👤 Qingzuo Zhao

The ability of liver to respond to changes in nutrient availability is essential for the maintenance of metabolic homeostasis. Autophagy encompasses mechanisms of cell survival, including capturing, degrading, and recycling of intracellular proteins and organelles in lysosomes. During negative nutrient status, autophagy provides substrates to sustain cellular metabolism and hence, tissue function. Severe negative energy balance in dairy cows is associated with fatty liver. The aim of this study was to investigate the hepatic autophagy status in dairy cows with severe fatty liver and to determine associations with biomarkers of liver function and inflammation. Liver and blood samples were collected from multiparous cows diagnosed as clinically healthy (n = 15) or with severe fatty liver (n = 15) at 3 to 9 d in milk. Liver tissue was biopsied by needle puncture, and serum samples were collected on 3 consecutive days via jugular venipuncture. Concentrations of free fatty acids and β-hydroxybutyrate were greater in cows with severe fatty liver. Milk production, dry matter intake, and concentration of glucose were all lower in cows with severe fatty liver. Activities of serum aspartate aminotransferase, alanine aminotransferase, glutamate dehydrogenase, and γ-glutamyl transferase were all greater in cows with severe fatty liver. Serum concentrations of haptoglobin and serum amyloid A were also markedly greater in cows with severe fatty liver. The mRNA expression of autophagosome formation-related gene ULK1 was lower in the liver of dairy cows with severe fatty liver. However, the expression of other autophagosome formation-related genes, beclin 1 (BECN1), phosphatidylinositol 3-kinase catalytic subunit type 3 (PIK3C3), autophagy-related gene (ATG) 3, ATG5, and ATG12, did not differ. More important, ubiquitinated proteins, protein expression of sequestosome-1 (SQSTM1, also called p62), and microtubule-associated protein 1 light chain 3 (MAP1LC3, also called LC3)-II was greater in cows with severe fatty liver. Transmission electron microscopy revealed an increased number of autophagosomes in the liver of dairy cows with severe fatty liver. Taken together, these results indicate that excessive lipid infiltration of the liver impairs autophagic activity that may lead to cellular damage and inflammation. Show less

The class III PI3-kinase (PIK3C3) is an enzyme responsible for the generation of phosphatidylinositol 3-phosphate (PI3P), a critical component of vesicular membrane. Here, we report that PIK3C3 deficiency in zebrafish results in intestinal injury and inflammation. In pik3c3 mutants, gut tube forms but fails to be maintained. Gene expression analysis reveals that barrier-function-related inflammatory bowel disease (IBD) susceptibility genes (e-cadherin, hnf4a, ttc7a) are suppressed, while inflammatory response genes are stimulated in the mutants. Histological analysis shows neutrophil infiltration into mutant intestinal epithelium and the clearance of gut microbiota. Yet, gut microorganisms appear dispensable as mutants cultured under germ-free condition have similar intestinal defects. Mechanistically, we show that PIK3C3 deficiency suppresses the formation of PI3P and disrupts the polarized distribution of cell-junction proteins in intestinal epithelial cells. These results not only reveal a role of PIK3C3 in gut homeostasis, but also provide a zebrafish IBD model. Show less

The ER forms contacts with other endomembrane systems to exchange materials (e.g., calcium and lipids) and also to modulate dynamic organelle processes, including fission, cargo sorting and movement. During autophagosome formation, dynamic contacts between the ER and the phagophore membrane are crucial for phagophore expansion and closure. Little is known about the mechanisms underlying the formation and disassembly of the ER contacts. We found that the ER-localized autophagy protein EPG-3/VMP1 plays an essential role in controlling ER-phagophore dissociation and also the disassembly of ER contacts with LDs, mitochondria and endolysosomes. VMP1 regulates the ER contact by activating the ER calcium channel ATP2A/SERCA (ATPase sarcoplasmic/endoplasmic reticulum Ca2+ transporting). CALM (calmodulin) acts as one of the downstream calcium effectors that controls the PIK3C3/VPS34 phosphatidylinositol (PtdIns) 3-kinase (PtdIns3K) activity to maintain these contacts. Our study provides insights into the molecular mechanisms which regulate ER contacts and generate autophagosomes. Show less

