling to facilitate tumorigenesis. This study expands the non-anaphase-promoting complex/cyclosome function of Cdh1 in regulating the NEDD4 family E3 ligases, and further suggested that enhancing Cdh1 to inhibit the E3 ligase activity of WWP2 could be a promising strategy for treating human cancers. Show less

Craniofacial anomalies (CFAs) characterized by birth defects of skull and facial bones are the most frequent congenital disease. Genomic analysis has identified multiple genes responsible for CFAs; however, the underlying genetic mechanisms for the majority of CFAs remain largely unclear. Our previous study revealed that the Wwp2 E3 ubiquitin ligase facilitates craniofacial development in part through inducing monoubiquitination and activation of the paired-like homeobox transcription factor, Goosecoid (Gsc). Here we report that Gsc is also ubiquitinated and activated by the APC(Cdh1) E3 ubiquitin ligase, leading to transcriptional activation of various Gsc target genes crucial for craniofacial development. Consistenly, neural crest-specific Cdh1-knockout mice display similar bone malformation as Wwp2-deficient mice in the craniofacial region, characterized by a domed skull, a short snout and a twisted nasal bone. Mechanistically, like Wwp2-deficient mice, mice with Cdh1 deficiency in neural crest cells exhibit reduced Gsc/Sox6 transcriptional activities. Simultaneous deletion of Cdh1 and Wwp2 results in a more severe craniofacial defect compared with single gene deletion, suggesting a synergistic augmentation of Gsc activity by these two E3 ubiquitin ligases. Hence, our study reveals a novel role for Cdh1 in craniofacial development through promoting APC-dependent non-proteolytic ubiquitination and activation of Gsc. Show less

Although many studies have investigated the association of the APOA5 -1131T/C polymorphism with coronary artery disease (CAD), definite conclusions have not been drawn. To understand the effects of the APOA5 -1131T/C polymorphism on the risk of developing CAD, we performed an updated meta-analysis in the Chinese population. Relevant studies published till April 2015 were identified from databases such as PubMed, Springer Link, Ovid, Chinese Wanfang Data Knowledge Service Platform, Chinese National Knowledge Infrastructure, and Chinese Biology Medicine. A total of 19 studies including 3983 patients and 4358 controls were involved in this meta-analysis. The crude OR with 95%CI was calculated to assess the strength of the association. With the pooled data from the studies included in this meta-analysis, we found a significant association between the APOA5 -1131T/C polymorphism and CAD risk in the Chinese population (C vs T: OR = 1.34, 95%CI = 1.16-1.54; CC vs TT: OR = 1.73, 95%CI = 1.30-2.30; CC vs TT and TC: OR = 1.51, 95%CI = 1.17-1.95; CC vs TC: OR = 1.30, 95%CI = 1.03-1.65). Stratified analyses according to the geographical location and source of controls revealed significantly increased risk in South China and in population-based studies. In conclusion, our meta-analysis provides substantial evidence that the APOA5 -1131T/C polymorphism might contribute to CAD development in the Chinese population. Show less

Pyrrolizidine alkaloids (PAs) are a group of phytotoxins that can induce human liver injury, particularly hepatic sinusoidal obstruction syndrome (HSOS). To date, the molecular targets of PA-induced HSOS are largely unknown. In this study, retrorsine (RTS), a known hepatotoxic PA, was used as a representative PA for proteomic studies. Toxicological assessment demonstrated that 35 mg/kg RTS (designated as RTS-L) caused early lesions of HSOS at 24 h after dosing. A proteomic approach revealed 17 up-regulated and 31 down-regulated proteins in RTS-L-treated rats. Subsequently, bioinformatic analysis suggested that two proteins, carbamoyl-phosphate synthase (CPS1) (p < 0.05) and ATP synthase subunit beta (ATP5B) (p < 0.01) were associated with RTS-L intoxication. Using immunohistochemical staining, we further verified the down-regulation of CPS1 and ATP5B in RTS-L-treated rats. These findings indicated that CPS1 and ATP5B were altered in the RTS-induced early lesions of HSOS in rats, and therefore, these two proteins and their involved pathways might play important roles in the initiation of HSOS. To the best of our knowledge, our study using a proteomic approach combined with conventional toxicological assessment is the first systems toxicology study on PA-induced HSOS. The results of this study provide novel findings on protein profiles in response to PA exposure, which can serve as a starting point to further investigate potential protein targets and their interactions with PAs to induce HSOS. Show less

