内毒素耐受和拟胆碱药物对巨噬细胞分泌肿瘤坏死因子-α的影响

目的:探讨内毒素耐受和拟胆碱药物卡巴胆碱对巨噬细胞肿瘤坏死因子-α(TNF-α)分泌的调节作用.方法:①分离小鼠腹腔巨噬细胞,将其与内毒素(LPS)作用不同时间(0、2、4、8 h);②小鼠腹腔巨噬细胞分为3组:空白对照组(RPMI1640常规培养),LPS耐受组(LPS预刺激20 h,再用LPS刺激4 h)和LPS不...     

Impact of body mass index on seminal oxidative stress

Male obesity has been linked with a reduction in sperm concentration and motility, an increase in sperm DNA damage and changes in reproductive hormones. Recent large observational studies have linked male obesity with a reduced chance of becoming a father. One of the potential underlying pathological mechanisms behind diminished reproductive performance in obese men is sperm oxidative stress. The primary aim of this study was to determine if sperm oxidative stress was more common in obese/overweight men. A total of 81 men had their body mass index (BMI) correlated with seminal reactive oxygen species (ROS) production (Nitro Blue Tetrazolium assay), sperm DNA damage (TUNEL), markers of semen inflammation (CD45, seminal plasma PMN elastase and neopterin concentration) and routine sperm parameters, together with reproductive hormones. The principal finding from this study was that oxidative stress did increase with an increase in BMI, primarily due to an increase in seminal macrophage activation. However, the magnitude of this increase was small and only of minor clinical significance as there was no associated decline in sperm DNA integrity or sperm motility with increasing ROS production. Increased BMI was also found to be significantly linked with a fall in sperm concentration and serum testosterone, and an increase in serum oestradiol.     

失血性休克大鼠血浆及腹腔巨噬细胞释放细胞因子改变

目的：研究大鼠失血性休克前后血浆及腹腔巨噬细胞释放细胞因子的变化。方法：将30只大鼠分为失血性休克组（20只）和对照组（10只）,观察大鼠休克前后血浆和腹腔巨噬细胞释放肿瘤坏死因子（TNF）－α,白细胞介素（IL）－1β,IL－6和IL－8改变。结果：失血性休克90min后大鼠腹腔巨噬细胞释放TNF－α,IL－1β,IL－6和IL－8显著减少（P＜0．01）,而血浆中这些细胞因子水平明显升高（P＜0．01）。结论：失血性休克对大鼠细胞免疫功能有抑制作用。     

巨噬细胞及肿瘤坏死因子在肝细胞癌中的表达及意义

目的检测巨噬细胞及肿瘤坏死因子-α（TNF-α）在肝细胞癌（HCC）中的表达,同时比较其与微血管密度指标CD31、vWF和细胞增殖指标（PCNA）、病理指标及预后的相关性。方法采用免疫组化、流式细胞术及原位杂交法,对53例肝细胞癌的标本进行检测,并将检测数据与病人的临床资料进行统计分析。结果癌组织中的巨噬细胞数目与肿瘤直径呈负相关,癌旁组织中的巨噬细胞数目与微血管密度呈正相关;HBsAg阳性及肝内门静脉浸润组的癌旁组织中TNF-α表达量较低,男性病人癌组织中TNF-α表达量较低;微血管密度计数结果显示,CD31与肝内门静脉浸润有关,vWF与肿瘤的TNM分期有关;PCNA与肿瘤TNM分期有关。结论在HCC中,巨噬细胞的作用呈双向性,TNF-α、vWF、CD31及PCNA指数均与预后有关。     

Diagnostic significance of the histopathology of bone marrow macrophages in forensic autopsies

Forensic pathologists often encounter autopsies that require an assessment of antemortem general conditions (e.g., infection, metabolic disorders). To establish evaluation clues for such cases, we quantitatively examined macrophages and the general pathology of bone marrow in samples from 180 forensic autopsy cases of decedents with various conditions. Hematoxylin-eosin staining, Berlin blue staining, and immunostainings for CD163, CD138, and CD61 were performed. We determined the numbers per field (density) of total macrophages, swollen macrophages, macrophages with hemophagocytosis, and hemosiderin-laden macrophages. Each density was standardized by identifying its ratio to the total number of macrophages. The decedents' background data (cause of death, other pathological findings, postmortem interval, antemortem symptoms, and presence of resuscitation) were extracted. No correlations were found between the postmortem interval and the other decedent data, indicating that these data are not affected by postmortem changes. In the group in which inflammatory disease was the cause of death, there were significant elevations in the ratio of the swollen macrophage density to total macrophages. Significantly higher ratios of the density of swollen and hemophagocytic macrophages were observed in the group in which conditions with a prolonged agonal period were the cause of death. The group with a return of spontaneous circulation to resuscitation showed a significantly higher ratio of macrophage density with hemophagocytosis. This study provides the first statistical analysis focused on bone marrow histopathology in forensic autopsies. The results will be useful for elucidating causes of death and agonal-period conditions.     

