CC to TT mutation in the mitochondrial DNA of normal skin: relationship to ultraviolet light exposure

We have previously reported that ultraviolet (UV)-specific (CC to TT) mutations in p53 gene can be detected in normal skin. This, however, cannot be used as a cumulative marker of UV exposure, since cells with the p53 mutation acquire a clonal growth advantage. Moreover, a large skin biopsy is necessary for each assay. In order to circumvent these problems, we have measured mitochondrial (Mt) DNA mutations; there are more than 1000 copies of the Mt genome per cell, and Mt genes are not directly involved in cell growth. We have established a sensitive allele-specific polymerase chain reaction (AS-PCR) assay capable of detecting one CC to TT mutation in Mt DNA among 10(7) wild-type genes using a mismatch allele-specific primer. With this assay, we found no mutation-positive samples from internal non-exposed tissue (stomach, colon, and blood) (0/50). In contrast, 17 out of 111 skin samples were positive: the mutation frequency in positive samples was around 10(7)-10(-6) (10-100 copies of mutant in 10(8) wild-type Mt DNA). In normal skin tissue, the prevalence of positive samples was higher in those from exposed sites (13/51) than in those from less-exposed sites (1/26) (p<0.05). However, a quantitative correlation between sunlight exposure and the accumulation of mutations was not found. We conclude that the UV exposure-associated CC to TT mutation in Mt DNA can be detected in normal skin, but that further studies are required to develop this as a quantitative marker for UV exposure.     

体外研究UGT2B7基因SNP rs28365062对霉酚酸酯代谢的影响

目的:体外研究尿苷二磷酸葡萄糖醛酸转移酶(UGT2B7)基因SNP rs28365062对霉酚酸酯(MMF)代谢的影响,明确该位点变异是否与MMF副作用具有潜在关系。方法:采用基因重组、定点突变技术构建UGT2B7基因SNP rs28365062不同等位基因过表达载体,POLO3000转染法将重组过表达载体转染HEK293细胞,采用液相色谱-质谱联用仪(LC/MS/MS)系统检测霉酚酸(MPA)代谢产物--酰基化葡萄糖醛酸化物(Ac MPAG)24小时的生成量来评估转染不同等位基因细胞的酶活性。结果:成功构建过表达载体p IRES2-EGFP-prom(A)和p IRES2-EGFP-prom(G)并转染至HEK293细胞。LC/MS/MS系统检测结果显示携带G等位基因的突变型UGT2B7代谢MMF产生Ac MPAG的能力降低,产物Ac MPAG 24小时的生成量仅为野生型的18.60%(P0.001)。结论:UGT2B7基因SNP rs28365062可显著影响MMF代谢产物Ac MPAG的生成,可能是导致病人对MMF产生不同程度副作用的因素之一。     

黄氏枝膜叶蜂生物学特性及金龟子绿僵菌对其幼虫的致病力

【目的】黄氏枝膜叶蜂Cladiucha huangbki是新发现的叶蜂科(Tenthredinidae)枝膜叶蜂属Cladiucha种类,以幼虫取食危害乳源木莲Manglietia yuyuanensis叶片,虫口密度大, 严重影响乳源木莲的生长。本研究旨在明确该虫的生物学特性,并筛选出对该虫具有高致病力的金龟子绿僵菌Metarhizium anisopliae菌株,为高效治理该虫提供理论基础和生物防治资源。【方法】以林间调查和室内饲养观察相结合的方式,观察黄氏枝膜叶蜂的各发育阶段形态特征和生物学特性(寄主植物、分布和生活史);采用孢子悬浮液喷雾法筛选高致病力金龟子绿僵菌菌株。【结果】林间和室内观察结果表明,黄氏枝膜叶蜂幼虫取食木兰科木莲属植物,在福建永安市一年发生1代,以老熟幼虫(预蛹)在土室中越夏、越冬;翌年3月上旬化蛹,3月下旬成虫羽化,4月上旬为雌虫产卵高峰期,4月中旬幼虫出现,4月下旬至5月上旬为幼虫危害高峰期,5月中旬老熟幼虫开始入土做土室,入土深度30~150 mm。室内金龟子绿僵菌不同菌株对黄氏枝膜叶蜂幼虫致病力测定结果表明,以1×10⁷孢子/mL悬浮液喷雾11 d后,不同菌株对其幼虫的校正死亡率均超过85%,其中MaQZ-01, MaYTTR-04-05, Ma3和MaFZ-13 4个菌株感染的叶蜂幼虫校正死亡率为100%。菌株MaFZ-08感染引起的黄氏枝膜叶蜂僵虫率最高,为60%;其次是MaYTTR-04-05,引起的僵虫率为50%。金龟子绿僵菌各菌株对黄氏枝膜叶蜂幼虫的致死中时间(LT₅₀)相差不大,在6 d左右,其中MaYTTR-04-05菌株的LT₅₀最短,为5.090 d。【结论】黄氏枝膜叶蜂在福建永安一年发生1代,5月中、下旬老熟幼虫入土做土室化为预蛹越夏越冬。本研究筛选出对黄氏枝膜叶蜂幼虫致病力强的金龟子绿僵菌MaYTTR-04-05菌株,在其防治上有良好的应用前景。     

