👤 Li Luo

Genome-wide association studies have identified single-nucleotide polymorphisms (SNPs) that increase the risk of developing non-alcoholic fatty liver disease (NAFLD). One purpose of this study was to determine the frequencies of NAFLD susceptibility SNPs in a non-Hispanic white and Hispanic population who attended a clinic in northeast Albuquerque, NM. Another goal was to determine associations with selected indicators in this New Mexican population. This cohort study involving 168 volunteer subjects in the NM population (88 non-Hispanic whites, 63 Hispanics, 4 Native Americans, 11 Asian Americans, 2 unreported ethnicity). Eight SNPs within 6 NAFLD susceptibility genes including PNPLA3 (rs738409), LYPLAL1 (rs12137855), APOC3 (rs2854116, rs2854117), GCKR (rs780094, rs741038), FABP2 (rs1799883), PEMT (rs7946) were analyzed by genotyping using the TaqMan genotyping assay (Applied Biosystems, Foster City, CA). Statistical analyses were carried out using statistical package SAS 9.3. The NAFLD allele frequencies were similar in non-Hispanic whites and Hispanics except for PNPLA3 (rs738409), FABP2 (rs1799883), and PEMT (rs7946). Eight SNPs in 5 NAFLD susceptibility genes were significantly associated OR marginally associated with selected indicators for NAFLD, metabolic syndrome, overweight, obesity, insulin resistance, type 2 diabetes, hypertension, dyslipidemia. No SNPs were significantly associated with the same indicator in both the non-Hispanic white and Hispanic groups. In this population of non-Hispanic whites and Hispanics, there were only heterozygotes for the APOC3 derived alle le whereas for all other genes tested, both heterozygotes and homozygotes were found. Associations of alleles with indicators of chronic disease were different in non-Hispanic whites compared to Hispanics. Show less

To explore the role of CD44 in monocyte adhesion to human brain microvascular endothelial cells (HBMECs) and monocyte migration across an in vitro model of blood-brain barrier (BBB) infected by Cryptococcus neoformans (Cn). An in vitro blood-brain barrier model was constructed using a transwell chamber covered with a HBMEC monolayer. The wild-type strain of Cn B4500FO2, TYCC645#32 strain with CPS1 gene deletion and PCIP strain with CPS1 complementation were chosen to infect the monolayer HBMECs. THP-1 cells were added to the upper chamber of transwell, and the relative migration rate was determined by counting the number of the cells entering the lower chambers. The inhibitory effects of anti-CD44 monoclonal antibody and the CD44 inhibitor bikunin were examined on THP-1 binding to and migration across HBMECs. Cn infection of the HBMECs caused markedly enhanced THP-1 cell adhesion and migration across the monolyers (P<0.01) dependent on Cn concentration and exposure time. Addition of anti-CD44 monoclonal antibody and bikunin significantly lowered THP-1 adhesion and migration rates in the BBB model with Cn-infected HBMECs (P<0.01) with a dose dependence of the antibody (within 0-1 µg) and inhibitor (within 0-20 nmol/L). Both THP-1 adhesion rate and migration rate were lowered in the BBB model infected with CPS1 gene-deleted Cn but increased in the model infected with the complemented strain compared with those in the wild-type strain-infected model. In the in vitro BBB model, CD44 expressed on HBMECs may play an essential role in monocyte adhesion to and migration across the BBB. The capsular hyaluronic acid may mediate Cn-induced monocyte adhesion and migration. Show less

MicroRNAs (miRNAs) are small, non-coding RNAs that modulate gene expression by negatively regulating the stability or translational efficiency of their target mRNAs. The aim of this study was to investigate the expression of microRNA-145 (miR-145) in human papillary thyroid cancer and its potential function. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to determine the expression level of miR-145 in ten papillary thyroid cancer and adjacent normal specimens. The function of miR-145 overexpression on the proliferation of human TPC1 thyroid cancer cells was conducted by MTT assays and by colony-formation assays. Western blot was used to validate the impact of miR-145 on the protein expression of the target gene. Luciferase reporter assays were employed to validate a putative target of miR-145. MiR-145 expression was relatively decreased in papillary thyroid cancer specimens compared with adjacent normal tissues (P<0.05). MTT assays and colony-formation assays showed that overexpression of miR-145 suppressed TPC1 cell growth. Luciferase assays using a reporter carrying a putative miR-145 target site in the 3' untranslated region of DUSP6 revealed that miR-145 directly targets DUSP6. Overexpression of miR-145 led to downregulation of DUSP6 at protein level as assessed by Western blot. Targeted knockdown of DUSP6 by siRNA significantly inhibited the proliferation of TPC1 cells. The overexpression of miR-145 inhibited TPC1 cellular growth by targeting DUSP6; this finding implies a better understanding of initiation and progression of papillary thyroid cancer. Show less

