Association of β-adrenergic receptor genes polymorphisms with incidence of subsequent cardiovascular events in Han Chinese patients with coronary artery disease / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Sequence variants in the β-adrenergic receptor (ADRB) genes have a close relationship with the development of coronary artery disease (CAD) and the patient's prognosis. However, there is a lack of data on the role of the variants in ADRBs genes in Han Chinese patients with CAD. We aimed to investigate the association of genetic variants in the ADRB1 and ADRB2 genes with the incidence of major adverse cardiac event (MACE) in Han Chinese patients with CAD.</p><p><b>METHODS</b>A total of 545 Han Chinese patients with CAD undergoing percutaneous coronary intervention (PCI) were recruited to the study and followed for one year. Three variant sites in ADRB1 (rs1801253) and ADRB2 (rs1042713 and rs1042714) were genotyped. The effect of the ADRB1 and ADRB2 genotypes on MACE within one year was assessed.</p><p><b>RESULTS</b>There were 47 cases of MACE during follow-up. There was no significant difference in the incidence of MACE among patients carrying different genotypes of the three variants in ADRB1 and ADRB2 (Log-rank, all P > 0.05). Cox regression analysis showed no association between three variants in ADRB1 and ADRB2 genes and the incidence of MACE during one-year follow-up, the adjusted hazard ratios (95% confidence interval) for rs1801253, rs1042713 and rs1042714 were 1.05 (0.54-2.02), 1.24 (0.58-2.64) and 1.66 (0.81-3.42), respectively.</p><p><b>CONCLUSION</b>Our data did not support a relationship between the three polymorphisms of ADRB1 (rs1801253) and ADRB2 (rs1042713 and rs1042714) genes and risk of subsequent cardiovascular events after PCI in Han Chinese patients with CAD.</p>

Screening and identification of auto-antigen RHDAG1 of rheumatic heart disease / 南方医科大学学报

<p><b>OBJECTIVE</b>To identify the candidate auto-antigen of rheumatic heart disease as a molecular marker for this disease.</p><p><b>METHODS</b>The total RNA of the heart tissue of patients with rheumatic heart disease was extracted and reverse-transcribed into long cDNA to construct the phage expression library. The library was screened using the serum from patients with active rheumatic fever, and the positive clone was identified and analyzed by bioinformatics and expressed in vitro. The expressed products were evaluated with Western blotting and its cross-reactivity was assessed.</p><p><b>RESULTS</b>The phage expression library of the heart tissue of patients with rheumatic heart disease was constructed, with the titer of the primary library of 3.3×10(6) pfu/ml, recombinant rate of 99%, and 81% of the inserted segments were larger than 1 kb. An auto-antigen RHDAG1 was identified by screening, which was homologous to keratin 18. RHDAG1 was detected in the serum of patients with active rheumatic fever and of those with rheumatic heart disease, but not in the serum of healthy subjects.</p><p><b>CONCLUSION</b>Phage display library can be an effective strategy to screen the auto-antigens of rheumatic heart disease. The auto-antigen RHDAG1 can be a candidate molecular biomarker of rheumatic heart disease and/or rheumatic fever.</p>

siRNAs targetting sphingomyelin phosphodiesterase 1 protect mouse oocytes from apoptosis / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the potential of siRNAs targeting sphingomyelin phosphodiesterase 1 (SMPD1) in protecting the oocytes from apoptosis, and explore new approaches to female fertility preservation.</p><p><b>METHODS</b>Chemically synthesized siRNA targeting SMPD1 were introduced into mouse oocytes retrieved by hyperstimulation, and the cell apoptosis was analyzed by comic assay 48 and 72 h later.</p><p><b>RESULTS</b>In the oocytes without any siRNA injection, oocyte DNA damage occurred after 24 h, and large amount of DNA fragments migrated from the cells 48 h later. In oocytes injected with siRNA003, DNA migration decreased significantly as compared with the control and the other two groups injected with siRNA001 and siRNA002 (P<0.01).</p><p><b>CONCLUSION</b>siRNA targeting SMPD1 may protect the oocytes from apoptosis, and has the potential for use in future female fertility preservation.</p>

Induced differentiation and signaling factor PTEN expression of 3T3-L1 adipocytes / 南方医科大学学报

