细菌预成酶联合检测在阴道微生态评价中的应用价值

目的．探讨需氧菌性阴道炎（AV）、细菌性阴道病（BV）五项联合检测在阴道微生态评价中的应用价值。方法用传统方法对5000例健康体检者阴道分泌物进行清洁度、阴道pH、革兰染色检查作阴道菌群分析；同时应用AV、BV五项联合检测进行细菌预成酶测定。结果5000例体检者中传统方法检测阴道菌群正常2675例（53．5％），阴道菌群异常2325例（46．5％）；阴道菌群异常组细菌预成酶检测结果阳性率明显高于阴道菌群正常组，差异有统计学意义（P〈0．05）。结论AV、BV五项联合检测灵敏度高，操作简便，可以作为快速筛查方法应用于临床，结合阴道清洁度等指标可系统评价阴道微生态状况。

阴道分泌物常规和预成酶谱五项联合检测分析

目的探讨女性阴道分泌物常规（形态学）和预成酶谱五项联合检测分析对阴道炎诊断的临床价值。方法采集该院妇科门诊患者阴道分泌物进行常规检查（白细胞、上皮细胞、阴道毛滴虫、假丝酵母菌），同时采用阴道分泌物预成酶谱五项联合检测。结果1000例阴道分泌物标本常规检9n,4联合预成酶谱分析检测检出阳性患者620例，阳性率为62％（G20／1000），在阳性标本中以需氧菌阴道炎（AV）最为常见（59.2％），同时AV也是合并细菌性阴道病（BV）和假丝酵母菌阴道炎（VVC）的主要病原菌；本文统计显示H2O2阳性率为73.8％，提示健康妇女中有大部分处于乳酸杆菌减少或缺失，阴道微生态处于亚失调状态。结论对阴道炎患者阴道分泌物同时进行常规和预成酶谱五项联合检测，能更好反映阴道炎患者的病源体感染状况，有利于临床医生对症治疗。

厌氧菌预成酶在大鼠肠道菌群失调诊断应用

目的探讨厌氧菌预成酶作为快速诊断肠道菌群失调的可行性。方法以盐酸林可霉素灌胃实验大鼠造成其肠道菌群失调模型后,同时对肠道菌群及厌氧菌预成酶进行连续、定量分析。结果随着抗生素的灌胃,实验大鼠肠道内的正常菌群在数量上发生较大变化,尤其是优势厌氧菌双歧杆菌及乳酸杆菌的数量在抗生素灌胃的2、4、6d与正常对照组相比差异均有统计学意义(P<0.05);预成酶中除少数酶未发生改变,大多数酶随着抗生素的灌胃数量都在减少甚至消失,且与正常对照组相比差异均有统计学意义(P<0.05)。结论厌氧菌预成酶作为快速诊断肠道菌群失调的方法有可行性,但仍需进一步的实验。

预成酶谱分析和涂片染色多功能检查在妇科感染诊治中的临床应用价值

目的研究基于预成酶谱分析和涂片染色多功能检查的阴道菌群多维度评价方法在妇科感染诊治中的临床应用价值。方法选取到该院做妇科白带检查的21175例患者作为研究对象，同一患者均同时采集白带标本做生化检查和涂片染色检查，生化检查采用新型预成酶谱分析技术检测，涂片染色多项检查采用妇科白带涂片多功能染色液染色镜检。结果预成酶谱生化五项阳性检出率：过氧化氢（H2O2）63．83％、唾液酸苷酶（SNa）7．73％、白细胞酯酶（LE）46．16％、β-葡萄糖醛酸苷酶（GUS）3．19％、凝固酶（GADP）46．16％；涂片染色多项检查的各项检出率：滴虫0．63％、念珠菌11．73％、淋球菌0．29％、纤毛菌2．63％、球菌1．82％、条件致病小杆菌17．00％、加特纳球杆菌24．89％、线索细胞17．18％、癌细胞0．23％、阴道清洁度Ⅲ-Ⅳ20．72％。结论联合妇科白带预成酶谱分析与涂片染色多功能检查可在短时间内检出女性常见下生殖道感染、宫颈炎、宫颈癌等异常情况，为全面诊断和治疗妇科疾病提供可靠依据，在大型妇科普查和癌症筛查方面具有重要的临床价值，尤其值得在基层医院推广应用。

