Local Expression of Vaginal Th1 and Th2 Cytokines in Murine Vaginal Candidiasis under Different Immunity Conditions

To investigate the expression of vaginal Th1 and Th2 cytokines in rats with experimental vaginal candidiasis under different immune conditions, ICR murine vaginal candidiasis model was established and immno-suppressed murine models of vaginal cadidiasis were established in estrogen-treated mice. Non-estrogen-treated mice were used as controls. The mRNA level of Th1 （IL-2）/Th2 （IL-4, IL-10, TGF-β1） cytokines in murine vaginal tissues was determined by RT-PCR. The cykotine in local tissues was increased to different extent under normal immune condition. IL-2 mRNA was increased during early stage of infection, while IL-10 was increased transiently during late stage of infection. TGF-β1 production was found to be increased persistently. At same time, the expression of IL-2 mRNA was suppressed in immno-suppressed group, and the level of IL-4, IL-10, and TGF-β1 were higher than the normal immunity group to different degree during infection. The high level of IL-2 mRNA during early stage of infection was associated with clearance of mucosal Candidia albicans （C. albicans）, and its expression suppressed leading to decreased clearance of mucosal C. albican in immuno-suppression. The over-expression of IL-4 and IL-10 could significantly enhance the susceptibility to C. albicans infection in mice.     

Effects of Propofol on The mRNA Expression of Toll-like Receptor 4 in BV-2 Cells During Mimic Ischemia-Reperfusion Injury In Vitro

This study investigated the effects of propofol on the mRNA expression of Toll-like receptor-4 (TLR4) in BV-2 cells during mimic ischemia-reperfusion (I/R) injury in vitro. BV-2 cells, a mouse microglia line, were cultured and divided into 4 groups at random: control group (group C), ischemia/reperfusion group (group I/R), low-dose propofol (25 μmol/L) intervention group (group PF25) and high-dose propofol (100 μmol/L) intervention group (group PF100). The mRNA expression of TLR4 and NF-κB was measured by means of RT-PCR. TNF-α levels in the supernatants of BV-2 cells were detected by ELISA. The results showed that the mRNA expression of TLR4 and NF-κB was significantly higher in groups I/R, PF25 and PF100 than in group C (P<0.01). And the TNF-α level in the supernatants was elevated in groups I/R, PF25 and PF100 as compared with that in group C (P<0.01). After pre-treatment with propofol, the mRNA expressions of TLR4 and NF-κB and the TNF-α level were significantly decreased in groups PF25 and PF100 in comparison to those in group I/R (P<0.01). And the decrease in those indicators was more significant in group PF100 than in group PF25 (P<0.01). It was concluded that propofol exerted brain-protecting effects during I/R injury by suppressing the mRNA expressions of TLR4 and NF-κB and deceasing the TNF-α level.     

Expression of NF-κB in hRPE Cells Stimulated by PDTC and IL-1β

The constitutive expression of nuclear-factor-κB (NF-κB) in human pigment epithelial (hRPE) cells cultivated in vitro and the possible changes when incubated with PDTC and IL-I were investigated. The synchronized hRPE cells in vitro were divided into two groups. In nonPDTC group, hRPE cells were exposed respectively to IL-1β and NS (for detecting the constitutive expressions of NF-κB in hRPE cells) ; In PDTC group, PDTC-pretreated hRPE cells were exposed respectively to IL-1β?Aand NS. (for detecting the constitutive expression of NF-κB in PDTC-pretreated hRPE cells). The expression of NF-κB in hRPE cells in two groups was detected by immunofluorescence stain and flow cytometry. The results showed that the constitutive expression of NF-κB in hRPE cells in vitro was 8.05 %, and increased to 30.26 % by IL-1β. After PDTC pretreatment, the constitutive expression of NF-κB in hRPE cells was decreased to 3.74%, and 3.66 % by IL-l,respectively. It was concluded that the expressions of NF-κB in hRPE cells could be increased significantly by IL-1βand depressed effectively by PDTC. Also, PDTC could significantly inhibit the activation of NF-κB induced by IL-1β.     

