👤 Lee Zou

LINGO-1 (leucine-rich repeat and Ig domain-containing, Nogo receptor-interacting protein) is an important component of the NgR receptor complex involved in RhoA activation and axon regeneration. The authors report on passive immunization with LINGO-1 polyclonal antiserum, a therapeutic approach to overcome NgR-mediated growth inhibition after spinal cord injury (SCI). The intrathecally administered high-titer rabbit-derived antiserum can be detected around the injury site within a wide time window; it blocks LINGO-1 in vivo with high molecular specificity. In this animal model, passive immunization with LINGO-1 antiserum significantly decreased RhoA activation and increased neuronal survival. Adult rats immunized in this manner show recovery of certain hindlimb motor functions after dorsal hemisection of the spinal cord. Thus, passive immunotherapy with LINGO-1 polyclonal antiserum may represent a promising repair strategy following acute SCI. Show less

Numerous single nucleotide polymorphisms (SNPs) have been found in recent genome wide association studies (GWAS) to be associated with subtle plasma triglyceride (TG) variation in normolipidemic subjects. However, since these GWAS did not specifically evaluate patients with rare disorders of lipoprotein metabolism--'hyperlipoproteinemia' (HLP)--it remains largely unresolved whether any of these SNP determinants of modest physiological changes in TG are necessarily also determinants of most HLP phenotypes. To address this question, we evaluated 28 TG-associated SNPs from GWAS in 386 unrelated adult patients with one of five Fredrickson phenotypes (HLP types 2A, 2B, 3, 4 and 5) and 242 matched normolipidemic controls. We found that several SNPs associated with TG in normolipidemic samples, including APOA5 p.S19W and -1131T>C, TRIB1 rs17321515, TBL2 rs17145738, GCKR rs780094, GALNT2 rs4846914 and ANGPTL3 rs12130333, were significantly associated with HLP types 2B, 3, 4 and 5. The findings indicate that: (i) the TG-associated Fredrickson HLP types 2B, 3, 4 and 5 are polygenic traits; (ii) these Fredrickson HLP types share numerous genetic determinants among themselves; and (iii) genetic determinants of modest TG variation in normolipidemic population samples also underlie--to an apparently even greater degree--susceptibility to these rare HLP phenotypes. Thus, the TG-associated Fredrickson HLP types 2B, 3, 4 and 5, although historically considered to be distinct are actually complex traits sharing among them several common genetic determinants seen in GWAS of normolipidemic population samples. Show less

To express and purify the fusion protein of extracellular domain of human Ig domain-containing, neurite outgrowth inhibitor (Nogo) receptor-interacting protein-1 (LINGO-1(aa76-319)) in prokaryotic cells and prepare the rabbit anti-LINGO-1 polyclonal antibody (pAb). The 732 bp DNA sequence of hLINGO-1(aa76-319) was obtained from pCMV-SPORT6 by PCR and inserted into pET30a(+) plasmid to construct the prokaryotic expression plasmid pET30a(+)-hLINGO-1(aa76-319), which was subsequently transformed into E.coli. The target fusion protein was expressed with IPTG induction and purified by Ni(2+)-NTA affinity chromatography column. The antiserum against hLINGO-1(aa76-319) was obtained from the rabbits immunized with hLINGO-1(aa76-319), and the titer of the pAb was determined using enzyme linked immunosorbent assay (ELISA) and its specificity identified using Western blotting. The prokaryotic expression plasmid pET30a(+)-hLINGO-1(aa76-319) was constructed successfully. Efficient expression of the target fusion protein was achieved with IPTG induction at the optimal concentration of 0.4 mmol/L and culture temperature at 37 degrees celsius; for 2.5 h. The hLINGO-1(aa76-319) fusion protein was effectively expressed in E.coli as inclusion bodies, and the soluble protein was obtained through denaturation and refolding procedures, and the purified fusion protein showed a purity above 90%. The titer of the anti-hLINGO-1(aa76-319) pAb obtained by immunizing the rabbits with the purified protein reached 1:1.6x10(6), and Western blotting confirmed its good specificity. The fusion protein hLINGO-1(aa76-319) with high purity has been obtained and the anti-hLINGO-1(aa76-319) pAb obtained shows a high titer and good specificity, which provide important experimental basis for further functional investigation of LINGO-1. Show less

