目的：探究氯喹（chloroquine,CQ）对脂多糖（lipopolysaccharide,LPS）所致Ⅱ型肺泡（alveolar type Ⅱ, AT2）细胞株损伤的影响。方法：用不同浓度LPS和CQ分别单独处理AT2细胞,用细胞计数试剂盒-8（cell counting kit-8,CCK-8）检测细胞活力以筛选LPS和CQ的作用浓度。根据细胞处理情况将实验分为对照组（不做任何处理的细胞）、LPS组、CQ组和CQ+LPS组4组。用蛋白质印迹法检测各组裂解的胱天蛋白酶3、Bcl-2、Bax的表达和Akt的磷酸化水平。之后增加磷脂酰肌醇-3-激酶（phosphatidylinositol-3-kinase, PI3K）/Akt信号通路抑制剂LY294002与LPS和CQ共同处理细胞,即LPS+CQ+LY294002组,通过蛋白质印迹法检测裂解的胱天蛋白酶3、Bcl-2和Bax的蛋白表达。结果：用终浓度为1、10 和50 mg/L的LPS作用24 h,当LPS浓度为50 mg/L时,AT2活力下降显著（P<0.05),故选择该浓度的LPS构建脓毒症细胞损伤模型。用终浓度0.5 μmol/L和5 μmol/L CQ作用4 h时,AT2细胞活力无明显改变（P>0.05),当CQ浓度达10 μmol/L,AT2细胞活力开始下降,浓度越高,细胞活力下降越明显,故选择5 μmol/L的CQ作为治疗浓度。用50 mg/L的LPS刺激后,细胞内裂解的胱天蛋白酶3表达增高,Bcl-2/Bax比值及Akt蛋白磷酸化减少;在加入5 μmol/L的CQ后,裂解的胱天蛋白酶3含量下降,Bcl-2/Bax比值和Akt蛋白磷酸化显著增加;当加入LY294002后,CQ的作用被消除,Bcl-2/Bax比值减少,裂解的胱天蛋白酶3表达增高。结论：5 μmol/L的CQ可以通过激活PI3K/Akt信号通路,抑制细胞线粒体凋亡,减轻LPS诱导的AT2细胞损伤。

Objective To explore the effect of chloroquine (CQ) on lipopolysaccharide (LPS)-induced damage to alveolar type Ⅱ(AT2) cell. Methods Different concentrations of LPS and CQ were used to treat AT2 cells separately, and cell counting kit-8(CCK-8) was used to detect cell viability to screen the concentration of LPS and CQ. After that, the AT2 cells were divided into 4 groups based on different treatment, including control (without treatment), LPS, CQ and LPS+CQ groups. The expression of cleaved caspase 3, Bcl-2, Bax and the phosphorylation level of Akt were detected by Western blotting after each group of cells were processed. After that, LY294002, the signaling pathway inhibitor of phosphatidylinositol-3-kinase(PI3K)/Akt, was added in the cells which were treated by LPS and CQ successively(LPS+CQ+LY294002 group), and the protein expression of cleaved caspase 3, Bcl-2 and Bax was detected by Western blotting in this group. Results The LPS in the concentration of 1, 10 and 50 mg/L was kept for 24 h. The activity of AT2 decreased significantly (P<0.05) when the LPS concentration was increased to 50 mg/L, so LPS in this concentration was used to stimulate AT2. AT2 cell viability did not change significantly (P>0.05) when they were treated with 0.5 μmol/L and 5 μmol/L CQ for 4 h. However, AT2 cell viability began to decrease and showed a concentration-dependent manner when the CQ concentration increased to 10 μmol/L. According to the cell response to CQ in different concentration, 5 μmol/L CQ was chosen to treat the cells. After stimulation with 50 mg/L LPS, the expression of cleaved caspase 3 in the cell increased, and the Bcl-2/Bax ratio and the phosphorylation of Akt decreased. After adding 5 μmol/L CQ, the cleaved caspase 3 dropped, and Bcl-2/Bax ratio and Akt protein phosphorylation rise significantly. After the PI3K inhibitor LY294002 was added, the effect of CQ was eliminated, which showed the Bcl-2/Bax ratio decreased, and the expression of cleaved caspase 3 increased. Conclusions CQ(5 μmol/L) could inhibit cell mitochondrial apoptosis by activating the PI3K/Akt signaling pathway, and alleviate the damage of AT2 cells induced by LPS.