流式细胞术用于药物抗真菌效应的检测

目的尝试建立用流式细胞术检测药物抗真菌效应的新方法。方法以碘化丙锭为荧光染料,用流式细胞术检测标准菌株近平滑念珠菌ATCC22019在不同浓度氟康唑（FCZ）作用下各管平均荧光强度,探讨最佳实验条件。结果真菌MFI值随着FCZ浓度的增加而增强,其变化具有浓度依赖性,较好地反映了药物的抗真菌效应。结论成功建立用流式细胞术检测药物抗真菌效应的新方法,流式细胞术在该领域的应用值得进一步研究。     

贫血伴白细胞和血小板减少患者CD59检测结果分析

CD59是一种发现于体内几乎所有细胞的膜抗原，可与C8及C9结合阻止膜攻击性复合体（MAC）形成，抑制补体造成的细胞破坏。阵发性睡眠性血红蛋白尿症（PNH）是由于位于X染色体上的PIG-A基因突变，使其编码的、在促进糖化磷脂酰肌醇锚（GPI）合成中第一步所需要的酶发生异常，进而导致锚蛋白家族类（如CD59、CD55等）的表达异常，因此，红细胞和粒细胞表面CD59减少或缺如，     

心衰患者T细胞CD25水平及卡维地洛的体外影响

目的:观察慢性心衰患者外周血T细胞CD25表达水平,探讨卡维地洛(Car)体外对外周血T细胞CD25表达的作用。方法:分离外周血单个核细胞,用卟啉醇肉豆蔻酸乙脂(PMA)+A23187作为刺激剂,加或不加Car,分组体外培养。以流式细胞仪检测T淋巴细胞活化抗原CD25/CD3。结果:(1)与正常人相比,慢性心衰患者T细胞CD25 表达水平明显升高(P<0.001);(2)与对照组相比,10μMCar可以抑制T淋巴细胞CD25抗原的表达(P<0.05)。结论:慢性心衰患者外周血T细胞CD25表达水平明显升高;Car体外可以抑制其表达。     

碘过量和多聚肌苷酸-聚胞苷酸及甲状腺球蛋白诱发小鼠甲状腺炎对Toll样受体3表达的影响

Objective To observe the effect of iodine excess(HI),polyinosinic-polycytidylic acid[Poly(I:C),Poly]and thyroglobulin(TG)on the thyroid of mice by the expression of Toll-like receptor 3(TLR3)to reveal the functional role of TLR3 in autoimmune thyroiditis.Methods Forty-two non-obese diabetic mice,body weight (20±3)g,were divided into six groups:control group,HI group,Poly group,TG group,HI+TG group,HI+Poly group. Fed with deionized water and injected intraperitoneally with physiological saline 0.1 ml each day for a week, the mice in control group were injected with physiological saline every other day at the same dose for 1 week before they were sacrificed; HI group drank 0.05% NaI water and were injected intraperitoneally with physiological saline same as control group; Poly group drank deionized water and were injected intraperitoneally with poly 0.1 ml (1 g/L)each day of the week, then the mice were injected with Poly every other day at the same dose for 1 week before they were sacrificed; TG group drank deionized water and were injected intraperitoneally with physiological saline same as control group, immunized with 0.1 mg TG by subcutaneously injecting and the immunization was enhanced after they were fed half dose for 4 and 8 weeks separately. In HI + Poly group, the treatment was the same as HI group and Poly group; HI + TG group: the treatment was the same as HI group and TG group. Eight weeks later, mice were sacrificed and thyroids were taken to make frozen sections, Hematoxylin-Eosin (HE) staining was employed to observe the morphological change of the thyroids. The expression of TLR3 of thyroids was observed under fluorescence microscope after Immumofluorescence using TLR3 antibody and TR3-positive cells were analyzed in the thyroid density. Results HE staining showed thyroids of Poly group had no inflammation under microscope.There were different degrees of inflammatory cell infiltration in HI group and TG group. The inflammatory cell infiltration and the damage of follicular thyroid of HI + TG group and HI + Poly group were serious, and the degrees of inflammation were higher over "++". Thyroid follicular epithelial cell with TLR3 expression could be seen in Poly group and HI group, meanwhile, there were TLR3 strong positive inflammatory cells in HI group under fluorescent microscope. Using stereological analysis of TLR3-positive cell density in the thyroid, the difference between groups was statistically significant(F=7.870, P<0.01 ). TLR3-positive cell density in the thyroid of HI + Poly group was higher[ (9.287 ± 0.522)mm2] than control group[ (0.062 ± 0.025)mm2, P < 0.01] significantly, meanwhile, the density in HI + Poly group was higher than HI group [ (2.574 ± 0.257 )mm2] and Poly group[ (1.361 ± 0.148 )mm2, all P < 0.01]. The density in HI + TG group[ (4.843±0.405)mm2] was higher than HI group and TG group[(1.601 ±0.268)mm2, all P < 0.01 )]. Conclusions Excessive iodine and thyroglobulin can induce thyroiditis, and stimulate the expression of TLR3 in the thyroid follicular epithelial, Poly aggravated thyroiditis induced by iodine excess in NOD mice; TLR3 positive inflammatory cells also appeared in inflammatory region, suggesting that TLR3 is involved in the pathogenesis of autoimmune thyroiditis     