预成酶谱分析技术诊断女性需氧菌阴道炎的临床价值

目的:探讨预成酶谱分析技术在女性需氧菌阴道炎(AV)诊断中的临床价值。方法:采用Donders湿片镜检法和预成酶谱分析技术分别对300例疑似AV患者的阴道分泌物进行检测分析。结果:300例就诊患者,使用Donders湿片镜检法检出AV患者181例,预成酶谱分析技术检出AV患者178例,两种方法检出AV阳性率差异无统计学意义(P>0.05)。以Donders湿片镜检法为诊断标准,预成酶谱分析技术诊断AV的敏感性为94.9%(169/178)、特异性为90.2%(110/119),二者的符合率为93.0%(279/300)。结论:预成酶谱分析技术诊断AV具有很好的敏感度和特异度,与Donders湿片镜检法符合率高,且该方法操作简便、反应快速,值得妇科门诊及社区医院使用。

红杉醇对2型糖尿病大鼠心肌NADPH氧化酶亚单位p22phox、p47phox及iNOS蛋白表达的影响

目的:探讨红杉醇(sequoyitol,Seq)对2型糖尿病大鼠心肌烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶亚单位p22phox、p47phox及诱导型一氧化氮合酶(iNOS)蛋白表达的影响。方法:雄性SD大鼠100只,分为正常组、模型组、Seq(12.5、25、50mg/kg)及阿卡波糖(Aca,20mg/kg)组,采用小剂量链脲佐菌素(STZ,30mg/kg)联合高糖高脂饲料建立2型糖尿病大鼠模型,HE染色观察心肌病理变化,Western Blot法检测心肌组织iNOS、p22phox及p47phox蛋白表达,比色法检测心肌组织一氧化氮(NO)、过氧化氢(H2O2)、超氧化物歧化酶(SOD)及总抗氧化能力(T-AOC)的含量。结果:与对照组相比,模型组大鼠空腹血糖明显升高,心肌肥厚明显加重,心肌组织iNOS、p22phox及p47phox的表达水平及NO、H2O2含量显著升高,而SOD和T-AOC水平明显降低。与模型组相比,Seq各剂量组和Aca组能不同程度地降低大鼠血糖,改善糖尿病大鼠心肌损伤,下调心肌iNOS、p22phox和p47phox的表达,降低心肌NO、H2O2含量,升高SOD、T-AOC水平。结论:Seq可能通过抑制心肌p22phox、p47phox及iNOS所介导的氧化应激损伤,进而对糖尿病心肌损伤起到保护作用。

急性冠脉综合征患者循环血单核细胞产生明胶酶增加

目的 :探讨单核细胞产生两种明胶酶 (MMP2 ,MMP9)与急性冠脉综合征 (ACS)发病间的关系。方法 :分离 38例不稳定心绞痛 (UA)、16例心肌梗死 (AMI) ,2 2例稳定性心绞痛 (SA)及 18例正常对照 (CONT)循环血单核细胞 ,在体外无血清培养 2 4h ,测定上清液中MMP2、MMP9蛋白含量 ,并观察心肌缺血及心肌梗死对其的影响。结果 :UA、AMI组MMP2分别高于CONT组 (2 6 5 8 5 2± 5 33 42和 1842 34± 317 17,P <0 0 0 1,单位为 μg/L ,下同 ;2 6 43 0 4± 6 41 47和 1842 34± 317 17,P <0 0 0 1)和SA组 (2 6 5 8 5 2± 5 33 42和 1919 16± 2 89 38,P <0 0 0 1;2 6 43 0 4± 6 41 47和 1919 16± 2 89 38,P <0 0 0 1)。UA、AMI组MMP9也分别高于CONT组 (481 41± 5 2 5 6和 30 1 2 6± 30 17,P <0 0 0 1;486 34± 37 48和 30 1 2 6± 30 17,P <0 0 0 1)和SA组 (481 41± 5 2 5 6和 2 98 86± 42 2 4,P <0 0 0 1;486 34± 37 48和 2 98 86± 42 2 4,P <0 0 0 1)。心肌缺血发作前后MMP2、MMP9无显著变化 ,心肌梗死 72h内仍保持在发病初期水平 ,梗死后多个时间点及其最大值与CK或Ck MB相应时间点及其最大值间无线性相关。结论 :急性冠脉综合征患者循环血单核细胞产生MMP2、MMP9增加 ,MMP2。

