磷酸二酯酶在金黄色葡萄球菌性乳腺炎中的表达分析

Expression analysis of phosphodiesterases in Staphylococcus aureus mastitis

为了探究磷酸二酯酶(PDEs)家族成员表达与金黄色葡萄球菌性乳腺炎的关系,试验采用PCR技术检测了PDE家族除PDE6、PDE10和PDE11基因外的各亚型在奶牛乳腺上皮细胞(bMECs)、小鼠乳腺上皮细胞(mMECs)及小鼠乳腺(mMG)中的表达情况,实时荧光定量PCR技术检测了金黄色葡萄球菌刺激下这些PDE家族亚型的表达差异及PDE4抑制剂RO-20-1724对金黄色葡萄球菌诱导的炎症因子表达的影响。结果表明:在bMECs中,PDE3B、PDE5A、PDE8B、PDE9A基因呈高表达;在mMECs中,PDE2A基因呈高表达;而在mMG中,PDE2A、PDE3A基因呈高表达。金黄色葡萄球菌刺激8 h后bMECs中PDE1C、PDE4A、PDE4B、PDE4C、PDE4D、PDE5A、PDE8A基因的相对表达量显著或极显著升高(P<0.05或P<0.01),而PDE1B、PDE7A、PDE7B基因相对表达量显著或极显著降低(P<0.05或P<0.01);mMECs中PDE1B、PDE1C、PDE2A、PDE3A、PDE4A、PDE4C、PDE4D、PDE5A基因相对表达量极显著升高(P<0.01),而PDE7A基因相对表达量极显著降低(P<0.01);mMG中PDE1B、PDE3A、PDE4C、PDE4D、PDE5A、PDE8A、PDE8B基因相对表达量显著或极显著升高(P<0.05或P<0.01),而PDE1C、PDE4B、PDE7B基因相对表达量极显著降低(P<0.01)。加入PDE4抑制剂RO-20-1724孵育8 h后可极显著降低金黄色葡萄球菌诱导的mMECs中IL-1β基因的相对表达量。说明乳腺上皮细胞中PDE家族的一些亚型表达存在种属差异,金黄色葡萄球菌感染后可引起不同PDE亚型表达发生改变。

To investigate the relationship between phosphodiesterase(PDEs)expression and Staphylococcus aureus mastitis,the expression of all subtypes of PDE family except PDE6,PDE10 and PDE11 in cow mammary epithelial cells(bMECs),mouse mammary epithelial cells(mMECs)and mouse mammary gland(mMG)was detected by PCR.Real-time fluorescence quantitative PCR was used to detect the expression difference of these PDE subtypes under the stimulation of Staphylococcus aureus and the effect of PDE4 inhibitor RO-20-1724 on the expression of inflammatory factors induced by Staphylococcus aureus.The results showed that PDE3B,5A,8B and 9A were highly expressed in bMECs,and PDE2A was highly expressed in mMECs,and PDE2A and 3A were highly expressed in mMG.The relative expression levels of PDE1C,PDE4A,PDE4B,PDE4C,PDE4D,PDE5A and PDE8A genes in bMECs were significantly increased after 8 h of Staphylococcus aureus stimulation(P<0.05 or P<0.01).The relative expression levels of PDE1B gene,7A gene and 7B gene were significantly decreased(P<0.05 or P<0.01).The relative expression levels of PDE1B,PDE1C,PDE2A,PDE3A,PDE4A,PDE4C,PDE4D and PDE5A in mMECs were significantly increased(P<0.01),while the relative expression levels of PDE7A were significantly decreased(P<0.01).The relative expression levels of PDE1B,PDE3A,PDE4C,PDE4D,PDE5A,PDE8A and PDE8B in mMG were significantly increased(P<0.05 or P<0.01),while the relative expression levels of PDE1C,PDE4B and PDE7B were significantly decreased(P<0.01).Incubation with RO-20-1724 for 8 h significantly decreased the relative expression of mMECs IL-1βgene induced by Staphylococcus aureus.These results indicated that there were species differences in the expression of some subtypes of PDE family in mammary epithelial cells,and Staphylococcus aureus infection could cause changes in the expression of different PDE subtypes.