To identify the disease-causing gene mutations and to reveal the relationship between the genotype and the phenotype in Chinese patients with hypertrophic cardiomyopathy (HCM). One hundred unrelated patients with HCM and 120 controls were enrolled in this study. The full encoding exons and flanking sequences of the cardiac myosin binding protein C gene (MYBPC3) were amplified with PCR and the products were sequenced. A novel missense mutation c.706T > C was identified in exon 6 of MYBPC3 gene in three HCM patients, which resulted a Serine (S) to Glycine (G) exchange at amino acid residue 236 (S236G). The clinical phenotypes of the three patients were different (2 obstructive HCM, 1 non-obstructive HCM). The 120 controls were normal in the genetic test. The novel S236G mutation in MYBPC3 gene was a hot-spot mutation in Chinese patients with HCM. Show less

To investigate the genotype-phenotype correlation in Chinese familial hypertrophic cardiomyopathy (HCM), peripheral blood samples were collected from 7 members of a Chinese HCM family, and 120 normal subjects were recruited as control. The full encoding exons and flanking sequences of the cardiac troponin T (TNNT2) gene, beta-myosin heavy chain (MYH7) gene and myosin binding protein C (MYBPC3) gene were amplified and the products were sequenced directly to detect the mutations. A missense mutation, c.1273G>A, was identified in exon 14 of the MYH7 gene in 4 members of the Chinese HCM family, which resulted a glycine (Gly) to arginine (Arg) exchange at amino acid residue 425. The 425th glycine amino acid residue is highly conservative across the different species. The clinical phenotypes among the family members who carried this mutation presented significant individual differences. The c.1273G>A mutation of the MYH7 gene might be the causal mutation of the familial HCM. The heterogeneity of phenotypes suggested that multiple factors may be involved in the pathogenesis of HCM. Show less

The significance of transcription factors PPAR alpha, LXR alpha, and their responsive/target genes for the pathogenesis of atherosclerosis in apolipoprotein E and low-density lipoprotein receptor double deficient (AL) mice fed with high fat and cholesterol (HF) diet were studied. C57BL/6J wild-type (WT) mice were used as control to the AL mice. Plasma lipid metabolites and morphological atherosclerotic lesions in aortic wall were determined. Semi- and real-time quantitative RT-PCR were used to measure gene expression patterns between AL mice and the controls, which were fed with HF or normal chow diet. The results showed that in AL mice fed with HF diet, plasma lipid levels, hepatic lipid accumulation, and atherogenesis together with upregulated PPAR alpha, LXR alpha, and their target genes, i.e., FAT, SCD1, FAS, Angptl3, and apoB100 significantly increased in a 12-week long feeding period. In contrast, apoAI, apoAIV, apoF, LPL, and SR-BI were decreased compared to chow-fed group. In WT mice, PPAR alpha, LXR alpha, FAS, Angpt13, CPT1, apoF, ACOX1, LPL, and SR-BI were increased with HF treatment, while apoAI and apoAIV were decreased markedly. The different changes of lipid metabolism-related genes between AL and WT mice, fed with HF diet or chow diet indicated that the mechanisms of dietary effects on gene mutant mice are different from those of intact WT mice. Since lipid metabolic system defected genetically in AL mice, we suggest that the changes of PPAR alpha, LXR alpha, and their target genes aggravated lipid metabolic disorder in the liver and further accelerated the development of atherosclerosis on a stress of HF diet feeding in AL mice. Show less

Recent genome-wide association (GWA) studies have identified new genetic determinants of complex quantitative traits, including plasma triglyceride (TG). We hypothesized that common variants associated with mild TG variation identified in GWA studies would also be associated with severe hypertriglyceridemia (HTG). We studied 132 patients of European ancestry with severe HTG (fasting plasma TG > 10 mmol/l), who had no mutations found by resequencing of candidate genes, and 351 matched normolipidemic controls. We determined genotypes for: GALNT2 rs4846914, TBL2/MLXIPL rs17145738, TRIB1 rs17321515, ANGPTL3 rs12130333, GCKR rs780094, APOA5 rs3135506 (S19W), APOA5 rs662799 (-1131T > C), APOE (isoforms) and LPL rs328 (S447X). We found that: (i) genotypes, including those of APOA5 S19W, APOA5 -1131T > C, APOE, GCKR, TRIB1 and TBL2/MLXIPL, were significantly associated with severe HTG; (ii) odds ratios for these genetic variables were significant in both univariate and multivariate regression analyses, irrespective of the presence or absence of diabetes or obesity; (iii) a significant fraction-about one-quarter-of the explained variation in disease status was associated with these genotypes. Therefore, common SNPs (single nucleotide polymorphisms) that are associated with mild TG variation in GWA studies of normolipidemic subjects are also associated with severe HTG. Our findings are consistent with the emerging model of a complex genetic trait. At the extremes of a quantitative trait, such as severe HTG, are found the cumulative contributions of both multiple rare alleles with large genetic effects and common alleles with small effects. Show less