大黄灵仙方调控胆管细胞炎症的作用及机制研究

目的 观察大黄灵仙方调控白细胞介素-1受体相关激酶(interleukin-1 receptor-associated kinase 1,IRAK)-1和IRAK-4以及成纤维生长因子-3激活酶1MAP3K7结合蛋白2(TGF-3 activator 1 map3k7-binding protein 2,TAB2)的表达水平,探讨其缓解胆管细胞炎症的作用机制,为预防肝胆管结石的形成及术后复发提供理论依据。方法 将45只SPF级健康雄性SD大鼠随机分为9组,空白组、模型组、大黄灵仙颗粒组、激活核转录因子κB(nuclear factor kappa-B,NF-κB)阻断剂组、p38丝裂活化蛋白激酶(mitogen-activated protein kinase, MAPK)阻断剂组、NF-κB阻断剂+p38MAPK阻断剂组、NF-κB阻断剂+大黄灵仙颗粒组、p38MAPK阻断剂+大黄灵仙颗粒组、NF-κB阻断剂+p38MAPK阻断剂+大黄灵仙颗粒组,每组大鼠5只。除空白组以外,其余各组大鼠于胆总管处一次性注射5 mg/kg内毒素脂多糖以构建肝内胆管感染动物模型。第7天灌胃结束后取材,取大鼠完整胆管树组织,采用蛋白免疫印迹法和实时荧光定量PCR检测IRAK-1、IRAK-4、TAB2蛋白及mRNA表达量。结果 (1)与空白组比较,模型组IRAK-1、IRAK-4、TAB2蛋白及mRNA表达量升高(P<0.05);(2)与模型组比较,大黄灵仙颗粒组、NF-κB阻断剂组、p38MAPK阻断剂组、NF-κB阻断剂+p38MAPK阻断剂组、NF-κB阻断剂+大黄灵仙颗粒组、IRAK-1、IRAK-4、TAB2的蛋白及mRNA表达量降低(P<0.05),NF-κB阻断剂+p38MAPK阻断剂+大黄灵仙颗粒组IRAK-1、IRAK-4、TAB2蛋白及TAB2 mRNA表达量均显著下降(P<0.05);(3)与大黄灵仙颗粒组比较,NF-κB阻断剂+大黄灵仙颗粒组、NF-κB阻断剂+p38MAPK阻断剂+大黄灵仙颗粒组IRAK-1、IRAK-4、TAB2的蛋白表达量和NF-κB阻断剂+p38MAPK阻断剂组的IRAK-1、IRAK-4、TAB2 mRNA表达量均下降(P<0.05)。结论 大黄灵仙方可缓解内毒素脂多糖诱导的大鼠胆管炎症反应,抑制炎症因子的异常表达,促使胆道恢复至非炎症状态,从而达到预防肝胆管结石的形成及术后复发的目的,其作用机制可能与抑制TLR4/NF-κB/MAPK信号通路下游IRAK-1、IRAK-4和TAB2因子的活化有关。

