乳腺癌细胞株中MKK4 的表达及其对细胞转移的影响和机理研究

目的:观察乳腺癌细胞株中MKK4蛋白的表达水平,研究MKK4蛋白表达对乳腺癌细胞运动能力及EMT标志物的影响,确定MKK4在肿瘤细胞EMT转化及肿瘤转移中的作用,为肿瘤转移机制研究提供一定的基础资料,为肿瘤防治奠定一定的理论基础。方法:通过体外细胞培养技术收集系列乳腺癌细胞株的培养裂解液,利用Western blot技术检测细胞培养裂解液中MKK4及EMT标志物的表达水平,构建MKK4表达水平与细胞转移能力的对应图;采用siRNA技术,干扰MKK4高表达乳腺癌细胞株MKK4的表达,Western blot技术观察MKK4低表达后,EMT标志物的变化,同时,构建MKK4质粒,转染MKK4低表达乳腺癌细胞株,Western blot技术观察MKK4高表达后,EMT标志物的变化。并采用MTT法、Transwell、划痕法观察MKK4高表达后细胞增殖、运动、迁移等能力的变化。结果:乳腺癌细胞株中MKK4蛋白的表达水平与乳腺癌细胞运动能力有一定的相关性,并与EMT标志物的表达具有相关性,MKK4蛋白表达越高,细胞运动能力越差。干扰或转染技术影响乳腺癌细胞株中MKK4的表达后,细胞EMT标志物的表达也相应变化,乳腺癌细胞株中MKK4高表达后,其细胞增殖明显抑制,运动迁移能力也相应下降。结论:乳腺癌细胞中MKK4蛋白的表达水平与乳腺癌细胞EMT转化及运动能力有一定的相关性,MKK4蛋白表达越高,细胞运动能力越差。     

通过染色体整合表达古菌乙酰化酶合成N-末端乙酰化胸腺肽β4

胸腺肽β4（Tβ4）是N-末端乙酰化的43肽,具有多种重要生物学功能.其生物合成存在两大难点,即乙酰化修饰和小分子肽的表达.本研究发现来自古菌Sulfolobus solfataricus的乙酰化酶ssArd1可以催化Tβ4的N-末端乙酰化修饰.利用Red同源重组技术将ssArd1基因表达盒整合至E.coli BL21（DE3）染色体的lpxM位点上,构建了可以实现Tβ4N-末端乙酰化修饰的新型宿主E.coli BDA.将Tβ4编码基因融合在改造的微型Spl DnaX Intein的N端,并在Intein的C端添加His标签,构建了表达载体pET-Tβ4-Intein.在E.coli BDA中表达的融合蛋白,经镍亲和层析纯化后用β-巯基乙醇诱导融合蛋白切割释放小分子多肽,获得了具有N-末端乙酰化的Tβ4.     

QTL mapping for combining ability in different population-based NCII designs: a simulation study

The NCII design (North Carolina mating design II) has been widely applied in studies of combining ability and heterosis. The objective of our research was to estimate how different base populations, sample sizes, testcross numbers and heritability influence QTL analyses of combining ability and heterosis. A series of Monte Carlo simulation experiments with QTL mapping were then conducted for the base population performance, testcross population phenotypic values and the general combining ability (GCA), specific combining ability (SCA) and Hmp (midparental heterosis) datasets. The results indicated that: (i) increasing the number of testers did not necessarily enhance the QTL detection power for GCA, but it was significantly related to the QTL effect. (ii) The QTLs identified in the base population may be different from those from GCA dataset. Similar phenomena can be seen from QTL detected in SCA and Hmp datasets. (iii) The QTL detection power for GCA ranked in the order of DH(RIL) based > F₂ based > BC based NCII design, when the heritability was low. The recombinant inbred lines (RILs) (or DHs) allows more recombination and offers higher mapping resolution than other populations. Further, their testcross progeny can be repeatedly generated and phenotyped. Thus, RIL based (or DH based) NCII design was highly recommend for combining ability QTL analysis. Our results expect to facilitate selecting elite parental lines with high combining ability and for geneticists to research the genetic basis of combining ability.     

