高钙拮抗氟对人牙乳头间充质细胞毒性的体外试验研究

目的观察高浓度钙对氟细胞毒性的影响.方法体外培养的人牙乳头间充质细胞给予不同浓度氟10～2 630 μmol/L处理,将有明显氟细胞毒性的实验组同时予高浓度钙(4.1,10 mmol/L)处理,用S四唑盐(MTT)比色法和透射电镜观察细胞活性和超微结构.结果 263,1 315,2 630×10-6mol/L氟对人牙乳头间充质细胞毒性明显,10×10-3mol/L和4.1×10-3mol/L钙对1 315×10-6mol/L浓度以下氟毒性有明显拮抗作用,对2 630×10-6mol/L氟毒性作用不确定.结论氟细胞毒性在一定范围内可被高钙拮抗.     

氟中毒大鼠血清碱性磷酸酶活性及钙,磷水平的动态变化

本实验复制了慢性氟中毒大鼠病理模型、对血清碱性磷酸酶（ＡＬＰａｓｅ）活性和钙、磷水平进行了动态观察。结果表明，血清碱性磷酸酶活性在中毒１４ｄ内升高，然后下降，在３０ｄ时达到谷底，到９０ｄ时又恢复到原水平。而血清钙、磷水平在１４ｄ以后才开始下降，３０ｄ达到谷底，９０ｄ恢复到原水平。这一变化规律在饮水含氟（ＮａＦ）达０．１ｍｇ／ｍＬ时最明显。本文用“Ｍｇ＾２＋假说”解释了这一变化规律。同时对氟中毒的损     

体外诱导人羊膜间充质干细胞分化为胰岛素分泌细胞

目的 探讨体外诱导人羊膜间充质干细胞(hAD-MSCs)向胰岛素分泌细胞分化潜能.方法 采用胰蛋白酶-胶原酶消化法分离提取hAD-MSCs,流式细胞术分析和免疫细胞化学染色行表型鉴定.取第3代按2.5× 106个/ml或5×105个/ml细胞密度接种6孔培养板或预置盖玻片的24孔培养板,在含10mmol/L尼克酰胺和N2补充物的无血清HG-DMEM培养基培养,未诱导组为含10％胎牛血清的LG-DMEM基础培养基.分别于体外诱导第7、14、21天采用免疫细胞化学法检测胰岛素和β2微球蛋白的表达,采用放射免疫法检测上清液中胰岛素含量,采用RT-PCR检测胰岛素mRNA和胰十二指肠同源异型盒因子1(PDX-1)mRNA的表达.结果 (1)hAD-MSCs高表达间充质干细胞表面标志CD29、CD44、CD73、CD166及波形蛋白;(2) hAD-MSCs诱导第7、14、21天胰岛素阳性细胞百分率分别为74.67％±1.53％、75.00％±1.00％、74.33％±1.53％,培养物上清液中胰岛素含量分别为(331.62±1.76)、(330.50±1.22)和(331.65±0.48) μIU/ml,各时点间比较无显著性差异(均P＞0.05),而未诱导组仍未见胰岛素阳性细胞;培养上清也未检测到胰岛素;(3) hAD-MSCs诱导前后均有PDX-1 mRNA和蛋白表达,胰岛素基因mRNA表达仅见于诱导组;(4) hAD-MSCs诱导组和未诱导组各时点均有β2微球蛋白表达,其阳性细胞百分率组间差异无显著性(均P＞0.05).结论 hAD-MSCs具有向胰岛素分泌细胞分化的能力,可能成为1型糖尿病细胞移植治疗的新的细胞供源.     