需氧菌性阴道炎的临床及微生态特征分析

目的:探讨妇科门诊需氧菌性阴道炎(aerobic vaginitis,AV)的临床及微生态特征。方法:回顾性分析4019例妇科门诊不同症状就诊者阴道微生态评价检测指标(菌群密集度、多样性、优势菌、病原体、AV评分、Nugent评分及五项菌群功能及炎症反应指标)及诊断结果。结果:4019例被检查者中AV占3.11%(125/4019)。AV患者中单纯AV占24.00%(30/125),AV混合感染占76.00%(95/125),其中AV合并外阴阴道假丝酵母菌病(VVC)占44.80%(56/125),AV合并滴虫阴道炎(TV)占27.20%(34/125),AV合并细菌性阴道病(BV)占2.40%(3/125),AV合并TV、VVC占0.80%(1/125),AV合并BV、TV占0.80%(1/125)。125例AV患者阴道乳杆菌功能均异常伴阴道炎症反应,其中优势菌群异常占84.80%(106/125)。结论:AV为常见阴道感染,易合并其他感染,尤以合并VVC及TV多见。微生态特征为需氧菌增多,乳杆菌功能异常或优势菌群被替代伴明显阴道炎症反应。因此,应用阴道微生态评价系统诊断AV时,注意微生态状况及有无合并其他类型阴道感染。

Developmental Changes of the FAS and HSL mRNA Expression and Their Effects on the Content of Intramuscular Fat in Kazak and Xinjiang Sheep

Twenty-four male Kazak sheep and 30 Xinjiang fine wool sheep at different ages were selected to investigate the development-dependent expression levels of fatty acid synthase （FAS） gene and hormone-sensitive lipase （HSL） gene in muscle and their effects on the contents of intramuscular fat （IMF）. Longissimus dorsal muscle was sampled to measure IMF and total RNA was extracted to determine FAS and HSL mRNA expression levels by real-time PCR. The results showed that： l） The IMF content increased continuously with growth and showed significant differences （P 〈 0.05） between different age groups in male Kazak sheep, but in Xinjiang fine wool sheep there was no such difference observed. Furthermore, the IMF contents in Kazak were much higher （P 〈 0.01） than that of the other breed from day 30 to 90. 2） FAS mRNA expression level was the highest （P 〈 0.05） on day 0 in Kazak sheep and then declined with growth, in the other breed the gene showed a d‘ecline-rise-decline-rise＇ expression manner as the animals grew. HSL mRNA expression level had a similar model in two breeds, in Kazak sheep it was the highest on day 0 （P 〈 0.05） and in Xinjiang fine wool sheep on day 30 （P 〈 0.01）, then both decreased after this term. 3） In male Kazak sheep, FAS and HSL mRNA expression level were both negatively related to IMF content （r= -0.485 （P = 0.02）, r= -0.423 （P = 0.05））, and the ratio of FAS/HSL expression exhibited significantly negatively related IMF contents. In male Xinjiang sheep, there were no obvious relationship between FAS and HSL expression and IMF content （P 〉 0.05）.

Isolation of S-adenosylmethionine Synthetase Gene from Suaeda salsa and Its Differential Expression Under NaCl Stress

AdoMet plays numerous roles of being the major methyl-group donor in trans-methylation reactions. To gain insight into the possible functions of the AdoMet protein of Suaeda salsa L. in response to salt stress, S adenosylmethionine synthetase gene (SAMS2) was analyzed. We isolated SAMS2 cDNA clone (AF321001) from a lambda -Zap cDNA library constructed from the halophyte S. salsa Pall aerial tissue treated with 400 mmol/L NaCl. SsSAMS2 was found to encode a S-adenolyl-L-methionine synthetase enzyme (AdoMet synthetase). The fragment was 1 531 bp with an open reading frame of 395 amino acids, the calculated molecular weight was about 43 kD. SsSAMS2 showed the highest homology to SAMS2 gene of Catharanthus roseus G. Don., with 93% identity in deduced amino acid sequence. Southern blotting analysis showed that SsSAMS2 might be a two-copy gene in S. salsa genome. Northern blot indicated that the cDNA was up-regulated by salt and other stresses. Enzyme activity assay indicated that the activity of SAMS2 increased under NaCl stress.