Expression of SODD and P65 in ALL of Children and Its Relationship with Chemotherapeutic Drugs

The expression of silience of death domains (SODD) and its clinical significance and relationship with phospho-NF-κB-p65 proteins in bone marrow cells of childhood acute lymphoblas- tic leukaemia (ALL) were explored, and the expression of SODD and phospho-NF-κB-p65 in Jurkat cells treated with chemotherapeutic drugs was detected in order to find a new chemotherapeutic target. The expression of SODD and phospho-NF-κB-p65 proteins in bone marrow cells was detected by immunohistochemistry in 25 children with ALL. The apoptosis rate was measured by An- nexin-V-Fluorescence/PI double-labeling flow cytometry and the expression of SODD and phos- pho-NF-κB-p65 proteins determined by Western blotting in the Jurkat cells. It was found that the ex- pression of SODD and active P65 in ALL was significantly higher than that in normal control group (P<0.05). The expression of the SODD and phospho-NF-κB-p65 proteins in the high-risk (HR) group was significantly higher than that in the standard-risk (SR) group (P<0.05). The Pearson rank correla- tion analysis revealed that there was a positive correlation between SODD and phospho-NF-κB-p65 expression (P<0.01, r=0.69). VCR could effectively induce the apoptosis of Jurkat cells, and down-regulate the expression of SODD and phospho-NF-κB-p65 proteins in a time-dependent man- ner, but DNR could not down-regulate the expression of SODD effectively. It was concluded that SODD may be closely related to the clinical classification and prognosis of ALL in children. The ex- pression of SODD and phospho-NF-κB-p65 had a definite synergistic relationship with the onset and development of ALL. VCR could down-regulate the expression of SODD and inhibit the NF-κB ac- tivation, which could recover the sensibility of apoptosis in leukemic cells.     

Expression of NF-κB in Schwann cells and its effect on motor neuron apoptosis in spinal cord following sciatic nerves injury in rats

Objective: To explore the expression of nuclear factor-kappa B (NF-κB) in Schwann cells (SCs) and its effect on motor neuron apoptosis in spinal cord following sciatic nerves injury in adult rats. Methods: Thirty-six adult Sprague-Dawley (SD) rats were divided randomly into normal control group (n=6), and sciatic nerves crushing group (n=30). and the later was further equally randomized into 5 subgroups: 1. 3. 7. 11. and 21 d post-injury groups. The expression of NF-κB of normal and injured nerves were examined by immunohistochemistry staining, and the apoptosis of motor neurons in spinal cord of lumbar 4 to lumbar 6 (L4-L6) was investigated by terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNED assay. Both were quantitated by image analysis. Results: In crushing group, except 21 d post-injury group, the expression of NF-κB was markedly higher than that in the normal control group (P<0. 05,P<0. 01). At 1 d after sciatic nerves crushing, the expression of NF-κB was obviously up-regulated, reached peak at 3 d. and recovered at 21 d. The same trend was observed in the time-course on motor neuron apoptosis after sciatic nerves injury. Correlation analyses revealed that motor neuron apoptosis was significantly and positively correlated with the expression of NF-κB following sciatic nerves injury (r=0. 976 0,P<0. 01). Conclusion: After injury of sciatic nerves, the presence and up-regulation of NF-κB in SCs may be involved in motor neuron apoptosis in L4-L6 spinal cord.     

肠炎清通过抑制NF-κB活性对大鼠肠黏膜起抗炎作用

目的 检测大鼠小肠炎模型肠黏膜通透性的改变,探讨肠炎清对大鼠小肠炎模型肠黏膜通透性的作用的机制.方法 应用氨甲碟呤制备大鼠小肠炎模型,实验设正常对照组、模型对照组、N-乙酰半胱氨酸(NAC,100 mg/kg)组及肠炎清组(CYQ,100mg/kg),每天灌胃给药1次,共6d.每天观察大鼠疾病活动指数(DAI)和肠黏膜损伤指数(CMDI),光镜下观察组织学改变并评分(HS).ELISA测定大鼠IL-10,RT-PCR方法检测大鼠肠黏膜内TNF-α、IL-β的表达,应用Western blotting测定各处理组细胞内磷酸化胞质IκB蛋白表达水平.结果 与正常组相比,小肠炎模型组DAI、CMDI、HS评分明显升高(P<0.01).肠炎清组DAI,CMDI,HS评分较模型组有明显下降(P<0.01).MTX组TNF-α、IL-1βmRNA达水平均较正常对照组明显增高,肠炎清组与NAC治疗组TNF-α、IL-1β表达水平低于MTX组,但IL-10的表达高于MTX组.肠炎清能抑制IκB的降解.结论 姜黄素对肠黏膜起保护作用的机制可能是抑制NF-κB活性,进而减少致炎细胞因子TNF-α、IL-1βmRNA表达,增强抑炎因子IL-10的表达来实现的

