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目的探讨可吸入颗粒物(PM10)所诱发的巨噬细胞毒性和前炎性细胞因子mRNA表达的改变与PM10中内毒素成分的关系。方法采用鲎试剂终点显色法测定未经处理的PM10(PM10+)和经130℃高热处理24 h的PM10(PM10-)中内毒素含量,将PM10+和PM10-以不同浓度(0、25、50、100、200、400μg/ml)分别染毒小鼠单核巨噬细胞系RAW264.7细胞24 h,采用MTT法测定细胞存活率,选取0、25、50、100、200μg/ml的PM10+和PM10-分别染毒RAW264.7细胞6、12、24 h,采用反转录-聚合酶链反应技术(RT-PCR)测定染毒各时相RAW264.7细胞白介素-6(IL-6)mRNA和肿瘤坏死因子-α(TNF-α)mRNA的表达水平。结果 PM10+和PM10-中内毒素含量分别为(58.7±6.4)EU/mg和(2.0±0.1)EU/mg。PM10+和PM10-对RAW264.7细胞的存活率影响不明显。染毒剂量在50μg/ml以上时,PM10+和PM10-均可使RAW264.7细胞IL-6 mRNA和TNF-αmRNA表达增加(P<0.05),随着染毒剂量的增加,IL-6 mRNA和TNF-αmRNA的表达均呈升高趋势,染毒6 h后,PM10-染毒组与PM10+染毒组相比,IL-6mRNA的表达减少(P<0.05)。结论 PM10对巨噬细胞的前炎性细胞因子mRNA表达的影响与其内毒素成分有关。
Objective To investigate the changes of macrophage cytotoxicity and pro-inflammatory cytokine mRNA expression induced by inhalable particulate matter (PM10) and the relationship of endotoxin components in PM10. Methods The endotoxin content in untreated PM10 (PM10 +) and PM10 (PM10-) treated with high heat treatment at 130 ℃ for 24 h was determined by the method of end-point colorimetry with 鲎 reagent. The concentrations of PM10 + and PM10- , 100, 200, 400μg / ml) were exposed to RAW264.7 cells of monocyte-macrophage cell line for 24 h respectively. The cell survival rate was determined by MTT assay. PM10 +, 0, 25, PM10-infected RAW264.7 cells were exposed to 6, 12 and 24 h respectively. The levels of interleukin-6 (IL-6) mRNA in RAW264.7 cells were determined by reverse transcription-polymerase chain reaction And tumor necrosis factor-α (TNF-α) mRNA expression levels. Results The endotoxin contents in PM10 + and PM10- were (58.7 ± 6.4) EU / mg and (2.0 ± 0.1) EU / mg, respectively. The effect of PM10 + and PM10- on the viability of RAW264.7 cells was not obvious. IL-6 mRNA and TNF-αmRNA expression in RAW264.7 cells increased with the dose of 50μg / ml or above (P <0.05). With the increase of the dose of IL-6 mRNA and The expression of TNF-αmRNA tended to increase. After 6 h exposure, the expression of IL-6 mRNA decreased in PM10-treated group compared with PM10 + -treated group (P <0.05). Conclusion The effect of PM10 on the mRNA expression of proinflammatory cytokines in macrophages is related to its endotoxin composition.