NFκB抑制剂对大鼠肝脏移植再灌注损伤的保护作用

目的　探讨大鼠肝脏移植后核因子κB(NFκB)活化对炎性介质表达和中性粒细胞浸润集聚的影响。方法　供受体鼠分别于术前 15min腹腔注射NFκB抑制剂脯氨酸二硫代氨基甲酸酯 (ProDTC 15mg/kg)。 结果　与对照组相比 ,应用ProDTC者移植后NFκBp6 5含量、肿瘤坏死因子 α(TNF α)、巨噬细胞炎性蛋白 2 (MIP 2 )、细胞间粘附分子 1(ICAM 1)mRNA表达和蛋白表达明显下降 ;髓过氧化物酶 (MPO)活性明显减低 (P均 <0 0 1)。ProDTC也显著降低天门冬氨酸转氨酶(AST)、丙氨酸转氨酶 (ALT)、乳酸脱氢酶 (LDH)水平 (P <0 0 1)。结论　ProDTC抑制移植后NFκB活化从而下调了TNF α、MIP 2、ICAM 1表达从而减轻中性粒细胞浸润及肝脏缺血再灌注损伤。     

Cyclooxygenase-2 Inhibition Improves Macrophage Function in Melanoma and Increases the Antineoplastic Activity of Interferon γ

Background: Melanoma inhibits macrophage tumoricidal activity and increases the expression of cyclooxygenase-2 (COX-2). In this study, we sought to determine whether inhibition of COX-2 could restore macrophage function and hence maximize the antitumor activity of the immune stimulant interferon (IFN).Methods: Peritoneal macrophages were exposed to B16 melanoma-conditioned medium for 24 hours with or without the COX-2 inhibitor NS-398 and then were stimulated with lipopolysaccharide and IFN. Cytotoxic activity, nitrite production, and cytokine production by the stimulated macrophages were measured. In addition, B16 melanoma cells were implanted intradermally into mice treated with IFN (14,000 U on alternate days) alone or with a combination of IFN and a COX-2 inhibitor (NS-398 or nimesulide). Mice were assessed for tumor growth and survival.Results: Macrophage cytotoxicity and nitrite production were significantly suppressed by melanoma-conditioned medium (P < .01). This was prevented by 200 M of NS-398 (P < .05). In vivo, combined treatment with IFN and a COX-2 inhibitor caused a significant inhibition of tumor growth (P < .01) and improved survival (P = .02) compared with controls.Conclusions: COX-2 inhibition reversed melanoma-induced suppression of macrophage function, and combined treatment of IFN plus a COX-2 inhibitor was maximally effective in reducing tumor growth and improving survival.     

巨噬细胞移动抑制因子在大鼠急性坏死性胰腺炎模型中的作用研究

目的初步探讨巨噬细胞移动抑制因子（MIF）在大鼠急性坏死性胰腺炎（ANP）发病中的作用。方法健康雄性SD大鼠60只,随机分成三组,分别为对照组（腹腔注射生理盐水）、ANP组（腹腔注射左旋精氨酸）和干预组（腹腔注射左旋精氨酸＋单克隆抗MIF抗体）,每组20只。采用左旋精氨酸改良方法建立ANP模型,分别于3、6、12、24h四个不同时点处死5只大鼠,剖腹后从肠系膜上静脉取血,ELISA法测定血清MIF、TNF-α、IL-1、IL-6和IL-8水平 碘比法测定血淀粉酶 切取胰腺组织依据Kusske标准行胰腺病理学评分 Western blot法测定胰腺组织NF-κBp65蛋白的表达。结果ANP组血淀粉酶及胰腺组织病理学评分各时点㈦对照组相比均显著升高,干预组㈦ANP组相比均显著降低（P〈0.01） 胰腺组织NF-κBp65蛋白的表达于造模后3h开始呈持续性上调,与对照组相比其表达显著增多,在干预组其表达水平㈦ANP组相比明显下调（P〈0.01） 血清MIF、TNF-α、IL-1、IL-6各时点在ANP组与正常对照组相比其水平均有明显升高,在干预组其水平㈦ANP组相比显著降低（P〈0.05） 血清IL-8在6、12、24h不同时点水平变化同上述因子,但在3h时点其水平无明显升高 MIF、胰腺组织病理学评分及胰腺组织NF-κBp65蛋白的表达,两两之间均成正相关（P〈0.05）。结论腹腔注射左旋精氨酸建立ANP的模型是稳定的 MIF可能通过调控核因子NF-κB的途径影响血清TNF-α、IL-1、IL-6及IL-8水平而在大鼠急性胰腺炎发病过程中起一定的作用。     

