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目的观察人羊膜匀浆上清液(human amniotic homogenate supernatant,hAHS)对脂多糖(lipopolysaccharide,LPS)致伤条件下大鼠肺泡Ⅱ型上皮细胞(alveolar typeⅡepithelial cell,AT-Ⅱ)增殖及肿瘤坏死因子α(tumor necrosis factor alpha,TNF-α)和白细胞介素1β(interleukin1 beta,IL-1β)分泌的影响。方法 1取健康胎盘来源的新鲜人羊膜制备hAHS,配制含hAHS体积分数不同的5组培养基(对照组、10%hAHS组、20%hAHS组、40%hAHS组、80%hAHS组)对大鼠AT-Ⅱ行分组培养,四甲基偶氮唑盐(microculture tetrazolium,MTT)法测不同时间点OD630值。2对对照组、LPS致伤组、40%hAHS干预组大鼠AT-Ⅱ行分组培养,MTT法和酶联免疫吸附剂测定(enzyme-linked immunosorbent assay,ELISA)法测不同时间点OD630值和培养液中大鼠TNF-α和IL-1β含量。结果 1对数期内前4组间各时间点比较,hAHS体积分数相对较高组别的OD630值也相对较高(即40%hAHS组>20%hAHS组>10%hAHS组>对照组),且第3~5天均有统计学差异(P<0.05),而hAHS含量更高的80%hAHS组,各时间点均低于40%hAHS组,且第3~5天有统计学差异(P<0.05)。2 hAHS干预组在24 h后各时间点的OD630值不仅均高于LPS致伤组,也均高于对照组,且均有统计学差异(P<0.05)。与此同时,hAHS干预组各时间点大鼠TNF-α和IL-1β含量均低于LPS致伤组,且二者均分别于前6 h各时间点组间差异有统计学意义(P<0.05)。结论 hAHS可显著促进体外正常培养条件下AT-Ⅱ的增殖,且对LPS致伤条件下AT-Ⅱ的增殖和TNF-α、IL-1β等促炎因子分泌具有保护和抑制作用。
Objective To observe the effects of human amniotic homogenate supernatant (hAHS) on proliferation of alveolar type Ⅱ epithelial cell (AT-Ⅱ) and tumor necrosis factor-α (TNF-α) induced by lipopolysaccharide (LPS) α (tumor necrosis factor alpha, TNF-α) and interleukin 1β (IL-1β) secretion. Method 1 Take fresh human amniotic membrane derived from healthy placenta to prepare hAHS, prepare 5 groups of medium with different hAHS volume (control group, 10% hAHS group, 20% hAHS group, 40% hAHS group, 80% hAHS group) The rats in AT-Ⅱ group were cultured in groups and the OD630 values at different time points were measured by MTT method. (2) Grouping of AT-II in control group, LPS-injured group and 40% hAHS intervention group, MTT assay and enzyme-linked immunosorbent assay (ELISA) The content of TNF-α and IL-1β in the culture medium of rats. Results In the logarithmic phase, the OD630 values of the groups with relatively high hAHS volume were also relatively high (ie, 40% hAHS group> 20% hAHS group> 10% hAHS group> control group) (P <0.05), while the hAHS group with higher hAHS content was lower than the 40% hAHS group at each time point, and there was a statistically significant difference between the 3rd and 5th days (P <0.05). The OD630 values of 2 hAHS intervention group at 24 h were not only higher than that of LPS injury group, but also higher than that of control group (P <0.05). Meanwhile, the levels of TNF-α and IL-1β in rats in hAHS intervention group were lower than those in LPS injury group at each time point, and the differences were statistically significant (P < 0.05). Conclusions hAHS can significantly promote the proliferation of AT-Ⅱ under normal culture conditions and protect and inhibit the proliferation of AT-Ⅱ and the secretion of proinflammatory cytokines such as TNF-α and IL-1β under LPS injury.