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目的：探究羧甲司坦（carbocysteine, S-CMC）在气道炎症中对组蛋白去乙酰化酶2（histone deacetylase2, HDAC2）表达的调控作用和机制。方法：建立脂多糖（lipopolysaccharide，LPS）诱导大鼠肺泡巨噬细胞（NR8383）炎症模型、短期烟熏Sprague Dawely （SD）大鼠气道炎症模型，采用酶联免疫吸附测定法（enzyme-linked immunosorbent assay，ELISA）检测炎症因子白细胞介素6 （interleukin-6，IL-6）和白细胞介素8（interleukin-8，IL-8）的水平，蛋白免疫印迹（Western blotting）及免疫组化染色检测HDAC2 的表达。结果：与对照组相比，模型组NR8383 细胞中HDAC2的表达明显降低至对照组的0.47 ± 0.11倍，细胞上清IL-6、IL-8 水平明显升高，分别为157.6± 15.0 pg/mL、378.0± 17.9 pg/mL；模型组SD 大鼠肺组织中HDAC2 的表达明显降低到对照组的0.42 ± 0.12 倍，气道灌洗液（bronchoalveolar lavage fluid，BALF）中IL-6、IL-8 分别为162.2 ± 51.4 pg/mL、331.4 ± 62.7 pg/mL，炎症因子水平明显升高。而与模型组比较，经S-CMC处理后细胞中HDAC2 表达明显上调至对照组的1.23 ± 0.05 倍，细胞上清中IL-6 为 92.3 ± 4.3 pg/mL，IL-8 为300.7 ± 17.7 pg/mL，炎症因子水平降低；肺组织中HDAC2 为对照组的0.78 ± 0.10 倍，表达水平明显升高，BALF中IL-6、IL-8 水平分别为100.6 ± 32.7 pg/mL，185.0 ± 50.4 pg/mL（P<0.05）炎症因子明显降低。组蛋白去乙酰化酶抑制剂曲古抑霉素A（trichostatin，TSA）能够抑制NR8383 细胞中HDAC2 的表达至对照组的0.19 ± 0.06 倍，增加IL-6（197.0 ± 42.6 pg/mL）、IL-8（567.0 ± 97.4 pg/mL）水平，该作用可以被S-CMC 所逆转（P <0.05）。另外，加入巯基供体二硫苏糖醇（dithiothreitol，DTT）可增强S-CMC 上调HDAC2 表达，降低IL-6、IL-8 的作用，而巯基耗竭剂丁硫氨酸亚砜亚胺（buthionine-sulfoximine，BSO）可减弱S-CMC 的作用（P <0.05）。进一步表明S-CMC 调控HDAC2 的过程与巯基相关。结论： S-CMC 可通过巯基上调HDAC2 的表达抑制气道炎症。

英文摘要:

Objective:To investigate the effects and mechanism of carbocysteine (S-CMC) on regulating airway inflammation via histone deacetylase2 (HDAC2).Methods:NR8383 cells stimulated by lipopolysaccharide (LPS) were used as the cell inflammatory model and Sprague Dawely (SD) rats exposed to cigarette smoking for 8 weeks were used as the animal model of airway inflammation. The levels of interleukin-6 (IL-6) and interleukin-8 (IL-8) were determined by enzyme-linked immunosorbent assay (ELISA). The expression of HDAC2 was examined by western blotting and immunohistochemistry.Results:HDAC2 expression was significantly decreased both in the cellular (0.47 ± 0.11 fold) and animal (0.42 ± 0.12 fold) models accompanied with increasing IL-6 (157.6 ± 15.0 pg/mL for NR8383 cells, 162.2 ± 51.4 pg/mL for BALF) and IL-8 (378.0 ± 17.9 pg/mL for NR8383 cells, 331.4 ± 62.7 pg/mL for BALF) levels compared with the control group. S-CMC can increase HDAC2 expression (to 1.23 ± 0.05 fold for NR8383 cells, 0.78 ± 0.10 fold for lung tissue compared with the control group) associated with decreasing levels of IL-6 (92.3 ± 4.3 pg/mL for NR8383 cells, 100.6 ± 32.7 pg/mL for BALF) and IL-8 (300.7 ± 17.7 pg/mL for NR8383 cells, 185.0 ± 50.4 pg/mL for BALF) (P<0.05). The decreased expression of HDAC2 (0.19 ± 0.06 fold) and increased IL-6 (197.0 ± 42.6 pg/mL), IL-8 (567.0 ± 97.4 pg/mL) levels with treatment of the histone deacetylase inhibitor trichostatin (TSA) were attenuated by S-CMC, significantly. The ability of S-CMC in increasing HDAC2 expression associated with decreasing levels of IL-6 and IL-8 was strengthened by dithiothreitol (DTT), a thiol-based reducing agent, while was attenuated by buthionine sulfoximine (BSO), a specific inhibitor of GSH synthesis.Conclusion:Our finding implied that S-CMC ameliorated the airway inflammation through up-regulating HDAC2 by affecting thiol levels.

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