MIF和Cyclin D1在肝细胞癌中的表达

目的 探讨巨噬细胞游走抑制因子(macrophage migration inhibitory factor,MIF)和细胞周期蛋白D1(cyclin D1)在原发性肝细胞癌(hepatocellular carcinoma,HCC)组织中的表达及两者在肝癌发病机制及细胞周期调控中的关系.方法 应用定量PCR及Westem blot技术检测MIF和Cyclin D1在肝癌组织和癌旁组织的表达.化学合成MIF siRNA,将PLC细胞和HepG2细胞分成对照组、MIF siRNA 50 nmol/L干预组、MIF siRNA 1130 nmol/L干预组,脂质体法转染肝癌细胞PLC和HepG2,定量PCR技术和Western blot检测MIF siRNA干扰后MIF和Cyclin D1 mRNA和蛋白的表达变化.结果 MIF和Cyclin D1蛋白在肝癌组织中的相对表达量为0.825±0.13和0.843±0.104;MIF和Cyclin D1 mRNA在肝癌组织中的相对表达量为癌旁组织的(7.31±1.85)倍和(4.27±1.05)倍,与癌旁组织相比,差异均具有统计学意义(FMIF=15.5,P<0.01;fCyclin D1=87.5,P<0.01).应用小RNA干扰技术转染PLC和HepG2肝癌细胞后MIF mRNA分别下调71.2%±7.2%、87.4%±2.9%、74.3%±8.9%、88.4%±4.6%(FPLC=315.5,P<0.01;FHHepG2=201.2,P<0.01);MIF蛋白分别下调为0.33±0.03、0.11±0.02、0.81±0.08、0.36±0.02,并呈剂量依赖关系(FPLC=43.9,P<0.01;FHepG2:133.4,P<0.01).伴随MIF mRNA和蛋白的表达F调Cyclin D1 mRNA分别下调68.2%±3%、78.1%±1.4%、65.8%±4.7%、77.3%±2.6%(FPLC=1569,P<0.01;FHepG2=480.4,P<0.01);Cyclin D1蛋白下调0.28±0.06、0.15±0.03、0.44±0.04、0.13±0.02,亦呈剂量依赖关系,与对照组相比差异有统计学意义(P<0.01),两干预组相比差异有统计学意义(FPLC=35.5,P<0.01;FHepG2=114.7,P<0.01).结论 MIF和Cyclin D1在肝细胞癌中过表达,MIF可能在转录水平调控Cyclin D1的表达,并促使肿瘤细胞通过细胞周期检测点,继续增值和分化,导致肿瘤的形成.     

人脾脏巨噬细胞总RNA的提取和产率研究

目的　探讨提取人脾脏巨噬细胞总RNA的方法并测算其产率。方法　收集 5例门静脉高压症脾亢患者的手术切除脾脏 ,经贴壁培养法分离纯化出巨噬细胞 (Mφ)后 ,采用TRIzol试剂一步法提取 5例脾脏Mφ样本的总RNA。分光光度法和琼脂糖凝胶电泳检测总RNA的产量和质量。最后计算出每 10 8个Mφ总RNA的产率。 结果　 5例样本Mφ总RNA的A2 60nm与A2 80nm比值平均为 1.83± 0 .13 ,提示总RNA纯度较高 ,电泳结果提示总RNA无降解。每 10 8个Mφ平均可抽提 (4 1.5 8± 2 3 .13 ) μg总RNA。 结论　采用TRIzol试剂一步法提取脾脏Mφ总RNA是可行的 ,收获的人脾脏Mφ总RNA纯度高、完整性好、量较多 ,可满足后续实验要求。