To study the disease-causing gene mutations in familial hypertrophic cardiomyopathy (HCM) in Chinese and to reveal the relationship between the genotype and the phenotype. Peripheral blood samples were collected from 12 members of a HCM family, and 120 healthy volunteers in China. PCR and double deoxygenation chain termination method were used to analyze the cardiac troponin T gene (TNNT2), beta-myosin heavy chain gene (MYH7) gene and myosin binding protein C gene (MYBPC3) and to detect mutations. Mutation G14452A was identified in exon 22 of MYH7 gene in 4 family members, causing the conversion of glycine (G) into glutamic acid (E). The onset ages and clinical manifestations of the family members carrying the mutation G823E, including 2 patients (the proband, male, with the onset age of 51, and his 26-year-old second son with the onset age of 20), and 2 carriers (his 31-year-old elder son and 29-year-old elder daughter), presented significant individual differences. The G823E mutation of MYH7 gene is the causal mutation of familial HCM. The heterogeneity of phenotypes suggests that multiple factors may be involved in the pathogenesis of HCM. Show less

To reveal genotype-phenotype correlation of disease-causing gene mutations in Chinese hypertrophic cardiomyopathy (HCM) pedigree. Peripheral venous blood samples were collected from two Chinese HCM families and 120 healthy subjects were recruited as normal control. The full encoding exons and flanking sequences of the cardiac troponin T gene (TNNT2), beta-myosin heavy chain gene (MYH7) and myosin binding protein C gene (MYBPC3) were amplified with the polymerase chain reaction method, DNA sequencing was used to detect the mutation. In ZZJ family, mutation G12101A was identified in exon 21 of MYBPC3 gene in 4 family members [the arginine (R) converted to histidine (H)]. In this pedigree, three out of eight family members were diagnosed as HCM and with a penetrance of 75%. In FHL family, mutation G15391A was identified in exon 23 of MYH7 gene in 3 family members [the glutamic acid (E) converted to lysine (K)]. In this pedigree, three out of six family members were diagnosed as HCM and with a penetrance of 100%. Echocardiography showed obstruction of left ventricular outflow tract in two out of the three HCM patients. Our results showed that the G12101A mutation of MYBPC3 gene is the causal mutation of familial HCM with mild phenotype. The G15391A mutation of MYH7 gene is the causal mutation of familial HCM with malignant phenotype and a penetrance of 100%. Screening mutations in the MYH7 gene should be viewed as a reasonable procedure in obstructive HCM patients. Show less

No data are available on survival analysis and longitudinal evolution of patients with gene mutations of beta-myosin heavy chain (MYH7) and myosin binding protein C (MYBPC3) in Chinese. To prospectively investigate whether different gene mutations confer distinct prognosis. We performed a prospective study in 70 HCM patients and 46 genetically affected family members without HCM-phenotype with direct DNA sequencing of MYH7 and MYBPC3, clinical assessments, and 5.8 +/- 1.8 years follow-up. After follow-up, more surgical intervention (8/52 versus 0/18, p < 0.001), higher sudden death risk (7/52 versus 0/18, p < 0.001) and shorter life span were found in patients with MYH7 mutations than in patients with MYBPC3 mutations (45.1 +/- 14.0 versus 73.5 +/- 7.5 years, p = 0.03). Seven of the 27 mutation carriers of MYH7 had clinical presentations of HCM, but no carriers of MYBPC3 mutations developed to HCM during follow-up. Maximal wall thickness was thicker in the patients carrying mutations in the global region of MYH7 than in those carrying mutations in the rod region of MYH7 (21.5 +/- 6.6 versus 15 +/- 6.1 mm, p < 0.05) at baseline. More sudden death (7/41 versus 0/11) and left ventricular dysfunction (NYHA Class III approximately IV, 17/32 versus 1/10) were identified in patients with mutations in the global region of MYH7 than in patients with other mutations. MYH7 mutations, especially in the global region, cause malignant clinical phenotypes. Show less

To study the disease-causing gene mutation in Chinese patients with familial hypertrophic cardiomyopathy (FHC) and to analyze the correlation between the genotype and the phenotype. Peripheral blood samples were collected from 40 members from a family affected with FHC, and 120 healthy volunteers. PCR was performed to analyze the exons and flanking introns of the cardiac troponin T gene (TNNT2), beta-myosin heavy chain gene (MYH7), and myosin-binding protein C gene (MYBPC3) and the products were sequenced. The clinical data including symptom, physical examination, echocardiography and electrocardiography were collected. A 14035c > t mutation, which causes a missense mutation (R130C) in exon 10 of TNNT2 gene were identified in 4 family members, including the proband, female, aged 53, with the onset at the age of 30. The 4 persons with the 14035c > t mutation, all FHC patients, presented left ventricular dysfunction with a penetrance of 100%. Two of the patients died of sudden cardiac death during follow-up. No mutation was identified in the MYH7 and MYBPC3 genes. The 14035c > t mutation of TNNT2 gene is the causal mutation of FHC which is associated with malignant phenotype with a penetrance of 100%. It is a reasonable procedure in HCM patients with malignant phenotype to screen mutation in the TNNT2 gene. Show less