To describe the expression profiles of FOXA1, DUSP6, and HA117 in different portions of the colon of patients diagnosed with Hirschsprung's disease (HSCR). Colon specimens were collected from 34 HSCR patients and grouped into 3 segments: proximal anastomosis, dilated segment and stenotic segment. Levels of FOXA1, DUSP6, and HA117 RNA were evaluated by real-time PCR. Levels of FOXA1 and DUSP6 protein were analyzed by immunohistochemistry and Western blotting. The levels of FOXA1 and DUSP6 RNA were significantly lower in the stenotic segment compared to proximal anastomosis (P < 0.05). The level of HA117 RNA was significantly higher in the stenotic segment compared to proximal anastomosis (P < 0.05). In proximal anastomosis, FOXA1 and DUSP6 were both expressed at the protein level in ganglion cells and nerve fibers between the circular and longitudinal muscles. In the stenotic segments, positive staining for FOXA1 and DUSP6 was diminished. The levels of FOXA1 and DUSP6 protein were significantly lower in the stenotic segment compared to proximal anastomosis (P < 0.05). Suppression of the FOXA1/DUSP6 signaling pathway may contribute to the development of HSCR. LncRNA HA117 may have an anti-differentiation function, and play a pivotal role in the progression of HSCR. Show less

The fruiting body of Ganoderma lucidum has been used as a traditional herbal medicine for many years. However, to the date, there is no detailed study for describing the effect of G. lucidum spores on oxidative stress, blood glucose level and lipid compositions in animal models of type 2 diabetic rats, in particular the effect on the gene expression profiles associated with glucose and lipid metabolisms. G. lucidum spores powder (GLSP) with a shell-broken rate >99.9 % was used. Adult male Sprague-Dawley rats were randomly divided into three groups (n = 8/group). Group 1: Normal control, normal rats with ordinary feed; Group 2: Model control, diabetic rats with ordinary feed without intervention; Group 3: GLSP, diabetic rats with ordinary feed, an intervention group utilizing GLSP of 1 g per day by oral gavages for 4 consecutive weeks. Type 2 diabetic rats were obtained by streptozocin (STZ) injection. The changes in the levels of glucose, triglycerides, total cholesterol and HDL-cholesterol in blood samples were analyzed after GLSP intervention. Meanwhile, gene expressions associated with the possible molecular mechanism of GLSP regulation were also investigated using a quantitative RT-PCR. The reduction of blood glucose level occurred within the first 2 weeks of GLSP intervention and the lipid synthesis in the diabetic rats of GLSP group was significantly decreased at 4 weeks compared to the model control group. Furthermore, it was also found that GLSP intervention greatly attenuated the level of oxidative stress in the diabetic rats. Quantitative RT-PCR analysis showed up-regulation of lipid metabolism related genes (Acox1, ACC, Insig-1 and Insig-2) and glycogen synthesis related genes (GS2 and GYG1) in GLSP group compared to model control group. Additionally, there were no significant changes in the expression of other genes, such as SREBP-1, Acly, Fas, Fads1, Gpam, Dgat1, PEPCK and G6PC1. This study might indicate that GLSP consumption could provide a beneficial effect in terms of lowering the blood glucose levels by promoting glycogen synthesis and inhibiting gluconeogenesis. Meanwhile, GLSP treatment was also associated with the improvement of blood lipid compositions through the regulation of cholesterol homeostasis in the type 2 diabetic rats. Show less

Both genetic predisposition and lifestyle factors are associated with the risk for obesity. Multiple obesity loci have been identified using genome-wide association studies mainly in European populations. The aims of this study were to examine the associations of these loci with obesity and gene×dietary behavior interactions among Chinese children and adolescents. Nineteen candidate SNPs were genotyped using Sequenom technology in the Chinese children (N=2977, 853 obese and 2124 controls, aged 7-17). Dietary behaviors were assessed using self-administered questionnaires. After adjusting for age, sex and multiple testing, MC4R rs17782313, SEC16B rs543874, MAP2K5 rs2241423 and KCTD15 rs11084753 were associated with obesity and obesity-related traits (all P<0.005), with odd ratios ranging from 1.22 to 2.15. Dose-response association was significant between genetic risk score, which was calculated by summing the risk alleles, and the risk of obesity (P<0.001). Multiplicative interaction was found between rs543874 and salt preference on obesity with an OR of 4.40 (95% CI, 1.12-17.30). Additive interactions with salt preference were found in rs17782313 and rs11084753. Our findings indicated that rs17782313, rs543874, rs2241423 and rs11084753 were associated with the risk for children obesity in China, and interaction of genetic variants with diet behaviors on obesity. Show less