碘诱发NOD和Balb/c小鼠甲状腺炎的实验研究

Objective To observe the different effects of iodine excess on inducing two strain mice thyroiditis. Methods NOD and Balb/c mice, each having 14 mice, were divided into NaI and control group. The mice were given 0.05% NaI water for 8 weeks in NaI group. RIA and ELISA were used respectively to detect TT4, TgAb, TPOAb and TSH level in serum. Morphology changes of thyroid and apoptosis of thyrocytes stained by immunohistochemistry were observed under light microscope. Lymphocytic proliferation of cervical lymph node and spleen to responding to Tg were detected by MTr method. Results After intake of iodine water for 8 weeks, NOD and Balb/c mice showed relative quality of thyroid in Nal group[(104.83±14.52), (155.79±20.77)mg/kg]obviously increased compared with control group[(71.80±20.42), (105.15±21.98)mg/kg, t values:-3.293,-4.429, all P< 0.01)], enlarged follicular lumen with colloid accumulation were observed in thyroid. Serum level of TT4 in Nal group [(29.52±4.42), (19.53± 2.35)nmol/L]to control group[33.40±5.38), (23.47±6.22)nmol/L]of NOD and Balb/c mice showed a decreasing tendency(t values: 1.374,1.567, all P > 0.05). TSH of Nal group showed an increasing tendency in Balb/c mice[(4.14±1.71)μg/L, compared with control [(3.55±1.41)μg/L, t values:-0.705, P > 0.05]and obviously increased in NOD mice [(6.98±0.66)μg/L, compared with control[(555±056)μg/L, t values:-3.562, P< 0.01], but no change of TgAb and TPOAb level in Nal group(1281,1364 cpm, 2.50×103, 0.14×103U/L were observed, compared with control(1297,1220 cpm, 3.17×103,0.03×103 U/L; Zvalues:-0.081,-0.703, -0.244,-1.293, all P > 0.05). In NOD mice NaI group, apoptosis of thyrocytes was more intense than Balb/c mice, obvious infiltration of lymphoeytes, disorganization and focus fibrosis was seen in thyroid. The cell amount of NaI group increased in NOD mice lymph node and spleen cells[(1.100±0.014), (1.076±0.033)]were more than that in the control group [(0.993±0.011), (1.005±0.003), t value:-11.672,-4.314, P < 0.01). Conclusions Iodine leads to enlargement of thyroid and malfunction of thyroid in Balb/c mice. Besides, NOD mice have generate inflammatory reaction in thyroid and produced sensitized lymphocytes to Tg. Iodine excess can induce NOD mice to occur autoimmune thyroiditis.     

瞬时转染和稳定转染对RNAi抑制乙型肝炎病毒S基因表达的影响

目的:构建针对乙型肝炎病毒(HBV)S基因的siRNA表达载体Psuper-S1和Psuper-S2,观察瞬时转染及稳定转染对RNAi抑制HBV基因表达作用的影响.方法:设计并合成针对HBV S基因的siRNA寡核苷酸,经退火形成双链后克隆入Psuper载体,构建成功的siRNA表达载体分别瞬时转染和稳定转染表达HBV的2.2.15细胞,对所得细胞上清中的HBsAg和HBeAg进行定量检测,RT-PCR检测siRNA对靶基因Mrna的抑制效果,并比较两种转染方式对RNAi抑制HBV基因表达作用的影响.结果:在稳定转染并经潮霉素筛选所得的单克隆细胞株中,Psuper-S1和Psuper-S2均能明显抑制HBsAg及HBeAg的分泌(P<0.01),抑制率分别为83%和78%,RT-PCR结果证实HBV的Mrna明显降低,而瞬时转染细胞在蛋白质水平上及Mrna水平上的结果则比稳定转染的结果逊色.结论:针对HBV S基因的siRNA能稳定、高效、特异地抑制HBV基因的表达,转染效率对于靶基因的RNA干涉作用有明显的影响.     

