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目的 探讨严重烧伤早期血管内皮细胞 (EC)损伤机制 .方法 以培养脐静脉 EC为模型 ,应用扫描电镜技术 ,3H-Td R特异性释放实验 ,观察了终浓度为 2 0 0 m L· L- 1 的早期烧伤患者血清 (EBPS)介导 PMN与 EC相互作用后 ,EC及PMN的表面结构 ,PMN与 EC粘附特性及 EC3H- Td R特异释放率的变化 .结果 正常人血清 (NHS)调理的 PMN为园形、规则 ,EBPS调理的 PMN形态不规则 .NHS对照组 PMN与 EC连接松散 ,EC与 EC连接紧密 ,EBPS刺激 EC与 NHS调理的 PMN相互作用 ,EC与 EC连接出现微小间隙 ,EC表面突起增多 ,PMN形态不规则 ,主要粘附于 EC间隙 .NHS刺激的 EC与 EBPS调理的 PMN相互作用后 ,PMN偏平状锚于 EC表面 .EBPS刺激的 EC与 EBPS调理的 PMN相互作用 ,PMN形态不规则 ,表面微绒毛显著减少 ,EC从膜上分离脱落 ,EC3H- Td R特异性释放率与对照组比较有显著差异(P<0 .0 1) .结论 EBPS可激活 EC和 PMN,激活的 EC与PMN相互作用 ,可引起 EC损伤 ,EC损伤依赖 PMN的参与 ,EBPS可能不是作为一种直接的损伤因素而是作为一种细胞激活剂参与了烧伤早期 EC的损伤过程 相似文献
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脂质体介导质粒转染角朊细胞的影响因素 总被引:4,自引:2,他引:2
探讨利用脂质体介导真核表达质粒转染体外培养的人角朊细胞的最佳转染条件。体外分离培养正常人角朊细胞 ,培养至 6 0 %、70 %、80 %、90 %及 10 0 %融合时 ,应用不同浓度的LipofectAMINE包被真核表达质粒pCMV·SPORT β gal,分别转染 6、8、10、12及 2 4h。细胞经转染后再培养 48h ,行 β 半乳糖苷酶原位染色 ,镜下观察并计算阳性转染率。被转染的阳性细胞 ,可见质粒 β 半乳糖苷酶基因的良好表达 ;当细胞融合率为 80 %、90 % ,以12 5 μL/ 10 0 μL的LipofectAMINE包被 1 5 μg/ 10 0 μL的pCMV·SPORT β gal,转染时间为 8h的转染率最高 ,可分别达到 (31 35± 1 35 ) %、(32 32± 2 4) %。该实验说明LipofectAMINE可有效地介导真核表达质粒转染培养的人角朊细胞 ;转染率与细胞生长状态、脂质体包被质粒的浓度比例、及转染时间直接相关。 相似文献
105.
目的:建立一种简单高效的毛囊隆突细胞的培养方法. 方法: 将5~7 mo终止妊娠胎儿头皮的毛干中上部剪下后Ⅰ型胶原酶消化,采用成纤维细胞和毛囊隆突细胞对胰酶的不同敏感性进行纯化,流式细胞仪进行细胞周期分析,MTT法检测细胞克隆率,免疫组化检测细胞K19的表达情况. 结果: 消化法原代培养可以每小时获得100个左右毛囊隆突,经姨酶消化不同时间可以纯化毛囊隆突细胞,流式细胞仪检测第二代细胞G1期占88.20%,细胞克隆形成率为18.2%,K19表达呈阳性. 结论: 建立了一种简单高效的毛囊隆突细胞的分离、培养方法,并证明其具有某些干细胞的特征性表现. 相似文献
106.
复合皮真皮-表皮连接区结构再生与功能重建 总被引:4,自引:0,他引:4
目的:为了解复合皮(CS)真皮-表皮连接(DEJ)区结构再生及其对CS功能重建的作用机制。方法:用经胰蛋白酶处理的异体乳猪真皮与自体表皮重组CS,并以自体表皮为对照(C组),对移植术后2~124周的活检标本作光镜和透射电镜观察。结果:①CS组表皮和真皮生长快、成熟早,乳头层组织疏松有序,毛细血管增生迅速,20~24周时,乳头和皮钉及以致密板为中心的半桥粒、锚丝、锚纤维等DEJ区结构形成,以靠近毛细 相似文献
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Objective To reproduce a model of heat injured KC in vitro and explore its apoptosis rate of KC due to heat injury at different temperature. Methods Human KCs were cultured in vitro, and they were incubated at 37, 41, 43,45, 48, and 51 ℃ respectively for 10 mins in water bath. Trypan blue staining and Hoechst 33258 fluorescence staining were used respectively to determine necrosis and apoptosis of KC. Rates of apoptosis and necrosis of KC were analyzed quantitatively by flow cytometer. The prolifera-tion activity of KC after heat injury was detected by MTT test. Results The results of trypan blue stai-ning, Hoechst 33258 fluorescence staining, and flow cytometer demonstrated that number of apoptotic and necrotic KC increased gradually along with a rise of water bath temperature. The rates of apoptosis and necro-sis of KCwererespectively(12.3±3.2)% and (14.1±1.6)% at 45℃, (27.7±5.1)% and (58.0± 4.2)% at 48 ℃. Rate of KC necrosis reached up to (83.0±5.3)% at 51℃. Inhibition of KC growth reached a stationary phase when the injurious temperature reached 45 ℃ as observed with MTT test. Con-clusions Heat injury can induce apoptosis and growth inhibition of KC in vitro. Incubating KC at 45 ℃ for 10 mins is a good condition to reproduce a model of heat injured KC in vitro. This model may be used to study the biological character and apoptosis of KC after burn injury. 相似文献
110.
Objective To study the effect of insulin in different concentrations on secretion function of growth factors of adipose-derived stem cells (ADSCs). Methods ADSCs were isolated from human ab-dominal adipose tissue and cultured. The immunophenotype and adipose induced-differenciation were identi-fied, and the third generation cells were collected. The collected cells were assigned to 1 × 10-8, 1 × 10-7, 1 × 10-6 mol/L insulin groups according to the concentration of added insulin. When cells grew into 70% confluence in conventional medium, ADSCs were cultured further in serum-free DMEM containing insulin in different concentrations for 3 days. ADSCs cultured in medium without insulin were used as control group. Secretion amount of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) of ADSCs were determined by enzyme-linked immunosorbent assay. The effects of the supernatant fluid of ADSCs' nutrient solution on the proliferation and collagen synthesis of the cultured fibroblast were detected by MTT chromatometry and hydroxyproline chromatometry. Results The secretion amounts of VEGF and HGF of ADSCs in 1 × 10-8 and 1 × 10-7 mol/L insulin groups [ (471±41, 762±66 ng/L), (643±64, 930±67 ng/L) , respectively ] were significantly higher as compared with those in control group (286±47, 577±84 ng/L) ( P <0.05 orP <0.01). No change occurred in the secretion amount of VEGF and HGF of ADSCs in 1±10-6 mol/L insulin group ( P >0.05 ). The supernatant fluid of ADSCs' nutrient medium of 1 ± 10-8 ,1 ± 10-7 mol/L insulin groups showed obvious stimulative effect on the proliferation and collagen synthesis of fibroblasts, and it was most obvious in the 1 ± 10-7 mol/L group ( P < 0.05 or P < 0. 01 ). Conclusions Insulin in the concentrations of 1 ± 10-8 and 1 ± 10-7 mol/L can notably promote ADSCs' function of secreting VEGF and HGF. 相似文献