Here, we investigated the role of LXRα in capsaicin mediated anti-inflammatory effects. Results revealed that capsaicin inhibits LPS-induced IL-1β, IL-6 and TNF-α production in a time- and dose-dependent manner. Moreover, capsaicin increases LXRα expression through PPARγ pathway. Inhibition of LXRα activation by siRNA diminished the inhibitory action of capsaicin on LPS-induced IL-1β, IL-6 and TNF-α production. Additionally, LXRα siRNA abrogated the inhibitory action of capsaicin on p65 NF-κB protein expression. Thus, we propose that the anti-inflammatory effects of capsaicin are LXRα dependent, and LXRα may potentially link the capsaicin mediated PPARγ activation and NF-κB inhibition in LPS-induced inflammatory response. Show less

Specificity protein 1 (SP1) is a ubiquitous transcription factor that plays an important role in controlling gene expression. Although important in mediating the function of various hormones, the role of SP1 in regulating milk fat formation remains unknown. To investigate the sequence and expression information, as well as its role in modulating lipid metabolism, we cloned SP1 gene from mammary gland of Xinong Saanen dairy goat. The full-length cDNA of the SP1 gene is 4376 bp including 103 bp of 5'UTR, 2358 bp of ORF (HM₂₃₆₃₁₁₎ and 1915 bp of 3'UTR, which is predicted to encode a 786 amino acids polypeptide. Phylogenetic tree analysis showed that goat SP1 has the closest relationship with sheep, followed by bovines (bos taurus, odobenus and ceratotherium), pig, primates (pongo, gorilla, macaca and papio) and murine (rattus and mus), while the furthest relationship was with canis and otolemur. Expression was predominant in the lungs, small intestine, muscle, spleen, mammary gland and subcutaneous fat. There were no significant expression level differences between the mammary gland tissues collected at lactation and dry-off period. Overexpression of SP1 in goat mammary epithelial cells (GMECs) led to higher mRNA expression level of peroxisome proliferator-activated receptor-γ (PPARγ) and lower liver X receptor α (LXRα) mRNA level, both of which were crucial in regulating fatty acid metabolism, and correspondingly altered the expression of their downstream genes in GMECs. These results were further enhanced by the silencing of SP1. These findings suggest that SP1 may play an important role in fatty acid metabolism. Show less

Mutations in CLN3 gene cause juvenile neuronal ceroid lipofuscinosis (JNCL or Batten disease), an early-onset neurodegenerative disorder that is characterized by the accumulation of ceroid lipofuscin within lysosomes. The function of the CLN3 protein remains unclear and is presumed to be related to Endoplasmic reticulum (ER) stress. To investigate the function of CLN3 in the ER stress signaling pathway, we measured proliferation and apoptosis in cells transfected with normal and mutant CLN3 after treatment with the ER stress inducer tunicamycin (TM). We found that overexpression of CLN3 was sufficient in conferring increased resistance to ER stress. Wild-type CLN3 protected cells from TM-induced apoptosis and increased cell proliferation. Overexpression of wild-type CLN3 enhanced expression of the ER chaperone protein, glucose-regulated protein 78 (GRP78), and reduced expression of the proapoptotic protein CCAAT/-enhancer-binding protein homologous protein (CHOP). In contrast, overexpression of mutant CLN3 or siRNA knockdown of CLN3 produced the opposite effect. Together, our data suggest that the lack of CLN3 function in cells leads to a failure of management in the response to ER stress and this may be the key deficit in JNCL that causes neuronal degeneration. Show less

Retinitis pigmentosa (RP) is a devastating form of retinal degeneration, with significant social and professional consequences. Molecular genetic information is invaluable for an accurate clinical diagnosis of RP due to its high genetic and clinical heterogeneity. Using a gene capture panel that covers 163 of the currently known retinal disease genes, including 48 RP genes, we performed a comprehensive molecular screening in a collection of 123 RP unsettled probands from a wide variety of ethnic backgrounds, including 113 unrelated simplex and 10 autosomal recessive RP (arRP) cases. As a result, 61 mutations were identified in 45 probands, including 38 novel pathogenic alleles. Interestingly, we observed that phenotype and genotype were not in full agreement in 21 probands. Among them, eight probands were clinically reassessed, resulting in refinement of clinical diagnoses for six of these patients. Finally, recessive mutations in CLN3 were identified in five retinal degeneration patients, including four RP probands and one cone-rod dystrophy patient, suggesting that CLN3 is a novel non-syndromic retinal disease gene. Collectively, our results underscore that, due to the high molecular and clinical heterogeneity of RP, comprehensive screening of all retinal disease genes is effective in identifying novel pathogenic mutations and provides an opportunity to discover new genotype-phenotype correlations. Information gained from this genetic screening will directly aid in patient diagnosis, prognosis, and treatment, as well as allowing appropriate family planning and counseling. Show less