The protein level of OCT4, a core pluripotency transcription factor, is vital for embryonic stem cell (ESC) maintenance, differentiation, and somatic cell reprogramming. However, how OCT4 protein levels are controlled during reprogramming remains largely unknown. Here, we identify ubiquitin conjugation sites of OCT4 and report that disruption of WWP2-catalyzed OCT4 ubiquitination or ablation of Wwp2 significantly promotes the efficiency of pluripotency induction from mouse embryonic fibroblasts. Mechanistically, disruption of WWP2-mediated OCT4 ubiquitination elevates OCT4 protein stability and H3K4 methylation level during the reprogramming process. Furthermore, we reveal that OCT4 directly activates expression of Ash2l-b, and that ASH2L-B is a major isoform of ASH2L highly expressed in ESCs and required for somatic cell reprogramming. Together, this study emphasizes the importance of ubiquitination manipulation of the reprogramming factor and its interplay with the epigenetic regulator for successful reprogramming, opening a new avenue to improve the efficiency of pluripotency induction. Show less

ANGPTL4 (angiopoietin-like protein 4) is a LPL (lipoprotein lipase) inhibitor and is present in high-density lipoprotein (HDL). However, it is not defined whether ANGPTL4 in HDLs could affect HDL metabolism and function in type 2 diabetes mellitus (T2DM). ANGPTL4 levels in the circulation and HDLs were quantified in nondiabetic participants (n=201, 68.7% females) and T2DM patients (n=185, 66.5% females). HDLs were isolated from nondiabetic controls and T2DM patients to assess cholesterol efflux Physically, ANGPTL4 in HDLs protected HDLs from hydrolysis. Resulting from increased circulating ANGPTL4 levels in T2DM, ANGPTL4 levels in HDLs were elevated but with compromised inhibitory effect on EL, leading to increased HDL hydrolysis and dysfunction. Show less

A data-independent acquisition (DIA)/parallel reaction monitoring (PRM) workflow was implemented to identify improved ovarian cancer biomarkers. Data-independent acquisition on ovarian cancer versus control sera and literature searches identified 50 biomarkers and indicated that apolipoprotein A-IV (ApoA-IV) is the most significantly differentially regulated protein. Parallel reaction monitoring with Targeted Ovarian Cancer Proteome Assay validated differential ApoA-IV expression and quantified 9 other biomarkers. Random Forest (RF) analyses achieved 92.3% classification accuracy and confirmed ApoA-IV as the leading biomarker. Indeed, all samples were classified correctly with an [ApoA-IV] breakpoint. The next best biomarkers were C-reactive protein, transferrin, and transthyretin. The Targeted Ovarian Cancer Proteome Assay suggests that ApoA-IV is a more reliable biomarker than had been determined by immunological assays and it is a better biomarker than ApoA-I, which is in the OVA1 test for ovarian cancer. This research provides a PRM/RF approach together with 4 promising biomarkers to speed the development of a clinical assay for ovarian cancer. Show less

ApoA4 exerts anti-inflammatory effects, but the mechanism remains unclear. SERPINA3 is a member of the serine proteinase inhibitor gene family, and has been shown to be involved in anti-inflammation and associated with a number of human diseases. In this study, we revealed that ApoA4 stimulates the gene expression of SERPINA3 in mouse hepatocytes both in vivo and in vitro, in a dose- and time-dependent manner. The transcriptional response of SERPINA3 to ApoA4 is regulated through the binding of ApoA4 with nuclear receptors NR4A1 and NR1D1 on the SERPINA3 promoter, which was verified with ChIP, Luciferase activity assay and RNA interference-mediated NR4A1 or NR1D1 gene knockdown. These data suggests that ApoA4 transcriptionally induced SERPINA3 expression via NR1D1 and NR4A1. Our findings may throw light on the function of ApoA4 in inflammatory responses and acute-phase reactions, as well as the development of SERPINA3 relative diseases. Show less