Abstract：

Objective To detect the changes in intestinal mucosal permeation in rats with methotrexate-induced small intestinal damage and investigate the protective effects of Changyanqing decoction. Methods Rat enteritis model was established by methotrexate (MTX) and sodium chloride.The rats were randomly divided into normal control group,model group,N-acetylcysteine(NAC) group and Changyanqing decoction group,and Changyanqing decoction(100 mg/kg)or saline was administered daily in the corresponding groups by gastric irrigation for 6 days.The disease activity index(DAI),colonic mucosal damage index(CMDI)and histological score(HS) of the rats were observed and evaluated.The levels of mRNA expressions of TNF-α and IL-1β were detected by semi-quantitative RT-PCR.The expression of IL-10 was detected by enzyme linked immunosorbent assay,and IκB expression was determined with Western blotting.Results Compared with the normal control group,the model group showed significantly increased DAI,CMDI and HS.The DAI,CMDI,and HS in rats treated with Changyanqing decoction were significantly decreased in comparison with those in the model group (P<0.01).The expressions of TNF-α and IL-1β were significantly higher in MTX-treated group than in the control group.The expression of TNF-α and IL-1β mRNA in the Changyanqing group and NAC group were significantly lower,but IL-10 significantly higher than those of the MTX group.In MTX group,obvious NF-κB activation was observed,whose expression was significantly stronger in the cell nuclei,and the IκB in the cytoplasm was markedly degraded.Conclusion Changyanqing decoction offers protection on intestinal mucosa by inhibiting NF-κB activation to reduce TNF-α and IL-1β mRNA expressions and increase IL-10 expression.     

Role of Nuclear Transcription Factor-κB in Endotoxin-induced Shock in Rats

To investigate the role of NF-κB in endotoxic shock in rats. the model of endotoxinshock rats was induced by intravenous infusion of lipopolysaccharidc (LPS). 1 h. 2 h. 4 h and 6 h after LPS injection, the activation of NF-κB in blood mononuclear cells and the content of TNF-α and IL-6 in plasma was detected by enzyme-linked immunoadsordent assay (ELISA). The level of mean arterial pressure (MAP) and the histopathological changes of lung and liver were also observed. The activation of NF-κB in mononuclear cells increased 1 h after LPS injection and reached its peak 2 h after the injection, and its level was higher than that of normal group. The level of TNF-α was increased 1 h after the infusion and peaked 2 h after the injection, and its level was higher than that of normal group after LPS infusion. The content of IL-6 increased gradually with time. the IL-6 level was higher than that of normal group after LPS injection. MAP was decreased gradually with time and its level was lower than that of normal group after LPS injection. Pathological examination showed that endotoxic shock could cause pulmonary alveolar hemorrhage, edema and infiltration of inflammatory cell in lung tissue and congestion, edema, capillary dilation and inflammatory cell infiltration in liver tissue. It is concluded that NF-κB can up-regulate the expression of TNF-α and IL-6 in plasma and play an important role in endotoxin induced shock in rats.     