RICE TRANSFORMATION WITH A PHYTOALEXIN GENE AND BIOASSAY OF THE TRANSGENIC PLANTS

Immature embryos, mature embryos and embryogenic calli of 6 rice (Oryza sativa L.) materials were transformed with particle bombardment. The plasmids pSSVstl and pVE5+ were used, both containing the phytoalexin gene from grapevine coding for stilbene synthase, but driven by 35S and its own promoter respectively. Through resistance selection for G418 (100 to 150 mg/L) or hygromycin (50 mg/L), 54 independent transgenic plants were isolated and further assessed by PCR, Southern blot and Dot blot analyses. The transgenic plants and their progenies were tested for resistance to blast ( Pyricularia oryzae ) and bacterial blight of rice ( Xanthomonas oryzae ). Preliminary results indicated that the stilbene synthase gene could enhance the resistance of transgenic plants and their progenies to both pathogens.

Comparative Study of Immunological Properties on Glutamine Synthetase Isozymes in Rice Plants

In rice (Oryza sativa L.) roots two glutamine synthetase (GS) isozymes, GSra and GSrb, were identified recently in the author's experiments, but the homology of both GSra and GSrb as well as their localization in the rice roots are unclear. In the present study, the purified GSra and GSrb from rice roots were used to immunize rabbits to obtain the respective antibodies. The immunodiffusion and immunoblotting experiments showed that the antibody against GSra or GSrb was specific for GS and its isozymes. The immunoprecipitation test indicated that the antibody of GSra or GSrb not only recognized its respective antigen, but also well recognized each other's antigen. GSra or GSrb antibody recognized also better cytosolic GS1 of rice leaves, but the recognization for chloroplast GS2 from rice or spinach (Spinacia oleracea Mill.) leaves was weaker. Our results indicate that GSra and GSrb from rice roots are quite similar in antigenicity and are extremely similar proteins and that both GSra and GSrb may also be a form of cytosolic GS just as the cytosolic GS1 of rice leaves.

Effect of Low Temperature Stress on Sweet Potato S-Adenosyl Methionine Synthetase Gene Expression

[Objective] The mRNA expression level changes of S-adenosylmethionine synthetase （SAMS） under low temperature stress was studied. [Method] Total RNA were extracted from leaves, stem and earthnut of sweet potato 0,12,24,48 and 72 h after low temperature treatement, mRNA expression level was analyzed by reverse expression and Real-time PCR technique. [Result] The expression quality of the gene extracted from leaves, stem and earthnut increased and the expression quality reached the peak point 24,72 and 72 h after low temperature treatment respectively. The expression change of earthnut was the biggest. [Conclusion] Low temperature was good for increasing mRNA expression of relevart genes.

Secreted Expression of S-adenosy-L-methionine Synthetase in Pichia pastoris

[Objective] The research aimed to study the secreted expression of S-edenosyl-L-methionine synthetase （SAMS） in Pichia pastoris. Method ] The gene coding SAMS, from the genomic DNA of Saccharomyces cerevisiae, was amplified by PCR and inserted into the secreted expression vector pPIC9K to get recombinant plasmid. The recombinant plasmid pPIC9K-sarr~ was integrated into Pichia pastoris GSl15 genome by electroporation and induced by methanol. The activity of the recombinant enzyme was measured using high-pedormance liquid chroma- tography （HPLC） by determining the production of S-adenosy-L-methionine （SAM） with the enzyme secreted. [ ResultJ The molecular weight of the expression protein identified by SDS-PAGE was about 50 kD, being larger than the theoretical molecular mass of SAMS, which might be due to the glycosytation in the process of secretion. Methanol-induction as well as preliminary purification could enhance the enzyme activity, espe- cially the latter, after which the specific activity of SAMS was improved to 61.48 U/rng. [Conclusion] SAMS with biological activity was secreted successfully in Pichia pastoris GSl15 for the first time. And it is the start for the genetic engineering strains to open up prospects for industrial production.

Effect of ^(12)C^(6+) Ions Beam Irradiation on Seed Germination and Enzymes Activity in Seedlings of Sweet Sorghum

[ Objective] The aim was to study the effect of 12C6 ＋ ions beam irradiation to two varieties of sweet sorghum on seed germination and some enzymes activity in seedlings with different doses, and provided a theoretical foundation for sweet sorghum breeding. [ Method] After germination, the germination potential, germination fraction and enzyme activity were detected, respectively. [ Result] The results showed that with the dose increased, the germination potential of sweet sorghum increased first and then decreased, while their germination fraction presented ＂shoulder like shape＂ ; the activity of LDH, SOD, CAT and GSH-Px increased first and then decreased with doses, they presented slight differences among different enzymes. [ Conclusion] Low dose radiation could accelerate germination of sweet sorghum seeds and enzyme activity could remain at a relatively high level. Enzyme activity decreased with high doses and the growth of sweet sorghum was inhibited.