Hypocalcemia is an important metabolic disease of dairy cows during the transition period, although the effect of hypocalcemia on biological function in dairy cows remains unknown. In this study, proteomic, mass spectrum, bioinformatics and western blotting were employed to identify differentially expressed proteins related to serum Ca concentration. Serum samples from dairy cows were collected at three time points: 3rd days before calving (day -3), the day of calving (day 0), and 3rd days after calving (day +3). According to the Ca concentration on day 0, a total of 27 dairy cows were assigned to one of three groups (clinical, subclinical, and healthy). Samples collected on day -3 were used for discovery of differentially expressed proteins, which were separated and identified via proteomic analysis and mass spectrometry. Bioinformatics analysis was performed to determine the function of the identified proteins (gene ontology and pathway analysis). The differentially expressed proteins were verified by western blot analysis. There were 57 differential spots separated and eight different proteins were identified. Vitamin D-binding protein precursor (group-specific component, GC), alpha-2-macroglobulin (A2M) protein, and apolipoprotein A-IV were related to hypocalcemia by bioinformatics analysis. Due to its specific expression (up-regulated in clinical hypocalcemia and down-regulated in subclinical hypocalcemia), A2M was selected for validation. The results were consistent with those of proteomic analysis. A2M was as an early detection index for distinguishing clinical and subclinical hypocalcemia. The possible pathogenesis of clinical hypocalcemia caused by GC and apolipoprotein A-IV was speculated. The down-regulated expression of GC was a probable cause of the decrease in calcium concentration. Show less

Recent genome-wide association studies (GWAS) have identified multiple loci associated with coronary artery disease (CAD) among predominantly Europeans. However, their relevance to multi-ethnic populations from Southeast Asia is largely unknown. We performed a meta-analysis of four GWAS comprising three Chinese studies and one Malay study (Total N = 2,169 CAD cases and 7,376 controls). Top hits (P < 5 × 10 Show less

Xuezhikang (XZK), an extract of Chinese red yeast rice, is recommended as an optimal choice for patients with coronary heart disease (CHD) with markedly elevated triglyceride (TG) levels. This study was designed to compare the hypotriglyceridemic effects between XZK and simvastatin. The role of apolipoprotein A5 (apoA5), a key regulator of TG metabolism and a target gene of peroxisome proliferator-activated receptor α (PPARα), was to be identified in XZK-related hypotriglyceridemic actions. For these goals, hypertriglyceridemia of rats was induced by a high-fructose diet. In order to investigate the hypotriglyceridemic effects of XZK and simvastatin on these animals based on an equivalent low-density lipoprotein cholesterol (LDL-C) lowering power, we titrated their doses (XZK 80 mg/kg/d versus simvastatin 1 mg/kg/d) according to plasma LDL-C reduction of rats. Similarly, we titrated the target doses of the two agents (XZK 500 μg/ml versus simvastatin 10 μM) according to hepatocyte LDL receptor expressions, and then compared the effects of the two agents on TG and apoA5 of hepatocytes in vitro. Our results showed that XZK (80 mg/kg/d) had higher hypotriglyceridemic performance than simvastatin (1 mg/kg/d) on these animals albeit their equivalent LDL-C lowering power. Higher plasma apoA5 levels and hepatic apoA5 expressions were observed in rats treated with XZK (80 mg/kg/d) than simvastatin (1 mg/kg/d). Further, XZK (80 mg/kg/d) contributed to higher hepatic PPARα expressions of rats than simvastatin (1 mg/kg/d). Although the two agents led to an equivalent up-regulation of LDL receptors of hepatocytes, more TG reduction and apoA5 elevation were detected in hepatocytes treated with XZK (500 μg/ml) than simvastatin (10 μM). However, PPARα knockdown eliminated the above effects of XZK on hepatocytes. Therefore, our study indicates that XZK has greater hypotriglyceridemic performance than simvastatin in the setting of an equivalent LDL-C lowering power, which is attributed to more apoA5 up-regulation by this agent via the PPARα signaling pathway. Show less

It has previously been demonstrated that apolipoprotein A5 (apoA5) can be internalized by human adipocytes and significantly decreases intracellular triglyceride content. In the present study, endocytosis of apoA5 by adipocytes under different conditions, and the underlying mechanism by which apoA5 regulates cellular triglyceride storage, was investigated. The results revealed that the apoA5 protein was detected in human subcutaneous abdominal adipose tissues. In addition, the uptake of apoA5 was attenuated in human obese adipose tissues and in cultured adipocytes with hypertrophy or insulin resistance. Low‑density lipoprotein receptor protein 1 (LRP1) knockdown in adipocytes resulted in a decrease in internalized apoA5 content, suggesting that LRP1 serves a role in apoA5 uptake. Treatment of adipocytes with apoA5 decreased the expression of the lipid droplet‑associated proteins such as cidec and perilipin. ApoA5‑treated adipocytes demonstrated an increase in lipolysis activity and expression of uncoupling protein 1, which is the molecular effector of thermogenesis in brown adipocytes. These results suggested that decreased triglyceride accumulation in adipocytes induced by apoA5 may be associated with enhanced lipolysis and energy expenditure, which may result from reduced expression of cidec and perilipin. In conclusion, the present study demonstrated a novel role of apoA5 in regulating the intracellular triglyceride metabolism of adipocytes. The results of the present study suggested that apoA5 may serve as a potential therapeutic target for the treatment of obesity and its related disorders. Show less