利拉鲁肽在人骨髓间充质干细胞诱导分化为胰岛素分泌细胞中的作用

目的 观察利拉鲁肽在入骨髓间充质干细胞( hBM-MSCs)向胰岛素分泌细胞(IPCs)方向分化诱导中的作用.方法 采用高糖、尼克酰胺和利拉鲁肽3阶段诱导方案对hBM-MSCs进行定向诱导分化,倒置显微镜下观察诱导过程中细胞的形态学变化,双硫腙染色法鉴定诱导后细胞,Western印迹法检测细胞胰腺十二指肠同源盒基因1( PDX-1)、葡萄糖转运蛋白2(GLUT2)、葡萄糖激酶(GK)、胰岛素的蛋白表达,ELISA检测细胞的胰岛素分泌水平.结果 添加10 nmol/L利拉鲁肽作用7d后诱导效率明显增加.诱导过程中细胞形态由长梭形逐渐变为圆形,并聚集生长,至诱导末出现大量圆形葡萄状聚集生长的胰岛样细胞团;双硫腙染色阳性细胞量、细胞PDX-1、GLUT2、GK、胰岛素的蛋白表达、细胞的基础和葡萄糖刺激的胰岛素分泌水平均逐渐增加(均P＜0.05).结论 在体外,高糖、尼克酰胺联合利拉鲁肽可使hBM -MSCs定向诱导分化为IPCs.     

人脐带血间充质干细胞对人胃癌MGC80-3细胞增殖的影响

目的探讨人脐带血间充质干细胞(h UCB-MSCs)对人胃癌MGC80-3细胞体外增殖的影响。方法原代培养h UCBMSCs,通过流式细胞仪进行细胞表型鉴定;采用MTT法测定不同浓度h UCB-MSCs条件培养基作用于胃癌MGC80-3细胞后,胃癌细胞24 h、48 h、72 h的抑制率;倒置相差显微镜观察胃癌MGC80-3细胞形态的变化;流式细胞技术测定胃癌MGC80-3细胞凋亡率。结果采用流式细胞仪表型分析获得的第4代h UCB-MSCs均质性好;MTT法显示,不同浓度h UCB-MSCs条件培养基处理MGC80-3细胞24 h、48 h、72 h后,各实验组抑制率均有不同程度升高;倒置相差显微镜观察显示细胞形态发生变化;流式细胞术分析结果显示,MSC-CM能诱导MGC80-3细胞发生凋亡,细胞凋亡率随着MSC-CM的升高而升高,各实验组与空白对照组比较及各实验组之间比较,差异均有统计学意义(P0.05)。结论 h UCB-MSCs能明显抑制人胃癌细胞株MGC80-3细胞的体外增殖与转移,且作用效果具有剂量和时间依赖性。     

体外原代培养脐带基质间充质细胞和人皮肤成纤维细胞

目的探讨体外原代培养脐带基质间充质细胞(UMC)和人皮肤成纤维细胞(HSF)的方法。方法用Ⅳ型胶原蛋白酶、Ⅰ型脱氧核糖核酸酶和0.25%胰蛋白酶消化法原代分离培养UMC细胞,用组织块贴壁法原代分离培养HSF细胞,镜下观察细胞的培养过程和生长状态。结果脐带组织分离培养第3天,有少量UMC贴壁生长,细胞膜周围有折光性,形态类似于成纤维细胞;皮肤组织块贴壁培养第7天,有HSF爬出,呈长梭形、不规则三角形。随着细胞培养时间延长,UMC和HSF数量逐渐增多,细胞生长状态良好。结论应用酶消化法和组织块贴壁法可成功分离培养出UMC和HSF。     

高海拔饮茶型氟中毒大鼠血清钙磷水平及碱性磷酸酶活性观察

目的探讨高海拔地区饮茶型氟中毒的发病机制：方法动态观察高海拔地区饮茶型氟中毒大鼠血清碱性磷酸酶(ALP)活性、钙(Ca)、磷(P)水平。结果砖茶水组ALP活性呈逐渐下降趋势．但仍高于对照组；砖茶水组血清Ca在30d时低于对照组，以后逐渐回升接近对照组水平；砖茶水组血清P在30d时高于对照组，以后逐渐下降接近对照组水平。结论高海拔饮茶型氟中毒可能存在较为复杂的发病机制。     

人脐带间充质干细胞在心力衰竭领域的临床研究进展

心力衰竭(HF)是各种心血管疾病的终末阶段,具有高患病率和病死率的流行病学特点.庞大的患者群体(我国约450万)和低生存率的预后特征给该疾病的治疗带来了巨大的挑战.目前HF的治疗包括药物治疗和非药物治疗.非药物治疗主要有心脏移植、左心室辅助装置置入术及细胞治疗等.近年来,包括间充质干细胞治疗在内的多种细胞疗法在修复受损...     