绿色糖单孢菌木质素过氧化物酶的分离纯化及鉴定

木质素过氧化物酶是一种重要的具有工业应用前景的木质素降解酶,但已报道真菌来源的木质素过氧化物酶只能在酸性低温条件下发挥作用,限制了其进一步的工业应用.通过培养一株耐热耐碱放线茵——绿色糖单孢茵发酵产酶,采用DEAE-Cellulose,CM-Cellulose和Superdex 75凝胶过滤层析等分离纯化方法,得到一种具有耐热耐碱特性的木质素过氧化物酶.经凝胶电泳检测其为单一蛋白,分子量为41 kD.最终纯化倍数达到20倍,活性回收率为6％.采用LTQ法对纯酶进行蛋白质归类鉴定,得到其部分氨基酸片段,为该酶的进一步分子生物学研究奠定基础.     

A two-stage meta-analysis identifies several new loci for Parkinson's disease

A previous genome-wide association (GWA) meta-analysis of 12,386 PD cases and 21,026 controls conducted by the International Parkinson''s Disease Genomics Consortium (IPDGC) discovered or confirmed 11 Parkinson''s disease (PD) loci. This first analysis of the two-stage IPDGC study focused on the set of loci that passed genome-wide significance in the first stage GWA scan. However, the second stage genotyping array, the ImmunoChip, included a larger set of 1,920 SNPs selected on the basis of the GWA analysis. Here, we analyzed this set of 1,920 SNPs, and we identified five additional PD risk loci (combined p<5×10⁻¹⁰, PARK16/1q32, STX1B/16p11, FGF20/8p22, STBD1/4q21, and GPNMB/7p15). Two of these five loci have been suggested by previous association studies (PARK16/1q32, FGF20/8p22), and this study provides further support for these findings. Using a dataset of post-mortem brain samples assayed for gene expression (n = 399) and methylation (n = 292), we identified methylation and expression changes associated with PD risk variants in PARK16/1q32, GPNMB/7p15, and STX1B/16p11 loci, hence suggesting potential molecular mechanisms and candidate genes at these risk loci.     

Martentoxin对TNF-α引起脐静脉内皮细胞释放一氧化氮的影响

目的：研究东亚钳蝎毒素Martentoxin对脐静脉内皮细胞释放NO的影响。方法：人脐带经胶原酶消化分离后获得脐静脉内皮细胞,经抗血管性血友病8因子鉴定后,用TNF-α建立NO释放模型,观察Martentoxin对脐静脉内皮细胞释放NO的影响。结果：①经6,12,24h作用后,10μM、100μM Martentoxin能降低TNF-α引起的NO释放增加（P〈0．05）。②Martentoxin降低TNF-α引起的iNOS活性增强并对TNF—α引起的内皮源性一氧化氮合酶mRNA下调具有一定保护作用（P〈0．05）。结论：Martentoxin抑制TNF-α引起的脐静脉内皮细胞释放NO增加。     

红霉素对慢性阻塞性肺疾病大鼠ICAM-1、MMP-9的表达影响

目的:观察慢性阻塞性肺疾病(COPD)大鼠肺组织中ICAM-1及MMP-9的表达及红霉素的干预作用。方法:复制COPD大鼠模型,并用红霉素干预,收集支气管肺泡灌洗液行细胞学计数和分类检查;采用HE染色观察病理形态变化;免疫组化法检测大鼠支气管肺组织ICAM-1、MMP-9的表达。结果:与模型组比较,干预组支气管肺组织中ICAM-1、MMP-9表达显著降低;模型组中ICAM-1、MMP-9的表达与BALF中白细胞总数及中性粒细胞数成正相关;ICAM-1与MMP-9的表达成正相关。结论:COPD大鼠肺组织中的ICAM-1、MMP-9表达明显升高,可能与COPD的发病机制有关;红霉素可降低ICAM-1、MMP-9的表达,可能是红霉素在COPD中抗炎症反应的作用机制之一。