Effects of interleukin-18 on asthmatic airway inflammation and nuclear fa ctor kappa-B in murine models

Objective To investigate the effects of Interleukin-18 (IL-18) on asthmatic airway infla mmation and nuclear factor kappa-B (NF-κB) in a murine asthmatic model. Methods BALB/C mice were randomly divided into three groups: group A(control group,n=10) ; group B (asthmatic model group, n=10); group C (IL-18 injection group, n=10) . The asthmatic model was established in group B and C by respiratory syncytial virus (RSV) killed by ultraviolet light. Saline solution (0.1 ml) and IL-18 (0.1 ml, 1 μg) was injected in groups B and C at seven time points (1, 2, 7, 8 , 9, 21, 22 d). The symptoms and the numbers of eosinophils and plasmacytes in the airways were observed and the expression of NF-κB activation in the lung w as analyzed by Immohistochemistry (IHC) and Western blot. Results The symtoms of group C were more severe than in groups A and B. Group A did no t have EOS and plasmacytes in the airway submucosal while the numbers of eosinop hils ［15±3 (average cell counts per microscopic visual field, the same below) ］ and plasmacytes (10±2) in group B were found to have increased significantly . But the numbers of eosinophils and plasmacytes in group C were decreased sig nificantly when compared with group B (6±2 and 2±1, respectively, both P&lt;0 .05). ISH showed that the expression of NF-κB activation in group B was stro nger than that in groups A and C. The amount of NF-κB inhibitor (IκB) in gro up A and group C were 3.5 times and 2.5 times more than that of group B respec tively via Western blot. Conclusion IL-18 can inhibit asthmatic airway inflammation in mice and its mechamism may b e due to the fact that IL-18 can inhibit the activation of NF-κB in the murin e asthmatic model.     

Caspase-3和Bcl-2在骨肉瘤中的表达及意义

目的：探讨Caspase-3及Bcl-2的异常表达与骨肉瘤发生、发展的关系。方法：应用免疫组织化学方法检测骨肉瘤、骨软骨瘤中Caspase-3和Bcl-2的表达,并结合临床资料对结果进行统计学分析。结果：在41例骨肉瘤患者中,Caspase-3阳性19例,阳性率为46．3％,低于骨软骨瘤中Caspase-3的表达;其中G1期骨肉瘤表达明显高于G2期（P〈0．05）;Bcl-2阳性表达14例,阳性率为34．1％,显著高于骨软骨瘤;其中G1期组的表达明显高于G2期组（P〈0．001）。14例Bcl-2阳性的骨肉瘤病例中,Caspase-3阳性13例,两者阳性比率为92．9％;27例Bcl-2阴性的病例中,Caspase-3阴性22例,两者阴性比率为81．5％。两者有很好的相关性（P〈0．001）。结论：Bcl-2对细胞凋亡的抑制是骨肉瘤的早期分子事件,同骨肉瘤的发生关系密切,Caspase-3表达下调同骨肉瘤的发生、发展有关,在骨肉瘤中细胞凋亡的抑制是两者相互作用的结果。     

砷染毒大鼠生精细胞Bcl-2表达的变化

目的观察不同剂量的三氧化二砷（As2O3）对成年大鼠生精细胞Bcl-2表达的影响。方法40只健康雄性SD大鼠随机分成4组，分别为0．375mg/kg、0．75mg/kg、1．5mg/kg 3个染毒组和对照组，灌胃法连续给药16w后对各组大鼠计算睾丸脏器系数、精子头计数，并计算每日精子生成量（DSP）。采用免疫组化法检测大鼠生精细胞Bcl-2表达的情况。结果①中剂量组（0．75mg/kg）和高剂量组（1．5mg/kg）睾丸精子头计数和DSP均低于对照组（P〈0．05）；②与对照组相比，中、高剂量组大鼠生精细胞Bcl-2阳性产物表达明显降低（P〈0．01）；③生精细胞Bcl-2阳性率与每日精子生成量呈正相关（r=0．589,P〈0．01）。结论一定剂量的As2O3可通过抑制生精细胞Bcl-2的表达而导致精子生成量的减少，产生雄性生殖毒性。     

DKK3和SFRP2基因甲基化在肿瘤中的研究进展

DKK3和SFRP2基因是W nt信号传导通路中的抑癌基因,其启动子区CpG岛的甲基化与乳腺癌、肠癌、肺癌、肝癌等许多恶性肿瘤的发生、发展密切相关,在恶性肿瘤中甲基化或高甲基化,在癌旁组织或正常组织中无甲基化或低甲基化,并且甲基化导致其基因表达明显下调。     