Class XIX myosin (Myo19) is a vertebrate-specific unconventional myosin, responsible for the transport of mitochondria. To characterize biochemical properties of Myo19, we prepared recombinant mouse Myo19-truncated constructs containing the motor domain and the IQ motifs using the baculovirus/Sf9 expression system. We identified regulatory light chain (RLC) of smooth muscle/non-muscle myosin-2 as the light chain of Myo19. The actin-activated ATPase activity and the actin-gliding velocity of Myo19-truncated constructs were about one-third and one-sixth as those of myosin-5a, respectively. The apparent affinity of Myo19 to actin was about the same as that of myosin-5a. The RLCs bound to Myo19 could be phosphorylated by myosin light chain kinase, but this phosphorylation had little effect on the actin-activated ATPase activity and the actin-gliding activity of Myo19-truncated constructs. Using dual fluorescence-labeled actin filaments, we determined that Myo19 is a plus-end-directed molecular motor. We found that, similar to that of the high-duty ratio myosin, such as myosin-5a, ADP release rate was comparable with the maximal actin-activated ATPase activity of Myo19, indicating that ADP release is a rate-limiting step for the ATPase cycle of acto-Myo19. ADP strongly inhibited the actin-activated ATPase activity and actin-gliding activity of Myo19-truncated constructs. Based on the above results, we concluded that Myo19 is a high-duty ratio molecular motor moving to the plus-end of the actin filament. Show less

In nonruminants, the alternative splicing of peroxisome proliferator-activated receptor γ (PPARG) generates PPARG1 and PPARG2 isoforms. Although transcriptional control differences between isoforms have been reported in human adipose tissue, their roles in ruminant mammary cells are not well known. To assess which of these isoforms is more closely associated with the regulation of mammary lipogenic pathways, their tissue distribution was analyzed and the expression of key genes regulating lipogenic gene networks was measured after overexpression of the 2 isoforms in goat mammary epithelial cells (GMEC). The expression of PPARG2 was markedly greater in adipose tissue, whereas PPARG1 is the main isoform in goat mammary tissue (ratio of PPARG1:PPARG2 was close to 37:1). As was reported in previous work, PPARG1 upregulated the transcription regulators SREBF1 and PPARG and the lipogenic genes FASN, ACACA, and SCD. Along with a tendency for greater expression of AGPAT6, DGAT1, and PLIN2, these data suggest that PPARG1 is the isoform controlling lipogenesis in mammary cells. Addition of the PPARG ligand rosiglitazone (ROSI) to GMEC overexpressing both isoforms upregulated the expression of LPL and CD36, which help control uptake of long-chain fatty acids into mammary cells. Other responses to ROSI addition to GMEC overexpressing PPARG1 and PPARG2 included upregulation of AGPAT6, DGAT1, INSIG1, SREBF1, and NR1H3. Although the data suggest that both PPARG1 and PPARG2 could affect mammary lipogenesis via control of gene expression when stimulated (e.g., by ROSI), the fact that PPARG1 is more abundant in mammary tissue and that its overexpression alone upregulated key lipogenic gene networks suggest that it is the more important isoform in goat mammary cells. Show less

To explore the relationship between the liver X receptor α gene (LXRα) rsl2221497 polymorphism and the susceptibility of coronary heart disease (CHD) and serum lipids and glucose levels. The single fluorescently labeled probes technique was used to detect the genotype of rsl2221497 in LXRα gene in 240 CHD patients and 250 healthy control subjects. The difference of genotype distribution between the two groups was analyzed using of Chi-square test. The serum lipids and glucose levels between the different genotypes were also compared. The risk of CHD in carriers with (AA + GA) genotype was 1.76 times as that in the GG genotype carriers (OR = 1.76, 95% CI: 1.18-2.87, P <0.05), and the risk of CHD in carriers with A allele increased 0.88 times compared to that in G allele carriers (OR = 1.88, 95% CI:1.21-3.43, P <0.01). Logistic regression analysis showed that after adjusting for other confounding factors, A allele was an independent risk for CHD. However, there were no differences in serum lipids and glucose levels between each genotype. The rsl2221497 polymorphism in LXRα gene was associated with susceptibility of CHD in Han population. Show less

The ubiquitin-proteasome system plays an important role in various celluar processes. WWP2, a recently identified ubiquitin E3 ligase, has been proved a multifunctional gene by degradation a series of targets via ubiquitin-dependent proteasome system, including PETN, Smads, Oct4, EGR2, TIRF and so. Hereafter, we reviewed the recent research process about the function of WWP2. Show less

SRG3 plays essential roles both in early mouse embryogenesis and in extra-embryonic vascular development. As one of the core components of the SWI/SNF-like BAF complex, SRG3 serves as the scaffold protein and its protein level controls the stability of the BAF complex, which controls diverse physiological processes through transcriptional regulation. However, little is known about how the protein level of SRG3 is regulated in mammalian cells. Previously, we identified a murine ubiquitin ligase (Wwp2) and demonstrated that it interacts with pluripotency-associated key transcription factor Oct4 and RNA polymerase II large subunit Rpb1, promoting their ubiquitination and degradation. Here, we report that Wwp2 acts as a ubiquitin ligase of SRG3. Our results show that Wwp2 and SRG3 form protein complexes and co-localize in the nucleus in mammalian cells. The interaction is mediated through the WW domain of Wwp2 and the PPPY motif of SRG3, respectively. Importantly, Wwp2 promotes ubiquitination and degradation of SRG3 through the ubiquitin-proteasome system. The expression of a catalytically inactive mutant of Wwp2 abolishes SRG3 ubiquitination. Collectively, our study opens up a new avenue to understand how the protein level of SRG3 is regulated in mammalian cells. Show less