<p><b>OBJECTIVE</b>To optimize the condition for inducing the differentiation of 3T3-L1 preadipocytes into adipocytes and study the expression of PTEN tumor suppression gene in this process, aiming to understand the regulatory role of PTEN in normal adipocyte differentiation and collect laboratory evidence for developing drugs targeting PTEN.</p><p><b>METHODS</b>The differentiation of 3T3-L1 preadipocytes cultured in high-glucose DMEM were induced according to 2 protocols with different combinations of dexamethasone, isobutylmethylxanthine (IBMX) and insulin, and the resultant adipocytes were identified by oil red O staining. The total proteins of 3T3-L1 were extracted and analyzed by Western blotting, and PTEN homology between mice and human was analyzed by bioinformatic method.</p><p><b>RESULTS</b>For optimized 3T3-L1 differentiation, 3T3-L1 cells were initially induced with the combination of 1 micromol/L dexamethasone, 0.5 mmol/L IBMX and 5 microg/ml insulin for 48 h, followed by treatment with 5 microg/ml insulin in 4.5 g/L glucose DMEM for 48 h, which resulted in high differentiation rate of 3T3-L1 cells (up to 90% on the 10th day) with unified morphology and size. PTEN expression varied quantitatively in the process of differentiation, especially low on the 12th day as compared with those measured on days 4, 6 and 9. The mice PTEN mRNA shared 96% homology and PTEN amino acid 100% homology with their human counterparts.</p><p><b>CONCLUSION</b>Endogenous PTEN expression is down-regulated during 3T3-L1 differentiation, suggesting that PTEN may enhance insulin sensitivity and promote adipogenesis under physiological conditions.</p>

Identification and tissue localization of intermediate filament protein in Angiostrongylus cantonensis / 南方医科大学学报

<p><b>OBJECTIVE</b>To identify the type of the intermediate filament (IF) protein of Angiostrongylus cantonensis and analyze its tissue localization.</p><p><b>METHODS</b>Recombinant pET-IF of antigen IF was expressed in E.coli with IPTG induction, and the expression products were purified by His.Bind column and identified for determining the type of the IF protein by Western blotting. Anti-IF antibody was prepared by multi-spot subcutaneous injection into mouse and used to detect the tissue slices of A. cantonensis by immunohistochemical analysis.</p><p><b>RESULTS</b>The antigen IF were correctly expressed and purified, and identified as a keratin located in the intestine wall and cytoplusma.</p><p><b>CONCLUSION</b>The antigen IF is distributed in the intestine wall of A. cantonensis.</p>

Schistosoma japonicum: construction of phage display antibody library and its application in the immunodiagnosis of infection / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>A monoclonal antibody would be an effective tool for the detection of circulating antigens in the serum of patients with schistosomiasis, but the traditional way of producing monoclonal antibodies is not cost-effective. The objective of this study was to find a new method for the large-scale production of monoclonal antibodies against Schistosoma japonicum (Sj).</p><p><b>METHODS</b>A phage display antibody library for Sj was constructed. To obtain a single-chain variable fragment antibody (scFv) against Sj, the library was screened with metabolic antigens from adult Sj worms (Sj-MAg) using enzyme-linked immunosorbent assay. The soluble scFvs selected were used to detect Sj antigens in the serum of acute and chronic schistosomiasis patients.</p><p><b>RESULTS</b>Six positive clones with good reactivity to Sj-MAg were obtained from the phage display antibody library of about 1.07 x 10(6) individual clones. Only two of these six clones bound specifically to Sj-MAg and were chosen for further analysis. Specific soluble anti-Sj-MAg scFvs were produced by inducing the 2 clones with isopropyl-D-thiogalactopyranoside. The characteristics of the scFvs were then determined. The results of Western blot showed that these scFvs could bind to Sj-MAg specifically and had a molecular weight of about 31 kD. When testing serum from schistosomiasis patients with one of the two specific scFvs, its sensitivity was found to be 60% and 37% in acute and chronic patients, respectively, with a specificity of 90%. When the two specific scFvs were combined, their sensitivity was found to be 75% and 57% in acute and chronic patients, respectively, with a specificity of 85%.</p><p><b>CONCLUSIONS</b>The results indicate that the scFvs are potentially useful for the diagnosis of schistosomiasis. The library construction also provides a useful tool for the further screening of other antibodies for both diagnostic and immunotherapeutic applications and for epitope analysis and vaccine design.</p>