针对尿激酶型纤溶酶原激活剂的siRNA的载体构建与表达

目的:构建针对尿激酶型纤溶酶原激活剂(uPA)mRNA的siRNA的表达载体,并观察其对转染细胞中的uPA表达的抑制作用. 方法:化学合成用于产生针对uPA的发卡状RNA的寡核苷酸,各64个碱基,退火后插入线性化的pSUPER载体的H1启动子下游. 重组载体经限制性酶切和测序. 把构建的载体转染乳腺癌细胞系MDA-MB 231,观察其对uPA的干扰作用. 结果:限制性酶切和序列测定表明成功构建了可以表达针对uPA的发卡状RNA表达载体,该载体表达的发卡状RNA能够有效的下调uPA的表达. 结论:成功的构建了针对uPA的发卡状RNA表达载体,为进一步研究uPA的功能和肿瘤的基因治疗打下基础.     

高血压脑出血患者脑脊液中游离 DNA 动态检测的意义

目的：探讨高血压脑出血患者脑脊液中游离DNA的动态变化与组织损伤的关系。方法54例脑出血患者按照格拉斯哥昏迷分级（GCS）评分分为轻度17例,中度21例和重度16例,分别采集发病后1、3、7、15 d 脑脊液2 mL,提取游离 DNA,采用荧光定量 PCR 技术检测游离 DNA 水平。结果脑出血组中游离 DNA 水平重度组显著高于轻度组和中度组;颅内压异常组高于正常组;颅内感染组高于非感染组。结论脑脊液游离 DNA 水平对于诊断脑出血颅脑损伤有一定诊断价值,而且有助于监控术后并发症的发生。     

腹腔镜经皮穿刺腹膜外腹股沟斜疝 结扎术的不良因素与处理

目的 总结并分析腹腔镜经皮穿刺腹膜外结扎术（LPEC）治疗小儿腹股沟斜疝时可能遭遇的各类不良因素,探讨适宜的处理方法和原则。方法 回顾分析该院2011年3月-2013年3月218例LPEC时遭遇到的各类不良因素、手术过程、处理方法及术后。结果 该组病例中82例出现不同程度的输精管辨认困难,其中4例最终无法辨认;69例遭受到腹膜褶皱的困扰;32例大龄或内环口开放巨大的病例;5例因为膀胱充盈,2例乙状结肠充盈;2例穿刺导致局部腹膜出现严重破损;2例大网膜疝入并与疝囊粘连;4例腹膜外气肿。术中采用各种方法灵活处理,最终212例完成LPEC手术,另外6例中转为传统小切口手术。该组无1例因术后复发及其他明显并发症出现。结论 绝大数小儿腹股沟斜疝可以通过LPEC治愈,但可能遭遇多种不良因素,灵活处理并遵守一定原则可以预防、降低或解除不良因素影响。     

出入境人群HIV感染者/AIDS患者血清生化指标分析

目的了解HIV感染后血清生化指标的变化,探讨HIV感染对机体组织的损伤程度。方法病例组为广西出入境人群发现的HIV感染者/AIDS患者,对照组为随机抽取的经广西口岸出入境正常人群。用全自动生化分析仪检测血清生化指标。结果各实验组ALT、ALP、BUN、Cre在正常范围;HBV/HCV/HIV混合感染组的AST明显高于HIV单感染组(P<0.01),而HIV单感染组与正常对照没有明显差异(P>0.05);各实验组γ-GT除吸毒传播感染组与正常对照没有明显差异(P>0.05)以外,其余各组都显著高于对照组(P<0.01);各实验组ALB除吸毒传播感染组与对照组没有明显差异(P>0.05)以外,其余各组都显著低于对照组(P<0.05)。结论单纯HIV感染对肝、肾组织似乎没有造成严重影响,HIV感染/AIDS者的白蛋白降低可能与慢性腹泻等消耗性疾病有关。