Multiple osteochondromas (MO) is an autosomal dominant hereditary disorder caused by heterozygous germline mutations in the exostonsin-1 (EXT1) or exostosin-2 (EXT2) genes. In this study, we screened mutations in the EXT1/EXT2 genes in four Chinese MO kindreds by direct sequencing. Three point mutations were detected, including a nonsense mutation in the EXT2 gene (c.544C > T) and two splice site mutations in the EXT1 and EXT2 genes, respectively (EXT1: c.1883 + 1G > A and EXT2: c.1173 + 1G > T). Although splice site mutations constitute at least 10% of all mutations that cause MO, there has been limited research on their pathogenic effect on RNA processing due to poor availability of patient RNA samples. In this study, ex vivo and in vivo splicing assays were used to investigate the effect of EXT1 and EXT2 mutations on aberrant splicing at the mRNA level. Our results indicate that identified splice site mutations can cause either cryptic splice site usage or exon skipping. Show less

To explore the function of PPAR γ in the goat mammary gland, we cloned the whole cDNA of the PPAR γ gene. Homology alignments revealed that the goat PPAR γ gene is conserved among goat, bovine, mouse, and human. Luciferase assays revealed that rosiglitazone enhanced the activity of the PPAR γ response element (PPRE) in goat mammary epithelial cells (GMECs). After rosiglitazone (ROSI) treatment of GMECs, there was a significant (P < 0.05) increase in the expression of genes related to triacylglycerol synthesis and secretion: LPL, FASN, ACACA, PLIN3, FABP3, PLIN2, PNPLA2, NR1H3, SREBF1, and SCD. The decreases in expression observed after knockdown of PPAR γ relative to the control group (Ad-NC) averaged 65%, 52%, 67%, 55%, 65%, 58%, 85%, 43%, 50%, and 24% for SCD, DGAT1, AGPAT6, SREBF1, ACACA, FASN, FABP3, SCAP, ATGL, and PLIN3, respectively. These results provide direct evidence that PPAR γ plays a crucial role in regulating the triacylglycerol synthesis and secretion in goat mammary cells and underscore the functional importance of PPAR γ in mammary gland tissue during lactation. Show less

Fetal alcohol spectrum disorders (FASD) results from ethanol exposure to the developing fetus and is the leading cause of mental retardation. FASD is associated with a broad range of neurobehavioral deficits which may be mediated by ethanol-induced neurodegeneration in the developing brain. An immature brain is more susceptible to ethanol neurotoxicity. We hypothesize that the enhanced sensitivity of the immature brain to ethanol is due to a limited capacity to alleviate cellular stress. Using a third trimester equivalent mouse model of ethanol exposure, we demonstrated that subcutaneous injection of ethanol induced a wide-spread neuroapoptosis in postnatal day 4 (PD4) C57BL/6 mice, but had little effect on the brain of PD12 mice. We analyzed the expression profile of genes regulating apoptosis, and the pathways of ER stress response (also known as unfolded protein response, UPR) and autophagy during these ethanol-sensitive and resistant periods (PD4 versus PD12) using PCR microarray. The expression of pro-apoptotic genes, such as caspase-3, was much higher on PD4 than PD12; in contrast, the expression of genes that regulate UPR and autophagy, such as atf6, atg4, atg9, atg10, beclin1, bnip3, cebpb, ctsb, ctsd, ctss, grp78, ire1α, lamp, lc3 perk, pik3c3, and sqstm1 was significantly higher on PD12 than PD4. These results suggest that the vulnerability of the immature brain to ethanol could result from high expression of pro-apoptotic proteins and a deficiency in the stress responsive system, such as UPR and autophagy. Show less

We previously demonstrated in streptozotocin-induced diabetic mice that deficiency or inhibition of aldose reductase (AR) caused significant dephosphorylation of hepatic transcriptional factor PPARα, leading to its activation and significant reductions in serum lipid levels. Herein, we report that inhibition of AR by zopolrestat or by a short-hairpin RNA (shRNA) against AR caused a significant reduction in serum and hepatic triglycerides levels in 10-week old diabetic db/db mice. Meanwhile, hyperglycemia-induced phosphorylation of hepatic ERK1/2 and PPARα was significantly attenuated in db/db mice treated with zopolrestat or AR shRNA. Further, in comparison with the untreated db/db mice, the hepatic mRNA expression of Aco and ApoA5, two target genes for PPARα, was increased by 93% (P < 0.05) and 73% (P < 0.05) in zopolrestat-treated mice, respectively. Together, these data indicate that inhibition of AR might lead to significant amelioration in hyperglycemia-induced dyslipidemia and nonalcoholic fatty liver disease. Show less