A major goal of biomedicine is to understand the function of every gene in the human genome. Loss-of-function mutations can disrupt both copies of a given gene in humans and phenotypic analysis of such 'human knockouts' can provide insight into gene function. Consanguineous unions are more likely to result in offspring carrying homozygous loss-of-function mutations. In Pakistan, consanguinity rates are notably high. Here we sequence the protein-coding regions of 10,503 adult participants in the Pakistan Risk of Myocardial Infarction Study (PROMIS), designed to understand the determinants of cardiometabolic diseases in individuals from South Asia. We identified individuals carrying homozygous predicted loss-of-function (pLoF) mutations, and performed phenotypic analysis involving more than 200 biochemical and disease traits. We enumerated 49,138 rare (<1% minor allele frequency) pLoF mutations. These pLoF mutations are estimated to knock out 1,317 genes, each in at least one participant. Homozygosity for pLoF mutations at PLA2G7 was associated with absent enzymatic activity of soluble lipoprotein-associated phospholipase A2; at CYP2F1, with higher plasma interleukin-8 concentrations; at TREH, with lower concentrations of apoB-containing lipoprotein subfractions; at either A3GALT2 or NRG4, with markedly reduced plasma insulin C-peptide concentrations; and at SLC9A3R1, with mediators of calcium and phosphate signalling. Heterozygous deficiency of APOC3 has been shown to protect against coronary heart disease; we identified APOC3 homozygous pLoF carriers in our cohort. We recruited these human knockouts and challenged them with an oral fat load. Compared with family members lacking the mutation, individuals with APOC3 knocked out displayed marked blunting of the usual post-prandial rise in plasma triglycerides. Overall, these observations provide a roadmap for a 'human knockout project', a systematic effort to understand the phenotypic consequences of complete disruption of genes in humans. Show less

We investigated simultaneously traditional and novel lipid indices, alone or in combination, in predicting coronary severity assessed by Gensini score (GS) in 1605 non-lipid-lowering-drug-treated patients undergoing coronary angiography. Firstly, levels of triglycerides (TG), total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), non high density lipoprotein cholesterol (non-HDL-C), apolipoprotein (apo) B, lipoprotein (a) [Lp(a)], proprotein convertase subtilisin/kexin type 9 (PCSK9), apoC3, small dense LDL (sdLDL) and large HDL were increased, while HDL-C and apoA1 levels were decreased as GS status (all p for trend <0.05). However, gender stratification analyses showed similar associations between lipids and GS in men but not in women. Secondly, multiple logistic regression analyses indicated that the 12 indices were predictive for high GS (≥24) but not for low GS (1-23) compared with normal coronary (GS = 0) except for TG (neither) and apoB (both). Finally, we found that interactions between two indices with mutually exclusive composition were positively associated with GS status except for couples of TC + apoC3, apoB/PCSK9/apoC3 + sdLDL-C. Concordant elevations in the two showed the highest predictive values for high GS (all p for trend <0.05). Therefore, lipid biomarkers were associated with coronary severity and their adverse changes in combination emerged greater risks in men but not in women. Show less

Plasma levels of proprotein convertase subtilisin/kexin type 9 (PCSK9), apolipoprotein C-III (apoC3) and small dense low density lipoprotein cholesterol (sdLDL-C), have been recently recognized as circulating atherosclerosis-related lipid measurements. We aimed to elucidate their associations with current dyslipidemias, and identify their levels at increased risk to dyslipidemia. A total of 1,605 consecutive, non-treated patients undergoing diagnostic/interventional coronary angiography were examined. Plasma PCSK9 and apoC3 levels were determined using a validated ELISA assay, and sdLDL-C was measured by the Lipoprint LDL System. Plasma levels of PCSK9, apoC3, and sdLDL-C were associated with the current dyslipidemias classification (all p<0.001). PCSK9 significantly conferred prediction of both hypercholesterolemia and combined hyperlipidemia at a level of 235 ng/ml; apoC3 levels for hypertriglyceridemia, hypercholesterolemia and combined hyperlipidemia were 80.0, 71.5, and 86.4 μg/ml, respectively; and sdLDL-C for hypertriglyceridemia, hypercholesterolemia, combined hyperlipidemia and hypo high density lipoprotein (HDL) cholesterolemia 3.5, 2.5, 4.5, and 2.5 mg/dl, respectively (all p<0.001 for area under the receiver-operating characteristic curve). In a polytomous logistic model comparing increasing LDL-C categories, the interactions with high PCSK9, apoC3, and sdLDL-C elevated gradually. Similarly, apoC3 and sdLDL-C showed elevated interaction with increased triglyceride categories, and only sdLDL-C showed interaction with decreased HDL cholesterol (HDL-C) categories. Furthermore, discordances of PCSK9, apoC3, and sdLDL-C with current dyslipidemias were observed. PCSK9, apoC3, and sdLDL-C showed significant interactions with current dyslipidemias, and were predictive in the screening. The substantial discordances with current dyslipidemias might provide novel view in lipid management and further cardiovascular benefit. Show less