细胞传代对SD大鼠骨髓间充质干细胞衰老的影响

目的探究体外传代培养对SD大鼠骨髓间充质干细胞(BMSCs)衰老的影响。方法全骨髓贴壁法分离培养大鼠BMSCs,待细胞汇合率达80%～90%时,进行消化传代及纯化培养。对不同代数细胞,进行倒置显微镜下观察细胞形态学及生长特性,绘制1～7 d生长曲线,流式细胞仪检测细胞表面抗原标志物(CD29、CD45和CD90)和细胞周期,β-半乳糖苷酶染色并记录阳性细胞数,Western印迹法检测衰老相关蛋白P53和P21表达情况。结果 SD大鼠BMSCs的CD29、CD90高表达(95%),CD45低表达(5%),细胞周期中G0/G1期占87.37%;相较于P1～5代细胞,P7～9代细胞增殖能力明显下降,β-半乳糖苷酶染色阳性细胞明显增多;随着传代次数的增加,BMSCs的P53、P21表达增高。结论体外培养传代次数的增加可导致SD大鼠BMSCs衰老。     

人胚肺间充质干细胞对损伤肺组织的修复作用

目的 研究人胚肺间充质干细胞(MSCs)对损伤肺组织的修复作用.方法 通过动物体内实验和细胞体外培养实验,观察人MSCs对肺上皮细胞的生长调控.结果 人胚肺MSCs能有效调控肺泡上皮的生长.结论 人胚肺MSCs及其条件培养液可以减轻人肺上皮损伤和加速修复,从而为临床治疗COPD、病毒性肺炎造成的肺损伤提供新的生物治疗方法.     

骨髓间充质干细胞在缺血性脑血管病治疗中的应用

缺血性脑血管病的病死率和致残率均很高,给社会和家庭带来了巨大负担。但目前在临床上无论是药物治疗还是外科手术治疗效果均不理想。最近的研究发现,骨髓间充质干细胞具有多向分化潜能,能跨胚层分化为神经细胞,其作为神经干细胞的来源,用于缺血性脑血管病的治疗具有广阔的前景。     

The Effects and Mechanism of GSA on Expression of MCP-1 in Cultured Human Umbilical Vein Endothelial Cells

Objectives To investigate the effects and mechanism of glycated serum albumin(GSA) on expression of Monocyte chemoattratant protein-1(MCP-1) in Endothelial Cells. Methods Human Umbilical Vein Endothelial Cells (HUVEC)are cultured with GSA of different concentrations and interfered by glycosylation products inhibitor Aminoguanidine (AG) and anti-oxidant N-acetylcy-steine (NAC), The expression of MCP-1 are evaluated by Immunocytochemistry and Sandwich ELISA. MDA content and SOD activity are determined by the technique of TBA and XOD respectively. Results GSA can stimulate MCP-1 production and secretion. Immunocytochemistry showed that after HUVECs were cultured with 50 mg/L GSA, expression of MCP-1 in group 4hrs, 8hrs and 12hrs was 1.3, 1.9 and 2.8 fold as much as that in control group (P < 0.01), and there was significant difference among the experiment groups(P < 0.01). Sandwich ELISA showed that expression of MCP-1 in three different groups was 1.6, 2.4 and 3.0 fold as much as that in control group(P < 0.01), and there was significant difference among the experiment groups(P < 0.01); GSA can cause the decrease of SOD activity(P < 0.05) and increase of MDA content(P < 0.01); AG and NAC can restrain obviously the expression of MCP-1 of HUVECs stimulated by GSA(P < 0.01); NAC can restrain the effect of GSA on SOD activity and MDA content in HUVECs (P < 0.05). Conclusions GSA can stimulate the expression of MCP-1 of endothelial cells by inducing endothelial cells oxidative stress.     