胃癌组织中Galectin-3及Bcl-2的表达

目的 研究Galectin-3在胃癌组织中的表达水平与临床病理之间的关系，初步探讨Galectin-3及Bcl-2在胃癌组织中表达的相关性。方法 用免疫组化方法检测了60例胃癌患者的组织中Galectin-3及Bcl-2的表达，同时随机选择其中的15例癌旁组织作为对照组进行检测。结果 胃癌组织中Galectin-3及Bcl-2的表达率分别为76．7％（46／60）和68．3％（41／60），远高于癌旁组织中的20％（3／15）和40％（6／15）。Galectin-3与Bcl-2在胃癌组织中的表达无显著性差异（P〉0．05）；Galectin-3的阳性表达与癌组织的分化程度无关，但在有淋巴结转移的癌组织中表达水平显著提高（P〈0．05）；胃癌中Galectin-3与Bcl-2的表达在临床高分期时显著增强（P〈0．05）。结论 Galectin-3及Bcl-2是良好的胃癌组织表面标记物，在进展期胃癌及有淋巴结转移的胃癌中表达增高，可用于胃癌的诊断，且Galectin-3与Bcl-2的表达有明显相关性，可能存在共同的抗凋亡细胞内信号通路。     

尖锐湿疣细胞凋亡与caspase-3、bcl-2的表达

目的探讨尖锐湿疣（CA）细胞凋亡与easpase-3、bcl-2表达及其相互关系。方法对于33例CA、5例巨大CA和8例正常上皮用TUNEL法原位检测角质形成细胞（KC）的凋亡，以免疫组化法检测caspase-3、bcl-2的表达。结果在CA和巨大CA两组之间，凋亡指数（AI）、easpase-3及bcl-2的阳性表达率均无统计学差异（P〉0．05）。CA组的Al与easpuse-3的表达水平呈极显著正相关（r=0．48979，P〉0．0001），与bcl-2的表达水平呈极显著负相关（r=0．51470，P〉0．0001），caspctse-3与bcl-2的表达水平呈极显著负相关（r=0．70165，P〉0．0001）。结论CA的KC中存在着明显的细胞凋亡现象，caslxtse-3和bcl-2可能参与了CA细胞凋亡的调节机制。     

Insulin Resistance and Type 2 Diabetes Mellitus: Its Relationship with the β3-Adrenergic Receptor

The β₃ subtype of adrenaline and noradrenaline receptors has been extensively characterized at structural and functional levels. Ligand binding and adenyl cyclase activation studies have helped to define their unique β-adrenergic profile. Humans, other larger mammals, and rodents share most of the characteristic β₃-adrenergic receptor properties, although obvious species-specific differences have been identified. Most studies in animal models have shown a distinct β₃-adrenergic receptor activity that results in an increase in energy expenditure, decrease of fat mass (especially of intra-abdominal fat), and increased glucose disposal efficiency. It is of interest that mild weight increase was shown to develop in female but not male mice, in whom the β₃-adrenergic receptor gene was disrupted. Recently, the incidence of a naturally occurring variant of the human β₃-adrenergic receptor was shown to correlate with hereditary obesity in Pima Indians and Japanese individuals. In Western obese patients, this phenotype increased the capacity to gain weight and develop type 2 diabetes mellitus. Studies of humans with the Trp⁶⁴Arg variant have shown controversial results. Many studies have failed to show any effect in heterozygous male subjects, and only modest effects in homozygous male subjects. In women, several studies have shown modest-to-significant effects regarding weight gain, intra-abdominal fat, and decreased insulin sensitivity in heterozygous and homozygous women. Other studies have failed to show any effect in heterozygous females.Disruptions in the activity of the β₃-adrenergic receptor in the homozygous male and the heterozygous or homozygous female appear to have a profound effect in animal models, but a limited consequence in human physiology. Association with obesity or diabetes in humans is still controversial. This difference between animal and human models may be explained by the different quantity and distribution of metabolically active brown adipose tissue in the two.     