Familial aggregation of hepatocellular carcinoma (HCC), the third leading cause of cancer death worldwide, has shown to be a common phenomenon. We investigated the association between the genetic background and HCC familial aggregation. Serum samples were collected from HCC family members and normal control family members for screening the differentially expressed protein peaks with the approach of surface-enhanced laser desorption ionization time-of-flight mass spectrometry. Potential genetically associated protein peaks were selected and further identified by matrix assisted laser desorption ionization-time of flight mass spectrometry. A panel of six protein peaks (m/z 6432.94, 8478.35, 9381.91, 17284.67, 17418.34, and 18111.04) were speculated to reflect the genetic susceptibility of HCC familial aggregation. Three of them (m/z 6432.94, 8478.35, and 9381.91) were selected to identify as the candidate proteins. Nine identified proteins, including mostly apolipoprotein family (ApoA1, ApoA2, ApoC3, ApoE) and serum amyloid A protein (SAA), were found overexpressed in the multiple HCC cases family members. The comparative proteomic profiles have suggested that genetic factors ought to be taken into account for familial aggregation of HCC. Show less

To further investigate the biological function of human novel gene CTRP4 by constructing the prokaryotic expression vector of human CTRP4, inducing the expression of and purifying hCTRP4-his protein in E.coli, and preparing polyclonal antibody against human CTRP4. Human CTRP4 gene was amplified by PCR, digested with enzymes, and subcloned into a his-tagged prokaryotic expression vector to generate a recombinant plasmid named pET-32a-hCTRP4. The pET-32a-hCTRP4 was transformed into E.coli BL21(DE3). The hCTRP4-his fusion protein was induced by IPTG, purified by Ni-NTA purification system, and analyzed by SDS-PAGE. The recombinant vector pcDNA3.1-myc/his(-)B-hCTRP4 expressing full-length human CTRP4 and purified prokaryotic protein hCTRP4 were used to immunize BALB/c mice to produce polyclonal antibody. The anti-serum was purified and the characteristics of the antibody were identified by ELISA, Western blotting, immunofluorescence cytochemistry and immunohistochemistry. The prokaryotic expression vector of pET-32a-hCTRP4 was constructed successfully. hCTRP4-his fusion protein was expressed in E.coli BL21(DE3) after IPTG induction. The titer of the anti-serum reached 1:20 000, and its specificity was proved by Western blotting. The results of immunofluorescence cytochemistry and immunohistochemistry indicated that CTRP4 was mainly localized in the cytoplasm of hepatic cells. hCTRP4-his fusion protein can be successfully expressed in E.coli. A specific polyclonal antibody against human CTRP4 has been successfully prepared. Show less

Nonalcoholic fatty liver disease (NAFLD) is associated with obesity, insulin resistance, type 2 diabetes, and dyslipidemia. The purpose of this study was to identify novel proteins and pathways that contribute to the pathogenesis and complications of NAFLD. C57BL/6J male mice were fed a 60% (HFD) or 10% (LFD) high or low fat diet. HFD induced obesity, hepatic steatosis and insulin resistance (euglycemic clamps, glucose infusion rate: LFD 50.5 ± 6.4 vs. HFD 14.2 ± 9.5 μg/ (g·min); Show less

The liver X receptor α (LXRα) is a nuclear receptor of the transcription factor and is known to play a crucial role in lipid metabolism processes such as bile acid and fatty acid synthesis in humans and rodents. However, very little information is available on the role of LXRα in the regulation of fatty acid synthesis in the goat mammary gland. In this investigation, a cDNA was isolated from the mammary gland of Xinong Saanen dairy goats and designated as goat LXRα. RT-PCR and RACE gave rise to the full-length cDNA of LXRα, which was comprised of 1654 bp and characterized by an ORF of 1344 bp and 5'- and 3'-UTR regions of 150 and 160 bp, respectively. The deduced amino acid sequence encodes 477 amino acids with a predicted molecular weight (MW) of 50.4kDa and a theoretical isoelectric point (pI) of 6.3. Additionally, homology search and sequence multi-alignment indicated that the putative goat LXRα amino acid sequence is very similar to those of cattle, mice, rats, swine, and humans. Bioinformatic predictions demonstrated that the LXRα protein is located in the nucleus, containing characteristic signatures of a nuclear receptor with DNA-binding domain (DBD) and ligand-binding domain (LBD). Real-time quantitative PCR suggested that LXRα was predominantly expressed in the small intestine, liver, spleen and mammary gland. Treatment of goat mammary gland epithelial cells (GMEC) with different concentrations (i.e., 0.01, 0.1, 1 μM) of T0901317, a synthetic agonist of LXRα, resulted in elevated sterol regulatory element binding protein 1 (SREBP1) and fatty acid synthase (FASN) mRNA levels in response to LXRα activation. The association between different T0901317 concentrations and fatty acid composition in GMEC also was examined using gas chromatography (GC). The results showed that activation of LXRα significantly increased GMEC C18:1 and C18:2 contents, but did not affect levels of saturated fatty acids (SFA). These discoveries are consistent with the notion that LXRα plays a key role in controlling lipogenesis and regulating synthesis of unsaturated fatty acids (UFA) in the mammary gland of goats, which may prove useful in regulation of milk fat production. Show less