诱导脐血间充质干细胞分化为神经细胞的研究进展

脐血间充质干细胞是从脐带血中分离和培养的一种多潜能成体干细胞,具有向神经细胞分化的潜能。近年来,脐血间充质干细胞治疗神经退行性疾病、神经损伤、卒中等神经系统疾病成为医学研究的热点之一,并取得了一定的进展。     

骨髓间质干细胞治疗脑梗死

脑梗死的发病率和致残率高,目前的治疗手段均未取得满意临床疗效。骨髓间质干细胞具有多向分化潜能,在体内和体外可转化为神经细胞,对脑梗死模型动物治疗后可取得神经功能改善。bcl-2具有抗凋亡作用,转染骨髓间质干细胞后,细胞内bcl-2高度表达可减少移植后干细胞凋亡。文章就骨髓间质干细胞治疗脑梗死的应用研究进展及前景进行了综述。     

The Effects of Glucocorticoids on Cultured Human Endothelial Cells

The effects of hydrocortisone, dexamethasone and prednisone on the morphology, replication, DNA synthesis, cell protein content and protein synthesis of cultured, human endothelial cells were evaluated. After culturing the cells with these glucocorticoids for 24-48 h, the cells covered a greater portion of the culture surface area. The mean surface area of the individual endothelial cell treated with glucocorticoids was 1.53 times greater than that of the untreated control endothelial cell. When compared with controls, the endothelial cover provided by the cells treated with glucocorticoids was more extensive and in many instances covered the entire culture surface. The change in morphology was associated with an increase in protein synthesis and protein content of the cells without an increase in DNA synthesis or cellular replication. Dexamethasone was approximately 10-fold more effective than hydrocortisone, while prednisone was the least effective. Aldosterone, DOCA, testosterone, progesterone, oestradiol and oestriol were ineffective. These studies indicate that glucocorticoids can alter the morphology and biochemistry of cultured endothelial cells and may have implications for the effects of steroids in the treatment of thrombocytopenic states and vascular disorders in man.     

Human Dental Pulp Cells Responses to Apatite Precipitation from Dicalcium Silicates

Unraveling the mechanisms behind the processes of cell attachment and the enhanced proliferation that occurs as a response to the presence of calcium silicate-based materials needs to be better understood so as to expand the applications of silicate-based materials. Ions in the environment may influence apatite precipitation and affect silicate ion release from silicate-based materials. Thus, the involvement of apatite precipitate in the regulation of cell behavior of human dental pulp cells (hDPCs) is also investigated in the present study, along with an investigation of the specific role of cell morphology and osteocalcin protein expression cultured on calcium silicate (CS) with different Dulbecco’s modified Eagle’s medium (DMEM). The microstructure and component of CS cement immersion in DMEM and P-free DMEM are analyzed. In addition, when hDPCs are cultured on CS with two DMEMs, we evaluate fibronectin (FN) and collagen type I (COL) secretion during the cell attachment stage. The facilitation of cell adhesion on CS has been confirmed and observed both by scanning with an electron microscope and using immunofluorescence imaging. The results indicate that CS is completely covered by an apatite layer with tiny spherical shapes on the surface in the DMEM, but not in the P-free DMEM. Compared to the P-free DMEM, the lower Ca ion in the DMEM may be attributed to the formation of the apatite on the surfaces of specimens as a result of consumption of the Ca ion from the DMEM. Similarly, the lower Si ion in the CS-soaked DMEM is attributed to the shielding effect of the apatite layer. The P-free DMEM group releases more Si ion increased COL and FN secretion, which promotes cell attachment more effectively than DMEM. This study provides new and important clues regarding the major effects of Si-induced cell behavior as well as the precipitated apatite-inhibited hDPC behavior on these materials.     