Morphology and ontogeny of the eodiscoid trilobite Sinodiscus changyangensis from the lower Cambrian of South China

Abstract: A large number of complete specimens together with numerous disarticulated sclerites of the eodiscinid trilobite Sinodiscus changyangensis Zhang in Zhou et al., 1977 have been collected from the lower Cambrian Shuijingtuo Formation in Changyang, Hubei Province, South China. An ontogenetic series is established based on the immature and mature exoskeletons including the previously unknown protaspides and meraspides, in particular. No further substages can be differentiated in the protaspid specimens herein. Changes that took place during the meraspid period include the addition of postcephalic segments and prominent pygidial larval notches in early meraspid development which became progressively less distinct and disappeared in degree 2. Two holaspid stages are recognized based on the addition of a new pygidial segment, indicating that the start of the holaspid phase preceded the onset of the epimorphic phase and accordingly, its developmental mode is attributed to the protarthrous pattern. The trunk segmentation schedule of S. changyangensis is discussed, which is similar to other primitive eodiscoid trilobites, that is, as the boundary between the thorax and pygidium migrated posteriorly, there is no change in the number of the trunk segments. The processes of liberation of the thoracic segment and segment insertion into the pygidium are separated from one another, and the two different mechanisms, somitogenesis and tagmosis, progress independently during the ontogenetic development of the postcephalic region of these primitive eodiscinids.     

急性铅应激诱导肝肾损伤及其分子机制初探

利用腹腔注射醋酸铅方法构建了铅染毒小鼠(Mus muscculus)模型,观察了染毒小鼠肝、肾的组织学变化,并通过免疫组织化学方法检测了染毒小鼠肝、肾组织中Caspase-3、Bcl-2和Bax蛋白的表达量.结果发现,急性铅染毒可诱导肝和肾组织学损伤,且在诱导肝细胞和肾细胞凋亡、损伤过程中,随时间的延长,Caspase-3的表达量逐渐增加,而Bcl-2与Bax两蛋白表达量的比值呈逐渐下降趋势,有一定的时效性,染毒48 h后,与对照组相比,均差异极显著,表明铅可能通过影响Caspase-3、Bcl-2和Bax的表达而诱导肝和肾细胞异常凋亡.     

自然环境中T-2毒素降解菌的筛选与鉴定

[目的]在自然环境中筛选和鉴定能够降解T-2毒素的菌株.[方法]取对虾养殖池水样、养殖池沉泥样品和对虾混合饲料中分离到的镰孢菌,于GYM产毒培养基中培养14 d后引入自然条件下气载细菌,继续培养至28 d,采用LC-MS/MS检测其中T-2毒素含量.然后结合稀释涂布、平板划线、革兰氏染色、镜检,从镰孢菌产毒培养液中毒素含量明显降低的菌悬液中筛选出T-2毒素降解菌,用16S rRNA分析方法对其进行系统发育分析及菌种鉴定,并验证两菌株的降毒能力与不同基质中二者的联合降毒能力.[结果]以从对虾养殖环境中分离到的5株镰孢菌作为试验菌种,在其产毒培养过程中分离到2株T-2毒素降解菌,16S rRNA鉴定结果分别为弯曲假单胞菌和尼泊尔葡萄球菌,对T-2毒素的降解率分别为90.9％和85.5％,但两者降毒能力并无显著差异(P＞0.05);其联合作用也有较好降毒效果,与两菌株单独作用无显著差异(P＞0.05),不同基质对其联合降毒作用影响不大(P＞0.05).[结论]新的T-2毒素降解菌的发现为进一步探明T-2毒素降解基因和开发T-2毒素生物降解酶提供了研究基础.     

多年生黑麦草温度胁迫差异表达转录因子的比较转录组分析

多年生黑麦草是禾本科冷季型草种,温度胁迫严重影响其分布和产量。转录因子调控基因表达在植物响应逆境胁迫中发挥重要作用。该研究以多年生黑麦草品种雅晴为材料,采用高通量RNA-seq技术,对热(40℃)、冷冻(-10℃)和对照(22℃)处理的样本进行转录因子应答分析,以探索多年生黑麦草转录因子对温度胁迫的响应规律,并筛选抗逆转录因子候选基因。结果显示:(1)共鉴定了694个转录因子unigenes,分属于AP2/ERF、GTF、HSF、MYB、NAC、WRKY、bHLH和bZIP等32个家族。(2)在热和冷冻胁迫下,ERF(AP2/ERF)、MYB、NAC和bZIP家族基因均以上调为主,而WRKY家族则多数下调。HSF、GTF和DREB(AP2/ERF)家族成员大多数受热诱导上调,而多数bHLH家族成员受冷冻胁迫上调。(3)功能富集结果表明,多年生黑麦草差异表达的转录因子基因主要参与植物激素信号转导、昼夜节律和病原体互作等通路。研究表明,多年生黑麦草中与胁迫适应相关的转录因子基因普遍上调表达,而与生长和抗病相关的基因多数下调。该研究筛选到大量抗逆转录因子候选基因,为分子育种提供抗逆基因资源。     