To study the disease-causing gene mutation in Chinese patients with hypertrophic cardiomyopathy (HCM) and to analyze the genotype and phenotype correlation. One family (n = 27) affected with HCM were chosen for the study. The full encoding exons and flanking sequences of beta-myosin heavy chain gene (MYH7) and cardiac myosin-binding protein C gene (MYBPC3) were amplified with PCR and the products were sequenced. The clinical data including symptom, physical, echocardiography and electrocardiography examinations were collected. We identified a 13261 G > A mutation, which causes a missense mutation (G758D) in exon 23 of MYBPC3 in 9 family members. One mutation carrier suffered from dilated cardiomyopathy (DCM) with asymmetric interventricular septal hypertrophy (14 mm). Another mutation carrier was diagnosed as HCM. The 13261 G > A mutation is associated with a DCM-like HCM and HCM phenotype in this Chinese family affected with HCM. Show less

To detect the disease-causing gene mutation of hypertrophic cardiomyopathy (HCM) in a Chinese family and to analyze the correlation of the genotype and the phenotype. One family affected with HCM was studied. The clinical data including symptom, physical examination, echocardiography and electrocardiography were collected. The full encoding exons and flanking sequences of beta-myosin heavy chain gene (MYH7) and cardiac myosin-binding protein C gene (MYBPC3) were amplified with PCR and the products were sequenced. A G8887A mutation, which is an acceptor splicing site of intron 15 (IVS15-1G > A) in MYBPC3 (gi: Y10129) was identified in 6 out of 11 family members. Three mutation carriers developed HCM at 48 - 75 years old with mild chest pain, chest distress and asymmetric septal hypertrophy (13 - 14 mm) and remaining mutation carriers are free of HCM. No mutation was identified in MYH7 gene. HCM caused by the IVS15-1G > A mutation is a benign phenotype. It is helpful to screen MYBPC3 gene mutation in late-onset HCM patients with mild symptoms. Show less

Clinical phenotype of hypertrophic cardiomyopathy exhibits significant inter- and intra-familial heterogeneities. To test if MYBPC3 polymorphism could modify the expression of cardiac hypertrophy, 226 patients with hypertrophic cardiomyopathy and 226 age- and sex-matched controls were recruited according to the diagnostic criteria of WHO. Genotyping was completed by using PCR, restrictive enzyme digestion, and sequencing. Three polymorphisms of MYBPC3 were studied, only the GG genotype at 18443 in exon 30 associated with thicker left ventricular wall (25.2+/-5.9 mm) in patient group, not the AA and AG genotypes (19.0+/-5.0mm, P<0.001). After multiple regression analysis for adjustment of age and sex, the association remained. No difference was found in the genotype distribution between control and patients. Our results point out that GG genotype of MYBPC3 might be a genetic risk factor for the expression of cardiac hypertrophic phenotype in the patients with hypertrophic cardiomyopathy. Show less

There are more than 1 million patients with hypertrophic cardiomyopathy (HCM) in China, but the genetic basis is presently unknown. We investigated 100 independent patients with HCM (proband 51, sporadic 49) by sequencing the three most frequent HCM-causing genes (MYH7, MYBPC3, TNNT2). Thirty-four patients (34%) carried 25 types of mutations in the selected genes, most (14/25) were newly identified. MYH7 and MYBPC3 accounted for 41% and 18% of the familial HCM, respectively. TNNT2 mutations only caused 2% of the familial HCM. These results suggested that MYH7 and MYBPC3 were the predominant genes responsible for HCM, and TNNT2 mutation less proportionally contributed to Chinese HCM. MYH7 mutations caused HCM at younger age, more frequent syncope and ECG abnormalities compared with MYBPC3 mutations. The patients carrying R663C, Q734P, E930K in MYH7 and R130C in TNNT2 expressed malignant phenotype. R403Q in MYH7, the most common hot and malignant mutation in Caucasians, was not identified in Chinese. We confirmed the diversity of mutation profile in different populations and suggest that a global registry of HCM mutations and their phenotypes is necessary to correlate genotype with phenotype. Show less