(2S,3R)-1-二甲氨基-3-(3-甲氧基苯基)-2-甲基戊-3-醇的合成工艺研究

目的 对(2S,3R)-1-二甲氨基-3-(3-甲氧基苯基)-2-甲基戊-3-醇合成工艺进行研究。方法 以3-戊酮为起始原料,经Mannich反应、手性拆分、Grignard反应等步骤合成(2S,3R)-1-二甲氨基-3-(3-甲氧基苯基)-2-甲基戊-3-醇,并对化学拆分进行工艺优化。结果 合成(2S,3R)-1-二甲氨基-3-(3-甲氧基苯基)-2-甲基戊-3-醇,总收率为30.6%。结论 工艺改进后提高了目标产物的收率,同时也减少了原料的浪费。     

稳定表达Trop-2基因的NIH3T3细胞系的构建和特性分析

目的:建立稳定表达人滋养层细胞表面抗原(Trop-2)NIH3T3细胞,分析过表达Trop-2对NIH3T3细胞的生长?增殖和侵袭特性的影响?方法:将Trop-2基因克隆到真核表达载体pcDNA3.1,转染NIH3T3细胞,通过G418筛选及RT-PCR鉴定获得稳定表达Trop-2的NIH3T3细胞(NIH3T3-Trop-2)?用MTS法检测NIH3T3-Trop-2细胞的增殖能力,软琼脂集落形成实验检测NIH3T3-Trop-2细胞的克隆形成能力,明胶酶谱法检测NIH3T3-Trop-2细胞的基质金属蛋白酶(MMP)-2和MMP-9的分泌及细胞划痕实验检测NIH3T3-Trop-2细胞的迁移能力?结果:稳定表达Trop-2的NIH3T3细胞在生长增殖?克隆形成及侵袭能力均较NIH3T3细胞强,细胞培养上清中的MMP-2和MMP-9增多?结论:Trop-2对细胞的增殖与迁移能力具有明显的促进作用?     

Skp2、Caspase-3在膀胱尿路上皮细胞癌组织中的表达

目的检测Skp2、Caspase-3在膀胱尿路上皮细胞癌（简称膀胱癌）组织中的表达,探讨其临床意义及相关性。方法采用免疫组化方法检测Skp2、Caspase-3蛋白在82例膀胱癌组织和16例正常膀胱黏膜组织中的表达,并收集临床资料,分析检测结果与临床病理的联系及二者的相关性。结果 Skp2在膀胱癌组织中的阳性率（43.9%）高于正常膀胱黏膜组织（12.5%）（P〈0.05）;Caspase-3在膀胱癌组织中的阳性率（39%）低于正常膀胱黏膜组织（81.3%）（P〈0.05）。膀胱癌细胞的恶性程度越高,Skp2蛋白表达越高,Caspase-3则无明显差异;与非肌层浸润性膀胱癌相比,肌层浸润性膀胱癌组织中Skp2蛋白的表达较高（P〈0.05）,而Caspase-3的表达则较低（P〈0.05）。膀胱癌组织中Skp2蛋白的表达与Caspase-3成负相关关系。结论 Skp2蛋白的表达与膀胱癌的分化、临床分期密切相关,与Caspase-3的表达成负相关;联合检测Skp2、Caspase-3有助于判定膀胱癌的侵袭能力。     

E2F3基因沉默载体pRNAT-U6.1-siE2F3/Neo的构建及表达

【目的】构建针对E2F3基因的shRNA表达载体,通过脂质体进行膀胱癌细胞内转染,监测转染效率。【方法】通过软件设计针对E2F3基因的寡核苷酸链,化学合成一对编码短发夹RNA序列,长度为64个碱基。核苷酸链经退火,克隆到经BamHI、HindⅢ双酶切的pRNAT-U6.1/Neo载体上,构建针对E2F3基因的RNA干扰质粒。通过双酶切、PCR鉴定及基因片段序列分析验证构建效果。通过不同浓度的Lipofectamin 2000及干扰质粒混合物进行膀胱癌细胞系5637细胞转染,通过绿色荧光的表达程度摸索出细胞最佳转染浓度及转染时间。【结果】重组构建的pRNAT-U6.1-siE2F3/Neo载体经PCR分析及插入基因片段序列分析,结果表明64个碱基成功插入到预计位点,并且序列完全一致。Lipofectamin 2000能够有效的进行pRNAT-U6.1-siE2F3/Neo质粒的转染。【结论】载体的成功构建及膀胱癌细胞系的高效转染,为进一步研究E2F3基因在膀胱肿瘤中的功能奠定基础。