Bioinformatics Analysis of Chalconesynthase Gene in Morus

[Objective] This study aimed to explore the molecular mechanism of mulberry pigment metabolism regulation. [Method] Chalconesynthase（CHS） gene was cloned from Morus（Moraceae） in silico. The amino acid sequence, physical and chemical properties, transmembrane structural domain, hydrophobicity/hydrophilicity,subcellular localization, secondary and tertiary structure of protein were predicted and analyzed by bioinformatics tools. [Result] The cDNA sequence of CHS gene was 1 365bp by splicing using the software DNAstar and it contained a complete ORF including 1 170 bp which encoded 389 amino acids. Bioinformatic analysis showed that CHS gene included specific peptide sequence RLMMYQQGCFAGGTVLR of chalcone synthase superfamily, but has no signal peptide, belonging to the non-secretory proteins, located inside of cytoplasm. Its molecular evolution is more conservative.[Conclusion] The results above provided foundation for the further studies of structure and function of CHS protein.

Germination Characteristics and Secondary Metabolism Regulation of Scutellaria baicalensis Georgi Seeds under Different Light Intensities

[Objective] This study aimed to investigate the primary and secondary metabolisms during the germination of Scutellaria baicalensis Georgi seeds under different light intensities. [Method] The activities of CHL, soluble sugar, PAL, C4H and CHS were determined with ultraviolet spectrophotometry. The secondary metabolites were detected by High Performance Liquid Chromatography （HPLC）. [Result] The results indicate that the germination of Scutellaria baicalensis Georgi seeds is not sensitive to light and the seedlings were very sensitive to light. The CHL, soluble sugar, PAL, C4H and CHS continuously increased with light intensity. The content of secondary metabolites also increased. [Conclusion] Light increased the formation of leaf photosynthetic pigment, thereby affecting the primary metabolites. The activities of PAL, C4H and CHS significantly increased with the development of light intensity. Finally the secondary metabolites of medicinal plants increased sharply. Therefore, the quality of Scutellaria baicalensis Georgi materials can be improved by increasing the light intensity moderately.

需氧菌阴道炎联合测定试剂盒的临床诊断价值

目的探讨需氧菌阴道炎（AV）联合测定试剂盒（AV法）对AV的诊断效能及临床价值。方法用AV法和AV评分标准法对妇科门诊就诊1150例患者的阴道分泌物进行测定，结果做对比分析。结果以AV评分标准法作为金标准，AV法检测AV的敏感度为90．30％，特异度为97．93％，准确度为97．04％，阳性预测值为85．21％，阴性预测值为98．71％，经z。检验，结果差异无统计学意义（x^2=1．44，P〉0．05），二种方法的检测结果一致性较高（Kappa值为0．86）。结论AV法简便、快速，其结果与AV标准法有很好的一致性，是一种值得推广的临床诊断AV的常规方法。

女性下生殖道感染病原菌检测方法的结果分析

目的探讨白带常规镜检法、细菌预成酶谱分析法及联合法3种方法在鉴别诊断女性常见下生殖道感染疾病中的作用。方法分别采用白带常规镜检法和细菌预成酶谱分析法对来我院就诊的4130例就诊者的阴道分泌物进行检测,检测结果结合临床表现和治疗效果进行汇总分析比较。结果采用白带常规镜检法检出阳性者1976例(检出率47.8%、准确率71.6%);采用细菌预成酶谱分析法检出阳性者1302例(检出率为31.5%、准确率58.1%);采用联合法检出阳性者2622例(检出率63.5%、准确率91.1%)。结论联合法集取白带常规镜检法和细菌预成酶谱分析法的优点,能一次采样即可准确鉴别5种女性常见下生殖道感染疾病。