Adequate intake of dietary fibers has proven metabolic and cardiovascular benefits, molecular mechanisms remain still limited. This study was aimed to investigate the effects of cereal dietary fiber on obesity-related liver lipotoxicity in C57BL/6J mice fed a high-fat/cholesterol (HFC) diet and underlying mechanism. Forty-eight adult male C57BL/6J mice were randomly given a reference chow diet, or a high fat/cholesterol (HFC) diet supplemented with or without oat fiber or wheat bran fiber for 24 weeks. Our results showed mice fed oat or wheat bran fiber exhibited lower weight gain, lipid profiles and insulin resistance, compared with HFC diet. The two cereal dietary fibers potently decreased protein expressions of sterol regulatory element binding protein-1 and key factors involved in lipogenesis, including fatty acid synthase and acetyl-CoA carboxylase in target tissues. At molecular level, the two cereal dietary fibers augmented protein expressions of peroxisome proliferator-activated receptor alpha and gamma, liver X receptor alpha, and ATP-binding cassette transporter A1 in target tissues. Our findings indicated that cereal dietary fiber supplementation abrogated obesity-related liver lipotoxicity and dyslipidemia in C57BL/6J mice fed a HFC diet. In addition, the efficacy of oat fiber is greater than wheat bran fiber in normalizing these metabolic disorders and pathological profiles. Show less

The aim of this study is to identify and validate protein change in the serum from PD patients. We used serum samples from 21 PD patients and 20 age-matched normal people as control to conduct a comparative proteomic study. We performed 2-DE and analyzed the differentially expressed protein spots by LC-MS/MS. In PD group 13 spots were shown to be differentially expressed compared to control group. They were identified as 6 proteins. Among these, 3 proteins were confirmed by Western blot analysis. It showed that the frequency of fibrinogen γ-chain (FGG) appeared 70% in PD, which could not be detected in control group. The protein of inter-alpha-trypsin inhibitor heavy chain H4 (ITI-H4) was found to exist two forms in serum. The full size (120 kDa) of the protein was increased and the fragmented ITI-H4 (35 kDa) was decreased in PD group. The ratio of full size ITI-H4 to fragmented ITI-H4 in PD patients was 3.85 ± 0.29-fold higher than in control group. Furthermore, fragmented Apo A-IV (∼ 26 kDa) was mainly detected in control group, while it was rare to be found in PD group. Above findings might be useful for diagnosis of PD. When the expressions of FGG and 120 kDa ITI-H4 are increase, as well as ∼ 26 kDa Apo A-IV disappear would provide strong evidence for PD. Show less

Apoptosis constitutes a system for the removal of aged, or damaged cells, which is regulated by the interplay of pro-apoptotic and antiapoptotic proteins. Previous study has shown that Juvenile Batten disease protein, CLN3, is antiapoptotic gene in NT2 neuronal precursor cells and a few types of cancers. However, in colorectal cancer, whether CLN3 also play its antiapoptotic role and the effect of targeted controlling CLN3 on the biological behavior of human colorectal cancer cell is unknown. We employed the sequence-specific siRNA silencing the CLN3 gene and investigated its effects on growth and apoptosis of colorectal cancer HCT116 cells, which has highest elevation of CLN3 expression among four colorectal cancer cell lines. After CLN3 specific siRNA transfection, mRNA and protein expression levels of CLN3 in HCT116 cells were noticeably decreased. Moreover, CLN3-siRNA inhibited the proliferation of colorectal cancer cells, promoted their apoptosis and induced G0/G1 cell cycle arrest. Our current study demonstrated that CLN3 was expressed in colorectal cancer cells at a high frequency. Moreover, CLN3 down-regulation with RNA interference can inhibit proliferation, apoptosis, and cell cycle progression of colorectal cancer cells. Our study represented a potential new approach to understanding the role of CLN3 in cancer and provides a potential novel strategy colorectal cancer therapy. Show less