人巨细胞病毒对培养人脐静脉内皮细胞增殖和凋亡的影响

目的 :探讨人巨细胞病毒 (CMV)在动脉粥样硬化发生和发展中的作用。　　方法 :采用流式细胞计数仪观察了人 CMV对离体培养的人脐静脉内皮细胞 (HUVEC)增殖和凋亡的影响。本实验分为 5组 ,分别为正常对照组 ,肿瘤坏死因子细胞凋亡组 ,无血清培养细胞凋亡组 ,CMV感染 肿瘤坏死因子细胞凋亡组 ,CMV感染 无血清培养细胞凋亡组。　　结果 :实验发现无血清培养和肿瘤坏死因子诱导的细胞凋亡率较正常对照组升高 ,而 CMV感染的细胞凋亡率明显降低 ;细胞计数的结果表明 CMV感染后 3天的 HUVEC细胞数是未感染的 5倍。流式细胞仪的检测结果表明 :CMV感染细胞的增殖指数明显高于正常对照组 ,CMV可使细胞耐受无血清培养条件 ,保持细胞增殖。　　结论 :人 CMV感染 HUVEC后能够改变 HUVEC的细胞增殖过程 ,抑制细胞凋亡 ,HUVEC的过度增生及 HUVEC感染引起的炎性反应可能影响内皮细胞的功能 ,甚至破坏内皮细胞 ,导致血栓形成、脂质代谢紊乱 ,最终形成动脉粥样硬化。     

移植冠心病患者骨髓间充质干细胞治疗心肌梗死大鼠的机制探讨

目的:在大鼠心肌梗死模型上移植冠心病患者来源的骨髓间充质干细胞,探讨人骨髓间充质干细胞改善心肌梗死后心功能的作用机制。方法:抽取冠心病患者骨髓,用人骨髓间质细胞专用培养液进行体外培养、扩增、标记。结扎大鼠冠状动脉左前降支制作心肌梗死模型,2周后使用超声筛选出合格的动物模型。然后动物随机分为细胞移植组(n=14)和培养液注射组(对照组,n=13)。在移植前后对心脏的收缩、舒张功能和心室重构指标进行超声评价和对比,取材后进行病理学检查和分子生物学检测。结果:移植后进行超声评价,治疗组较对照组射血分数值和缩短分数值显著改善。免疫组化显示移植细胞在宿主体内可以向肌源性细胞分化。逆转录—聚合酶链反应(RT-PCR)显示细胞治疗组梗死区心肌Ⅰ型、Ⅲ型胶原基因表达水平明显高于对照组,而在非梗死区低于对照组。细胞治疗组梗死区间质细胞源因子、血管内皮生长因子、Bcl-2基因表达水平均明显高于对照组。结论:冠心病患者骨髓间质细胞移植后可以提高心肌梗死后心脏收缩功能,减轻左心室重塑。作用机制可能为新血管生成增加、细胞外基质改变、旁分泌、抗凋亡和心肌再生等共同作用的结果。     

骨髓间充质干细胞治疗缺血性脑损伤的研究进展

骨髓间充质干细胞在体外特定的条件下,可诱导分化成神经细胞,这一生物学特性为缺血性脑损伤的细胞和基因治疗带来了新的希望,为神经系统疾病的治疗和康复展示了美好的前景。文章对其研究现状及进展做了综述。     

Osteoblast-like Cell Proliferation,ALP Activity and Photocatalytic Activity on Sintered Anatase and Rutile Titanium Dioxide

This study aimed to create a biomaterial from titanium dioxide (TiO₂), which has been known to have photocatalytic and bone formation promoting effects. I expected that anatase titanium dioxide-based implants could promote bone augmentation and induce bone formation. Powdery anatase TiO₂ was compression molded and sintered at 700, 800, 900, and 1000 °C to prepare sintered compact samples. X-ray diffraction and scanning electron microscopy were used to observe the surface of these samples. Furthermore, mouse osteoblast-like cells (MC3T3-E1 cell line) were seeded on the samples sintered at different temperatures, and cell proliferation was observed to evaluate the cell proliferation of the samples. The sample sintered at 700 °C was composed of anatase TiO₂. The samples sintered at 800 °C and 900 °C were confirmed to consist of a mixture of anatase and rutile TiO₂ crystalline phases. Moreover, the sample sintered at 700 and 800 °C, which contained anatase TiO₂, showed remarkable photocatalytic activity. Those samples sintered at 1000 °C were transformed to the rutile TiO₂. The cell proliferation after 7–14-days culturing revealed that cells cultured on the 700 °C sample decreased in number immediately after initiation of culturing. The cells cultured on TiO₂ sintered at 900 °C markedly proliferated over time with an increase in the alkaline phosphatase activity, showing good MC3T3-E1 cell compatibility of the samples. The sample sintered at 1000 °C, which is rutile TiO₂, showed the highest increase.