氨基酸定点突变提高灵芝蛋白LZ-8热稳定性的研究

通过氨基酸定点突变技术提高灵芝免疫调节蛋白LZ-8的热稳定性。通过分子动力学模拟结合温度因子预测对LZ-8氨基酸突变位点进行理性设计,在毕赤酵母X33菌株内构建并表达LZ-8突变体蛋白,采用HeLa细胞生长抑制实验和差示量热扫描分析检测并比较了LZ-8突变前后生物学活性及热力学参数。结果显示,LZ-8 N-端α螺旋为理论预测的温度敏感区域,在该区域进行F8W和R9K氨基酸双位点突变,突变后的LZ-8热稳定性提高,相变温度Tm上升0.92℃,相转变焓值ΔH提高23.14 kJ/mol,但突变后LZ-8生物学活性基本不变,LZ-8和LZ-8突变体对HeLa细胞生长抑制的IC50值分别是2.238μg/mL和2.407μg/mL。通过理性设计氨基酸突变位点,获得了稳定性提高但生物学活性不变的灵芝免疫调节蛋白LZ-8的突变体。     

干旱与盐胁迫对花生根际土壤细菌群落结构和花生产量的影响

以花生品种花育25号为试验材料,采用盆栽试验研究了开花期干旱和盐分胁迫对花生生长发育和荚果产量的影响,并运用高通量测序技术分析干旱、盐胁迫及旱盐双重胁迫下,花生根际土壤细菌群落结构的变化特征。结果表明: 不同胁迫处理花生根际土壤细菌群落均以变形菌门、放线菌门、Saccharibacteria、绿弯菌门、蓝藻菌门和酸杆菌门为主。干旱、盐胁迫及旱盐双重胁迫均不同程度降低了变形菌门和放线菌门的相对丰度,但显著提高了蓝藻菌门的含量,且旱盐双重胁迫较其单一胁迫引起的根际蓝藻菌门丰度变化更显著。土壤细菌功能预测分析显示,信号转导机制、防御机制及翻译后修饰、蛋白质周转和分子伴侣等相关功能在旱盐双重胁迫的细菌中活性更强,可能对花生生长及胁迫应答具有重要影响。统计学分析显示,开花期干旱、盐胁迫和旱盐双重胁迫严重影响花生生长发育,并显著降低产量。研究结果可为通过改良土壤微生物环境来提高植物胁迫耐受性提供参考。     

4 630例绝经后妇女阴道微生态状况分析

目的分析绝经后妇女阴道微生态状况,为临床诊疗提供实验室依据。方法以2018年1月至2019年9月在我院妇科门诊就诊的4 630例绝经后妇女作为研究对象,对其阴道微生态检查结果进行回顾性分析。结果 4 630例患者中阴道微生态正常者409例(8.83%),阴道微生态异常者4 221例(91.17%)。阴道微生态异常者中不能通过镜检明确病原菌者1 758例(37.97%),包含优势菌异常1 406例(30.37%)、菌群抑制351例(7.58%)、菌群增殖过度1例(0.02%);通过镜检明确病原菌者2 463例(53.20%),包含细菌性阴道病(BV)1 237例(26.72%)、需氧菌性阴道炎(AV)1 001例(21.62%)、外阴阴道假丝酵母菌病(VVC)159例(3.43%)、滴虫性阴道炎(TV)66例(1.43%)。不同年龄段绝经后妇女阴道微生态构成存在差异,其阴道清洁度(Ⅲ/Ⅳ)、菌群密集度(未见/Ⅰ)、菌群密集度(Ⅳ)、菌群多样性(未见/Ⅰ)、菌群多样性(Ⅳ)、优势菌(无)、优势菌(G~+大杆菌)、优势菌(G~-弧形菌)、优势菌(G~+球菌)、VVC、TV、β-氨基半乳糖苷酶(+)、唾液酸苷酶(+)、过氧化氢(+)、阴道pH4.5、微生态异常发生率在不同年龄段之间的分布差异均有统计学意义(均P0.05)。结论阴道微生态异常在妇科门诊绝经后妇女中发生率较高,且不同年龄段妇女阴道微生态异常构成存在差异,提示应结合年龄治疗阴道疾病,同时积极恢复阴道微生态环境。