Estrogen has pleiotropic effects on the cardiovascular diseases, yet the underlying mechanisms remain incompletely understood. Cholesterol efflux is a key mechanism through which to prevent foam cell formation and the development of atherosclerosis. Recent studies highlight the role of vascular smooth muscle cell (VSMC)-derived foam cells in atherogenesis. However, it remains unclear whether estrogen promotes cholesterol efflux from VSMCs and inhibits VSMC-derived foam cell formation. In the present study, we demonstrated that 17β-estradiol (E2) markedly enhanced cholesterol efflux to apolipoprotein (apo)A-1 and high-density lipoprotein (HDL) and attenuated oxidized low-density lipoprotein (ox-LDL) induced cholesteryl ester accumulation in VSMCs, which was associated with an increase in the expression of ATP-binding cassette transporters ABCA1 and ABCG1. The upregulation of ABCA1 and ABCG1 expression by E2 resulted from liver X receptor (LXR)α activation, which was confirmed by the prevention of the expression of ABCA1 and ABCG1 after inhibition of LXRα with a pharmacological inhibitor or small interfering RNA (siRNA). Furthermore, E2 increased LXRα, ABCA1 and ABCG1 expression in VSMCs via the estrogen receptor (ER), and the involvement of ERβ was confirmed by the use of selective ERα or ERβ antagonists (MPP and PHTPP) and agonists (PPT and DPN). These findings suggest that E2 promotes cholesterol efflux from VSMCs and reduces VSMC-derived foam cell formation via ERβ- and LXRα-dependent upregulation of ABCA1 and ABCG1 and provide novel insights into the anti-atherogenic properties of estrogen. Show less

The Crumbs (Crb) complex, formed by Crb, PALS1, and PATJ, is evolutionarily conserved in metazoans and acts as a master cell-growth and -polarity regulator at the apical membranes in polarized epithelia. Crb intracellular functions, including its direct binding to PALS1, are mediated by Crb's highly conserved 37-residue cytoplasmic tail. However, the mechanistic basis governing the highly specific Crb-PALS1 complex formation is unclear, as reported interaction between the Crb tail (Crb-CT) and PALS1 PSD-95/DLG/ZO-1 (PDZ) domain is weak and promiscuous. Here we have discovered that the PDZ-Src homolgy 3 (SH3)-Guanylate kinase (GK) tandem of PALS1 binds to Crb-CT with a dissociation constant of 70 nM, which is ∼ 100-fold stronger than the PALS1 PDZ-Crb-CT interaction. The crystal structure of the PALS1 PDZ-SH3-GK-Crb-CT complex reveals that PDZ-SH3-GK forms a structural supramodule with all three domains contributing to the tight binding to Crb. Mutations disrupting the tertiary interactions of the PDZ-SH3-GK supramodule weaken the PALS1-Crb interaction and compromise PALS1-mediated polarity establishment in Madin-Darby canine kidney (MDCK) cysts. We further show that specific target binding of other members of membrane-associated guanylate kinases (MAGUKs) (e.g., CASK binding to neurexin) also requires the presence of their PDZ-SH3-GK tandems. Show less