Leptin, a key hormone in regulating energy homeostasis, is mainly produced by adipocytes. Cogent evidence indicates a unique role of leptin in the promotion of liver fibrosis. Hepatic stellate cell (HSC) activation is a pivotal step in the process of liver fibrosis. Sterol regulatory element binding protein (SREBP)-1c, a critical transcription factor for lipid synthesis and adipocyte differentiation, functions as a key transcription factor in inhibition of HSC activation. SREBP-1c is highly expressed in quiescent HSCs and downregulated upon HSC activation. The aim of this study is to examine the effect of leptin on SREBP-1c gene expression in HSCs in vitro and in vivo and elucidate the underlying mechanisms. The results of the present study demonstrated that leptin strongly inhibited SREBP-1c expression in HSCs in vivo and in vitro. p38 MAPK was involved in leptin regulation of SREBP-1c expression in cultured HSCs. Leptin-induced activation of p38 MAPK led to the decreases in liver X receptor (LXR)-α protein level, activity and its binding to the SREBP-1c promoter, which caused the downregulation of SREBP-1c expression. Moreover, leptin inhibition of SREBP-1c expression via p38 MAPK increased the expression of alpha1(I) collagen in HSCs. Our results might provide new insights into the mechanisms of the unique role of leptin in the development of liver fibrosis and might have potential implications for clarifying the molecular mechanisms underlying liver fibrosis in diseases in which circulating leptin levels are elevated such as nonalcoholic steatohepatitis, type 2 diabetes mellitus and alcoholic cirrhosis. Show less

The contribution of genetic variants to body mass index (BMI) during adolescence across multiethnic samples is largely unknown. We selected genetic loci associated with BMI or obesity in European-descent samples and examined them in a multiethnic adolescent sample. In 5103 European American (EA), 1748 African American (AfA), 1304 Hispanic American (HA) and 439 Asian American (AsA) participants of the National Longitudinal Study of Adolescent Health (Add Health; ages 12-21 years, 47.5% male), we assessed the association between 41 established obesity-related single-nucleotide polymorphisms (SNPs) with BMI using additive genetic models, stratified by race/ethnicity, and in a pooled meta-analysis sample. We also compared the magnitude of effect for BMI-SNP associations in EA and AfA adolescents to comparable effect estimates from 11 861 EA and AfA adults in the Atherosclerosis Risk in Communities study (ages 45-64 years, 43.2% male). Thirty-five of 41 BMI-SNP associations were directionally consistent with published studies in European populations, 18 achieved nominal significance (P<0.05; effect sizes from 0.19 to 0.71 kg m(-2) increase in BMI per effect allele), while 4 (FTO, TMEM18, TFAP2B, MC4R) remained significant after Bonferroni correction (P<0.0015). Of 41 BMI-SNP associations in AfA, HA and AsA adolescents, nine, three and five, respectively, were directionally consistent and nominally significant. In the pooled meta-analysis, 36 of 41 effect estimates were directionally consistent and 21 of 36 were nominally significant. In EA adolescents, BMI effect estimates were larger (P<0.05) for variants near TMEM18, PTER and MC4R and smaller for variants near MTIF3 and NRXN3 compared with EA adults. Our findings suggest that obesity susceptibility loci may have a comparatively stronger role during adolescence than during adulthood, with variation across race/ethnic subpopulation. Show less

Hepatocellular carcinoma (HCC) is believed to arise from tumor-initiating cells (T-ICs), although little is known about their stem cell-like properties. We quantified levels of p28(GANK) (Gankyrin), OV6, and Oct4 in 130 human HCC samples using immunohistochemistry. Magnetic-activated cell sorting was used to isolate OV6+ HCC cells. T-IC properties were evaluated by quantitative reverse-transcription polymerase chain reaction, flow cytometry, and spheroid formation. We used a coimmunoprecipitation assay to study interactions among p28(GANK), Oct4, and WWP2. Tumorigenicity and pulmonary metastasis were examined in nonobese diabetic and severe combined immunodeficient mice. In HCC samples, high levels of p28(GANK) correlated with expansion of OV6+ tumor cells; the combination of high levels of p28(GANK) and OV6 was associated with progression of HCC. p28(GANK) was predominantly expressed in liver T-ICs, isolated by magnetic sorting, and undifferentiated primary HCC spheroids. Increased levels of p28(GANK) in T-ICs increased their percentages in HCC samples, expression of stem cell genes, self-renewal potential, chemoresistance in vitro, and tumorigenicity and ability to develop into pulmonary metastases in mice. Conversely, knockdown of p28(GANK) reduced their T-IC properties. p28(GANK) likely activates liver T-ICs by impeding ubiquitination and degradation of the transcription factor Oct4 by WWP2. In support of this concept, levels of p28(GANK) correlated with those of Oct4 in HCC samples. p28(GANK) activates and maintains liver T-ICs in HCCs by preventing degradation of Oct4. Inhibitors of p28(GANK) might therefore be developed to inactivate T-ICs and slow tumor progression. Show less

