高效液相法检测内毒素对肺泡Ⅱ型上皮细胞能量负荷的影响

Determination of the effect of lipopolysaccharide on the energy charge of alveolar type Ⅱ epithelial cells of rat by HPLC

目的建立高校液相法(HPLC)检测肺泡Ⅱ型上皮细胞能量负荷的方法,并观察内毒素对肺泡Ⅱ型上皮细胞能量负荷的影响。方法大鼠肺泡Ⅱ型上皮细胞成功分离纯化后随机分为两组:①对照组(PBS);②LPS组(1μg·mL-1)。药物干预4h候后用HPLC法测定两组ATP、ADP、AMP含量,根据腺嘌呤核苷酸总量(TAN)计算公式:TAN=ATP+ADP+AMP,能荷(energy charge,EC)计算公式:EC=(ATP+0.5×ADP)/TAN,计算细胞能荷值。结果 LPS组较空白组明显增高(P<0.05)。结论高校液相法检测ATP、ADP、AMP具有操作简单、分离效果好、灵敏度高等特点。内毒素能破坏肺泡Ⅱ型上皮细胞代谢平衡,引起能量利用障碍,可能与内毒素性肺损伤相关。

OBJECTIVE To determine the effect of lipopolysaccharide on the energy charge of alveolar type Ⅱ epithelial cells of rat by HPLC.METHODS ATⅡepithelial cells were isolated and purified,and divided randomly into two groups：control group（PBS） and endotoxin（LPS,1μg·mL-1） group.After 4 hours of stimulation,ATP,ADP,AMP,total adenine nucleotides（TAN） and energy charge（EC） were measured by high performance liquid chromatography（HPLC）.RESULTS The EC of ATⅡepithelial cells was higher in LPS group（P0.01 vs control group）.CONCLUSION HPLC is a simple,fast,accurate method for determination of the energy charge of alveolar type Ⅱ epithelial cells of rat.The lipopolysaccharide can destroy the balance of metabolism,and leat to ineffective use of energy of ATⅡepithelial cells.This may be associated with LPS-induced lung edema.