| Canine endometritis is a common obstetric disease in pet clinical and it serious threaten the health of pets. Caffeic acid is a phenolic acid compound and which has anti-inflammatory and anti-bacterial effect.In this article,through caffeic acid resistant the inflammation of canine endometrial epithelial cells(CEEC) induced by Lipopolysaccharide(LPS),to explore the anti-inflammatory effect of caffeic acid and to provide the theoretical basis for the new endometritis treatment drug explore and development.Our project includ the following exprements:This article obtained CEEC by enzyme digestion method; the effect of LPS on the viability of CEEC was tested by MTT, and the LPS-injured CEEC model was preliminarily established; the caffeic acid toxicity test was tested by MTT, and the safe dose rang was determined and three caffeic acid doses were choosed; the effect of different concentration of caffeic acid on the viability of LPS-induced CEEC was tested by MTT; the effect of different concentration of caffeic acid on the level of proinflammatory cytokine in LPS-induced CEEC’s culture media was detected by ELISA method; the level of nitric oxide in CEEC’s culture media was determined by Nitric Oxide Assay Kit; the effect of different dose of caffeic acid on the CEEC apoptosis induced by LPS was tested by Acridine Orange and Ethidium Bromide staining method.Following results are concluded:LPS significantly inhibited cell viability in a time-and dose-dependent manner. The inflammatiory CEEC model could be established by 50μg/m L LPS treated for 12 h. After CEEC was treated with various concentrations of caffeic acid for 12 h, compared with control group, the inhibited rate of 1ng/m L~200μg/m L caffeic acid groups had no significantly difference. After CEEC was treated with various concentrations of caffeic acid for 24 h, 1ng/m L~25μg/m L caffeic acid groups had no significantly difference. We choosed 25μg/m L, 50μg/m L, 100μg/m L in safe range as caffeic acid low, middle and high group.After CEEC was pretreated by middle, high caffeic acid for 3h, the cells viability was significant increased compared with model group and had no significant difference compared with control group. The level of NO, IL-6 and TNF-α in model group had significantly increased compared with control group, however the caffeic acid groups had significantly decreased. After AO/EB staining, under the fluorescence microscope, a large number of apoptotic cells in model group, and in the control group, most of the cells was living cells, and in caffeic acid group, the apoptotic cells is less than that of model group. |