Colorectal cancer (CRC) is one of the most common malignancies worldwide with poor prognosis. Studies have showed that abnormal microRNA (miRNA) expression can affect CRC pathogenesis and development through targeting critical genes in cellular system. However, it is unclear about which miRNAs play central roles in CRC's pathogenesis and how they interact with transcription factors (TFs) to regulate the cancer-related genes. To address this issue, we systematically explored the major regulation motifs, namely feed-forward loops (FFLs), that consist of miRNAs, TFs and CRC-related genes through the construction of a miRNA-TF regulatory network in CRC. First, we compiled CRC-related miRNAs, CRC-related genes, and human TFs from multiple data sources. Second, we identified 13,123 3-node FFLs including 25 miRNA-FFLs, 13,005 TF-FFLs and 93 composite-FFLs, and merged the 3-node FFLs to construct a CRC-related regulatory network. The network consists of three types of regulatory subnetworks (SNWs): miRNA-SNW, TF-SNW, and composite-SNW. To enhance the accuracy of the network, the results were filtered by using The Cancer Genome Atlas (TCGA) expression data in CRC, whereby we generated a core regulatory network consisting of 58 significant FFLs. We then applied a hub identification strategy to the significant FFLs and found 5 significant components, including two miRNAs (hsa-miR-25 and hsa-miR-31), two genes (ADAMTSL3 and AXIN1) and one TF (BRCA1). The follow up prognosis analysis indicated all of the 5 significant components having good prediction of overall survival of CRC patients. In summary, we generated a CRC-specific miRNA-TF regulatory network, which is helpful to understand the complex CRC regulatory mechanisms and guide clinical treatment. The discovered 5 regulators might have critical roles in CRC pathogenesis and warrant future investigation. Show less

Stress-induced cardiomyocyte apoptosis contributes to the pathogenesis of a variety of cardiovascular diseases, but how stress induces cardiomyocyte apoptosis remains largely unclear. The present study aims to investigate the effects of Axin1 up-regulated 1 (Axud1), a novel pro-apoptotic protein, on the cardiomyocyte survival and the underlying mechanisms. To this end, a rat model under restraint stress (RS) was established and in vitro stress-induced cardiomyocytes culture was achieved. Our data showed that Axud1 was upregulated in the rat myocardia after exposure to RS. Anti-apoptotic Bcl-2 was decreased, whereas pro-apoptotic Bax and Cleaved caspase-3 (Cc3) were increased in a time-dependent manner. The Wnt/β-catenin signaling was observed to be interestingly activated in heart undergoing RS. In addition, the treatment of norepinephrine (NE) to in vitro cardiomyocytes increased Axud1 level and induced cell apoptosis. Wnt/β-catenin signaling was consistently activated. Knockdown of Axud1 using specific siRNA blunted NE-induced cardiomyocytes apoptosis and also inactivated the Wnt/β-catenin signaling. XAV-939, an inhibitor of Wnt/β-catenin signaling, partially reversed the pro-apoptotic effect of NE. In conclusion, Axud1 accelerated stress-induced cardiomyocytes apoptosis through activation of Wnt/β-catenin signaling pathway. Our data provided novel evidence that therapeutic strategies against Axud1 or Wnt/β-catenin signaling might be promising in relation to RS-induced myocardial injury. Show less

