Cloning and expression of human interleukin-26 in Escherichia coli / 生物工程学报

To clone human interleukin-26 (hIL-26) and express it in E. coli efficiently. Two pairs of primers were synthesized according to the hIL-26 gene reported on GenBank. The hIL-26 gene was cloned by nest PCR following the first round RT-PCR from human peripherial blood monocytes total RNA, and then the PCR product was cloned into pMD18-T vector. Colony PCR, restriction analysis and sequence analysis showed that the gene cloned was the same as the reported hIL-26. The recombinant was cut with BamHI and EcoR I to obtain the hIL-26 fragment, and then the fragment was inserted into pBV220 which was cut with the same enzymes. The recombinant expression vector was induced to express hIL-26 at 42 degrees C, SDS-PAGE analysis showed that the recombinant protein accounted for up to 20% of the whole protein of E. coli, and the protein was also confirmed by Western blotting. Purity of the protein was found to be above 90% after purified with molecular sieve. After renaturalized with glutathione buffer, the promoting effect of it on the production of IFN-y in PBMC was detected by RT-PCR. A recombinant bacterial strain for expressing hIL-26 with biological activity was constructed successfully.

An analysis for the phenotype and genotype of autosomal dominant polycystic kidney disease from two Chinese families / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To scan for mutations of polycystic kidney disease 1 gene (PKD1) in Chinese population in order to find some features about Chinese patients and a better approach to detect mutations.</p><p><b>METHODS</b>Twenty-five PKD-affected individuals from twenty-one unrelated genealogies and sixteen controls participated in the study. Thirty-five blood samples and six tissues were obtained after receiving informed consent and were in accordance with institutional ethical guidelines. Genomic DNA was isolated from peripheral blood using standard procedures. PCR amplification of genomic DNA was performed to generate the aimed fragments. Amplified fragments were analyzed by denaturing gradient gel electrophoresis (DGGE). A GC clamp was attached to the 5' primer. After that, the abnormal fragments were sequenced on freshly amplified specific PCR products with the dideoxynucleotide chain termination method. Sequencing was performed for all samples to evaluate DGGE.</p><p><b>RESULTS</b>Aimed fragments of exons 44 and 45 were amplified. DGGE detected eleven abnormal PCR fragments. Two novel mutations were identified by sequencing, included one nonsense mutation (C12217T) and one frameshift (12431delCT). In addition, one polymorphism (A50747C) was identified. The mutation detection rate is 8% in our study.</p><p><b>CONCLUSION</b>Two novel pathogenic mutations were detected, including one nonsense mutation (C12217T) and one frameshift (12431delCT).</p>

Study on the (TAAAA)n repeat polymorphism in sex hormone-binding globulin gene and the SHBG serum levels in putative association with the glucose metabolic status of Chinese patients suffering from polycystic ovarian syndrome in Shandong province / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the relationship of (TAAAA)n repeat polymorphism in the promoter of the sex hormone-binding globulin (SHBG) gene and SHBG serum levels to the glucose metabolic status of Chinese polycystic ovary syndrome (PCOS) patients in Shandong province.</p><p><b>METHODS</b>GeneScan method was used to detect and identify (TAAAA)n repeat number (alleles) and genotypes for 156 controls and 157 patients who were divided into normal glucose tolerance without hyperinsulinemia (NIR group) and with hyperinsulinemia (HI group) and abnormal glucose metabolic (AGM) group according to the results of oral glucose test and insulin resistant test; IRMA was used to measure serum SHBG for part of them.</p><p><b>RESULTS</b>Five alleles containing (TAAAA) 6-10 repeats and 14 genotypes including 6/6, 6/7, 6/8, 6/9, 6/10, 7/7, 7/8, 7/9, 7/10, 8/8, 8/9, 8/10, 9/9, 9/10 repeats genotypes were present in the subjects. Genotype distribution of 6/10 repeats genotype is lower in PCOS than that in control, and 8/9 repeats genotype vice versa (P < 0.01); among PCOS subgroups, the eight repeat genotypes in NIR group is more frequent than that in HI group (P < 0.01), and 7/9 genotype distribution in AGM group is higher than that in NIR group and HI group(P < 0.05-0.01). The serum SHBG levels in homozygous genotype groups exhibit a sequence of 8/8 > 9/9 > 6/6, 7/7 repeats and the fall of serum SHBG trend is in reversed relation with the increase in body mass index (BMI), Homa-IR, and blood pressure. Serum SHBG levels in AGM exhibit a sequence of HI group < NIR group < control but show no statistical difference between both groups.</p><p><b>CONCLUSION</b>This study reveals that the repeat number, alleles, genotypes and their distributions in Chinese women are very different from these in foreigners. Some special genotypes and low serum SHBG levels may be associated with PCOS and its glucose metabolic status; some special genotypes may influence Chinese serum SHBG and need more studies, but both SHBG gene polymorphism genotype and serum SHBG are not good indicators to find out the PCOS individual at high risk.</p>

Influence of chemotherapy on Th1/Th2 cytokine switching in stomach cancer patients / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To observe the influence of chemotherapy on the switching of Th1/Th2 cytokines in stomach cancer patients.</p><p><b>METHODS</b>Th1/Th2 cytokine genes expressed by peripheral blood mononuclear cells of stomach cancer patients before and after chemotherapy were detected by RT-PCR.</p><p><b>RESULTS</b>The expression of Th2 cytokines was dominant in patients before chemotherapy, and the dominancy became less marked after chemotherapy.</p><p><b>CONCLUSION</b>The immune deviation with Th2 predominance in stomach cancer patients has a tendency to become reversed after chemotherapy.</p>