The link between lipoprotein metabolism and Alzheimer's disease (AD) has been established. Apolipoprotein A-IV (apoA-IV), a component of lipoprotein particles similar to apolipoprotein E, has been suggested to play an important role in brain metabolism. Although there are clinical debates on the function of its polymorphism in AD, the pathologic role of apoA-IV in AD is still unknown. Here, we report that genetic ablation of apoA-IV is able to accelerate AD pathogenesis in mice. In a mouse model that overexpresses human amyloid precursor protein (APP) and presenilin 1, genetic reduction of apoA-IV augments extracellular amyloid-β peptide (Aβ) burden and aggravates neuron loss in the brain. In addition, genetic ablation of apoA-IV also accelerates spatial learning deficits and increases the mortality of mice. We have found that apoA-IV colocalizes within Aβ plaques in APP/presenilin 1 transgenic mice and binds to Aβ in vitro. Subsequent studies show that apoA-IV in this model facilitates Aβ uptake in the Aβ clearance pathway mediated by astrocytes rather than the amyloidogenic pathway of APP processing. Taken together, we conclude that apoA-IV deficiency increases Aβ deposition and results in cognitive damage in the mouse model. Enhancing levels of apoA-IV may have therapeutic potential in AD treatment. Show less

The NF-κB and IL6/STAT3 pathways are major participants in tumor-promoting inflammation. C1qTNF related protein (CTRP) is a family with multiple physiological functions, but their involvement in tumor-promoting inflammation has received little attention. For the first time, we have identified CTRP4 as a novel secretary protein by N-terminal sequencing. Moreover, recombinant CTRP4 can effectively induce the activation of both NF-κB and IL6/STAT3 signaling pathways in the pattern similar to that of classical cytokine. By western blot analysis, we detected the upregulation of CTRP4 in response to IL6. Importantly, functional research revealed that CTRP4 could promote tumor cell survival and tumor resistance against apoptosis induced by chemotherapeutics. These results strongly suggest that CTRP4 is a novel tumor-promoting inflammatory regulator. Our findings might provide a meaningful indication for cancer research. Show less

Neuronal nitric oxide synthase (nNOS) is mainly expressed in neurons, to some extent in astrocytes and neuronal stem cells. The alternative splicing of nNOS mRNA generates 5 isoforms of nNOS, including nNOS-α, nNOS-β, nNOS-µ, nNOS-γ and nNOS-2. Monomer of nNOS is inactive, and dimer is the active form. Dimerization requires tetrahydrobiopterin (BH4), heme and L-arginine binding. Regulation of nNOS expression relies largely on cAMP response element-binding protein (CREB) activity, and nNOS activity is regulated by heat shock protein 90 (HSP90)/HSP70, calmodulin (CaM), phosphorylation and dephosphorylation at Ser847 and Ser1412, and the protein inhibitor of nNOS (PIN). There are primarily 9 nNOS-interacting proteins, including post-synaptic density protein 95 (PSD95), clathrin assembly lymphoid leukemia (CALM), calcium/calmodulin-dependent protein kinase II alpha (CAMKIIA), Disks large homolog 4 (DLG4), DLG2, 6-phosphofructokinase, muscle type (PFK-M), carboxy-terminal PDZ ligand of nNOS (CAPON) protein, syntrophin and dynein light chain (LC). Among them, PSD95, CAPON and PFK-M are important nNOS adapter proteins in neurons. The interaction of PSD95 with nNOS controls synapse formation and is implicated in N-methyl-D-aspartic acid-induced neuronal death. nNOS-derived NO is implicated in synapse loss-mediated early cognitive/motor deficits in several neuropathological states, and negatively regulates neurogenesis under physiological and pathological conditions. Show less

Essential tremor (ET) has been hypothesized to be a risk factor for the development of Parkinson disease (PD). Recently, rs9652490 variant in the leucine-rich repeat and Ig domain containing 1 gene (LINGO1) was found to be associated with ET susceptibility. To evaluate whether the same variant is associated also with PD susceptibility, we investigated the association between the LINGO1 rs9652490 variant and PD phenotype in Caucasian and Chinese PD subjects. We found no significant differences in genotypic and allele distribution between patients and control subjects (χ(2)=1.931, p=0.381 for genotypic distribution; χ(2)=0.001, p=0.973 for allele distribution), suggesting this variant is not associated with PD. Show less