Whole-genome and exome sequence data can be cost-effectively generated for the detection of rare-variant (RV) associations in families. Causal variants that aggregate in families usually have larger effect sizes than those found in sporadic cases, so family-based designs can be a more powerful approach than population-based designs. Moreover, some family-based designs are robust to confounding due to population admixture or substructure. We developed a RV extension of the generalized disequilibrium test (GDT) to analyze sequence data obtained from nuclear and extended families. The GDT utilizes genotype differences of all discordant relative pairs to assess associations within a family, and the RV extension combines the single-variant GDT statistic over a genomic region of interest. The RV-GDT has increased power by efficiently incorporating information beyond first-degree relatives and allows for the inclusion of covariates. Using simulated genetic data, we demonstrated that the RV-GDT method has well-controlled type I error rates, even when applied to admixed populations and populations with substructure. It is more powerful than existing family-based RV association methods, particularly for the analysis of extended pedigrees and pedigrees with missing data. We analyzed whole-genome sequence data from families affected by Alzheimer disease to illustrate the application of the RV-GDT. Given the capability of the RV-GDT to adequately control for population admixture or substructure and analyze pedigrees with missing genotype data and its superior power over other family-based methods, it is an effective tool for elucidating the involvement of RVs in the etiology of complex traits. Show less

High levels of plasma high-density lipoprotein-cholesterol (HDL-C) are inversely associated with the risk of atherosclerosis and other cardiovascular diseases; thus, pharmacological inhibition of cholesteryl ester transfer protein (CETP) is considered to be a therapeutic method of raising HDL-C levels. However, many CETP inhibitors have failed to achieve a clinical benefit despite raising HDL-C. In the study, we generated transgenic (Tg) rabbits that overexpressed the human CETP gene to examine the influence of CETP on the development of atherosclerosis. Both Tg rabbits and their non-Tg littermates were fed a high cholesterol diet for 16 weeks. Plasma lipids and body weight were measured every 4 weeks. Gross lesion areas of the aortic atherosclerosis along with lesional cellular components were quantitatively analyzed. Overexpression of human CETP did not significantly alter the gross atherosclerotic lesion area, but the number of macrophages in lesions was significantly increased. Overexpression of human CETP did not change the plasma levels of total cholesterol or low-density lipoprotein cholesterol but lowered plasma HDL-C and increased triglycerides. These data revealed that human CETP may play an important role in the development of atherosclerosis mainly by decreasing HDL-C levels and increasing the accumulation of macrophage-derived foam cells. Show less

Cholesteryl ester transfer protein (CETP) has been identified as a potential target for cardiovascular disease (CVD) for its important role in the reverse cholesteryl transfer (RCT) process. In our previous work, compound Show less

N,N-Substituted amine derivatives were designed by utilizing a bioisosterism strategy. Consequently, twenty-two compounds were synthesized and evaluated for their inhibitory activity against CETP. Structure-activity relationship (SAR) studies indicate that hydrophilic groups at the 2-position of the tetrazole and 3,5-bistrifluoromethyl groups on the benzene ring provide important contributions to the potency. Among these compounds, compound 17 exhibited excellent CETP inhibitory activity (IC Show less

The association between cholesteryl ester transfer protein (CETP) TaqIB polymorphism and ischemic stroke (IS) risk has generated conflicting results. To investigate whether the TaqIB polymorphism of the CETP gene was associated with the risk of IS, a meta-analysis was performed. Studies were retrieved by searching PubMed, Web of Science, the Chinese National Knowledge Infrastructure, the Chinese Wanfang Database, and the Chinese VIP Database before January 16, 2017. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were used to assess the association. Depending on the heterogeneity the fixed-effects model or the random-effects model was used. A total of 6 case-control studies were identified with 1494 cases and 1370 controls. Overall, an association of CETP TaqIB polymorphism with IS was found in the 4 genetic models (B2B2 versus B1B1: OR = .63, 95% CI = .51-.79, P < .001; B1B2 + B2B2 versus B1B1: OR = .75, 95% CI = .64-.87, P < .001; B2B2 versus B1B2 + B1B1: OR = .70, 95% CI = .57-.85, P < .001; B2 versus B1: OR = .78, 95% CI = .70-.87, P < .001). In the subgroup analysis by ethnicity, similar risks were also observed in Asian population. This meta-analysis indicates that CETP TaqIB polymorphism is associated with IS risk, and the B2 allele is a protective factor for IS. Show less