We used a complete spinal cord transection model and locomotor function, histological, and immunohistochemical examinations to evaluate the effects of local injection of lentivirus/LINGO-1-short hairpin RNA (VL) on rats with spinal cord injury (SCI). To demonstrate the neuroregenerative and neuroprotective effects of LINGO-1 RNAi on complete transection SCI rats. LINGO-1 has been reported as a negative regulator of axonal sprouting and its antagonist was determined to improve functional outcomes in SCI rats. However, it has not been assessed whether blockade of LINGO-1 mediated by lentivirus vectors could stimulate neural recovery after SCI. Complete spinal cord transection was made at T10 level. Suspension of lentivirus vectors encoding LINGO-1-short hairpin RNA was injected into the lesion gap. Controls received control vectors in the same manner and the sham group was subjected to laminectomy only. The Basso-Beattie-Bresnahan scale and surface righting reflex test were used to evaluate functional outcomes. Finally, the spinal cords were harvested for histological and immunohistochemical analysis. The treatment with VL improved Basso-Beattie-Bresnahan scores and surface righting reflex after SCI. Tissue repair was facilitated and the cavity area was significantly decreased in VL-treated animals. More sprouting and myelinated nerve fibers were detected within the injured site in the VL group as compared with the control. In addition, the number of survival neurons and oligodendrocytes around the epicenter was notably higher under the VL condition. Local injection of lentivirus/LINGO-1-short hairpin RNA after complete transection of spinal cord resulted in meaningful histological and functional outcomes in rats. The mechanism of VL protection may be related to its promotion of axonal sprouting, remyelination, and cell survival. Show less

Lingo-1 is selectively expressed on both oligodendrocytes and neurons in the central nervous system (CNS) and serves as a key negative regulator of nerve regeneration, implying a therapeutic target for spinal cord injury (SCI). Here we described a strategy to knock-down Lingo-1 expression in vivo using lentiviral vectors encoding Lingo-1 short harpin interfering RNA (shRNA) delivered by Pluronic F-127 (PF-127) gel, a non-cytotoxic scaffold and gene delivery carrier, after the complete transection of the T10 spinal cord in adult rats. We showed administration of PF-127 encapsulating Lingo-1 shRNA lentiviral vectors efficiently down-regulated the expression of Lingo-1, and exhibited transduction efficiency comparable to using vectors alone in oligodendrocyte culture in vitro. Furthermore, similar silencing effects and higher transfection efficiency were observed in vivo when Lingo-1 shRNA was co-delivered to the injured site by PF-127 gel with lower viral concentrations. Cografting of gel and Lingo-1 RNAi significantly promoted functional recovery and nerve regeneration, enhanced neurite outgrowth and synapses formation, preserved myelinated axons, and induced the proliferation of glial cells. In addition, the combined implantation also improved neuronal survival and inhibited cell apoptosis, which may be associated with the attenuation of endoplasmic reticulum (ER) stress after SCI. Together, our data indicated that delivering Lingo-1 shRNA by gel scaffold was a valuable treatment approach to SCI and PF-127 delivery of viral vectors to the spinal cord may provide strategy to study and develop therapies for SCI. Show less

The individuals carrying melanocortin-1 receptor (MC1R) variants, especially those associated with red hair color, fair skin, and poor tanning ability (RHC trait), are more prone to melanoma; however, the underlying mechanism is poorly defined. Here, we report that UVB exposure triggers phosphatase and tensin homolog (PTEN) interaction with wild-type (WT), but not RHC-associated MC1R variants, which protects PTEN from WWP2-mediated degradation, leading to AKT inactivation. Strikingly, the biological consequences of the failure of MC1R variants to suppress PI3K/AKT signaling are highly context dependent. In primary melanocytes, hyperactivation of PI3K/AKT signaling leads to premature senescence; in the presence of BRAF(V600E), MC1R deficiency-induced elevated PI3K/AKT signaling drives oncogenic transformation. These studies establish the MC1R-PTEN axis as a central regulator for melanocytes' response to UVB exposure and reveal the molecular basis underlying the association between MC1R variants and melanomagenesis. Show less