Clinical trials have been conducted for the neuronal ceroid lipofuscinoses (NCLs), a group of neurodegenerative lysosomal diseases that primarily affect children. Whereas clinical rating systems will evaluate long-term efficacy, biomarkers to measure short-term response to treatment would be extremely valuable. To identify candidate biomarkers, we analyzed autopsy brain and matching CSF samples from controls and three genetically distinct NCLs due to deficiencies in palmitoyl protein thioesterase 1 (CLN1 disease), tripeptidyl peptidase 1 (CLN2 disease), and CLN3 protein (CLN3 disease). Proteomic and biochemical methods were used to analyze lysosomal proteins, and, in general, we find that changes in protein expression compared with control were most similar between CLN2 disease and CLN3 disease. This is consistent with previous observations of biochemical similarities between these diseases. We also conducted unbiased proteomic analyses of CSF and brain using isobaric labeling/quantitative mass spectrometry. Significant alterations in protein expression were identified in each NCL, including reduced STXBP1 in CLN1 disease brain. Given the confounding variable of post-mortem changes, additional validation is required, but this study provides a useful starting set of candidate NCL biomarkers for further evaluation. Show less

The lack of high-throughput methods to analyze the adipose tissue protein composition limits our understanding of the protein networks responsible for age and diet related metabolic response. We have developed an approach using multiple-dimension liquid chromatography tandem mass spectrometry and extended multiplexing (24 biological samples) with tandem mass tags (TMT) labeling to analyze proteomes of epididymal adipose tissues isolated from mice fed either low or high fat diet for a short or a long-term, and from mice that aged on low Show less

This study documents the disparate therapeutic effect of N-carbamyl-l-glutamate (NCG) in the activation of two different disease-causing mutants of carbamyl phosphate synthetase 1 (CPS1). We investigated the effects of NCG on purified recombinant wild-type (WT) mouse CPS1 and its human corresponding E1034G (increased ureagenesis on NCG) and M792I (decreased ureagenesis on NCG) mutants. NCG activates WT CPS1 sub-optimally compared to NAG. Similar to NAG, NCG, in combination with MgATP, stabilizes the enzyme, but competes with NAG binding to the enzyme. NCG supplementation activates available E1034G mutant CPS1 molecules not bound to NAG enhancing ureagenesis. Conversely, NCG competes with NAG binding to the scarce M792I mutant enzyme further decreasing residual ureagenesis. These results correlate with the respective patient's response to NCG. Particular caution should be taken in the administration of NCG to patients with hyperammonemia before their molecular bases of their urea cycle disorders is known. Show less

Amyloid-β (Aβ) is a key neuropathological hallmark of Alzheimer's disease (AD). Postsynaptic density protein 93 (PSD-93) is a key scaffolding protein enriched at postsynaptic sites. The aim of the present study was to examine whether PSD-93 overexpression could alleviate Aβ-induced cognitive dysfunction in APPswe/PS1dE9 (APP/PS1) mice by reducing Aβ levels in the brain. The level of PSD-93 was significantly decreased in the hippocampus of 6-month-old APP/PS1 mice compared with that in wild-type mice. Following lentivirus-mediated PSD-93 overexpression, cognitive function, synaptic function, and amyloid burden were investigated. The open field test, Morris water maze test, and fear condition test revealed that PSD-93 overexpression ameliorated spatial memory deficits in APP/PS1 mice. The facilitation of long-term potentiation induction was observed in APP/PS1 mice after PSD-93 overexpression. The expression of somatostatin receptor 4 (SSTR4) and neprilysin was increased, while the amyloid plaque load and Aβ levels were decreased in the brains of APP/PS1 mice. Moreover, PSD-93 interacted with SSTR4 and affected the level of SSTR4 on cell membrane, which was associated with the ubiquitination. Together, these findings suggest that PSD-93 attenuates spatial memory deficits and decreases amyloid levels in APP/PS1 mice, which might be associated with Aβ catabolism, and overexpression of PSD-93 might be a potential therapy for AD. Show less

BACKGROUND Mutations of DNA topoisomerase II (TOP2A) are associated with chemotherapy resistance, whereas dual-specificity phosphatase 6 (DUSP6) negatively regulates members of the mitogen-activated protein (MAP) kinase superfamily to control cell proliferation. This study assessed TOP2A and DUSP6 single nucleotide polymorphisms (SNPs) in non-small cell lung cancer (NSCLC) patients for association with chemoradiotherapy responses and prognosis. MATERIAL AND METHODS A total of 140 Chinese patients with histologically confirmed NSCLC were enrolled and subjected to genotyping of TOP2A rs471692 and DUSP6 rs2279574 using Taqman PCR. An independent sample t test was used to analyze differences in tumor regression after radiotherapy versus SNP risk factors. Kaplan-Meier curves analyzed overall survival, followed by the log-rank test and Cox proportional hazard models. RESULTS There were no significant associations of TOP2A rs471692 and DUSP6 rs2279574 polymorphisms or clinicopathological variables with response to chemoradiotherapy (p>0.05). Comparing overall survival of 87 patients with stage I-III NSCLC treated with radiotherapy or chemoradiotherapy to clinicopathological variables, the data showed that tumor regression, weight loss, clinical stage, and cigarette smoking were independent prognostic predictors (p=0.009, 0.043, 0.004, and 0.025, respectively). Tumor regression rate >0.34 was associated with patent survival versus tumor regression rate ≤0.34 (p=0.007). CONCLUSIONS TOP2A rs471692 and DUSP6 rs2279574 SNPs were not associated with chemoradiotherapy response, whereas tumor regression, weight loss, clinical stage, and cigarette smoking were independent prognostic predictors for these Chinese patients with NSCLC. Show less

Multiple osteochondromas (MO) is an autosomal skeletal disease with an elusive molecular mechanism. To further elucidate the genetic mechanism of the disease a three‑generation Chinese family with MO was observed and researched, and a novel frameshift mutation (c.335₃₃₆insA) in the exotosin 1 (EXT1) gene of one patient with MO was observed through exome sequencing. This was further validated by Sanger sequencing and comparison with 200 unrelated healthy controls. Immunohistochemistry and multiple sequence alignment were performed to determine the pathogenicity of the candidate mutation. Multiple sequence alignment suggested that codon 335 and 336 in the EXT1 gene were highly conserved regions in vertebrates. Immunohistochemistry revealed that EXT1 protein expression levels were decreased in a patient with MO and this mutation compared with a patient with MO who had no EXT1 mutation. Owing to the appearance of c.335₃₃₆insA in exon 1 of EXT1, a premature stop codon was introduced, resulting in truncated EXT1. As a result integrated and functional EXT1 was reduced. EXT1 is involved in the biosynthesis of heparan sulfate (HS), an essential molecule, and its dysfunction may lead to MO. The novel mutation of c.335₃₃₆insA in the EXT1 gene reported in the present study has enlarged the causal mutation spectrum of MO, and may assist genetic counseling and prenatal diagnosis of MO. 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

Noncompaction cardiomyopathy is characterized by the presence of extensive trabeculations, which could lead to heart failure and malignant arrhythmias. How trabeculations resolve to form compact myocardium is poorly understood. Elucidation of this process is critical to understanding the pathophysiology of noncompaction disease. Here we use genetic lineage tracing to mark the Nppa Show less

Vascular smooth muscle cell proliferation, migration, and dedifferentiation are critical for vascular diseases. Recently, it was demonstrated that Notch receptors have opposing effects on intima formation after vessel injury. Therefore, it is important to investigate the specific regulatory pathways that activate the different Notch receptors. There was a time- and dose-dependent activation of Notch1 by angiotensin II and platelet-derived growth factor in vascular smooth muscle cells. When phospholipase Cγ1 (PLCγ1) expression was reduced by small interfering RNA, Notch1 activation and Hey2 expression (Notch target gene) induced by angiotensin II or platelet-derived growth factor were remarkably inhibited, while Notch2 degradation was not affected. Mechanistically, we observed an association of PLCγ1 and Akt, which increased after angiotensin II or platelet-derived growth factor stimulation. PLCγ1 knockdown significantly inhibited Akt activation. Importantly, PLCγ1 phospholipase site mutation (no phospholipase activity) did not affect Akt activation. Furthermore, PLCγ1 depletion inhibited platelet-derived growth factor-induced vascular smooth muscle cell proliferation, migration, and dedifferentiation, while it increased apoptosis. In vivo, PLCγ1 and control small interfering RNA were delivered periadventitially in pluronic gel and complete carotid artery ligation was performed. Morphometric analysis 21 days after ligation demonstrated that PLCγ1 small interfering RNA robustly attenuated intima area and intima/media ratio compared with the control group. PLCγ1-Akt-mediated Notch1 signaling is crucial for intima formation. This effect is attributable to PLCγ1-Akt interaction but not PLCγ1 phospholipase activity. Specific inhibition of the PLCγ1 and Akt interaction will be a promising therapeutic strategy for preventing vascular remodeling. Show less