Effects And Mechanism Of Free Fatty Acids On Inhibition Of KLF4 Expression In Obesity Related Endometrial Carcinoma

Object:In this study,we cultured endometrial cancer cell Ishikawa in vitro,determined whether high-level palmitic acid(PA)can promote the proliferation,invasion and migration of endometrial cancer cells by inhibiting KLF4 expression,which explore its possible molecular mechanism.It provides a new theory and basis for the clinical search for molecular targets for the development and progression of obesity-related endometrial cancer.Methods:(1)We cultured endometrial cancer cell Ishikawa in vitro and stimulated cancer cells with 20 ?M and50 ?M PA.(2)Transfected with an expression plasmid or si RNA,after up and down regulating KLF4,down-regulating DNMT1 and DNMT3 b,q RT-PCR and Western Blot were used to detect m RNA and protein expression levels of DNMT1,DNMT3 b,KLF4,P53,Ki67 and MMP2 by using cell transfection and RNA interference techniques.(3)CCK-8,Transwell and scratch assays were used to examine the effects of Ishikawa cell proliferation,invasion and migration.(4)Statistical analysis of data was performed using SPSS 17.0 software.Data analysis was performed by t-test and rank sum test,which comparison between groups was performed by ANOVA analysis of variance.P<0.05 was considered statistically significant.Results:1.PA can affect the expression of KLF4 and related genes,and lead to changes in the biological behavior of endometrial cancer cells(1)20 ?M,50 ?M PA stimulated Ishikawa cells for 24 hours,the results of 20 ?M group showed,compared with the control group,the expression of KLF4 m RNA was significantly decreased(P<0.05).The expression of Ki67 and MMP2 m RNA was significantly increased(P<0.05).The results of the 50 ?M group showed that compared with the control group,the m RNA and protein expression levels of KLF4 and P53 were significantly decreased(P<0.05).The m RNA and protein expression levels of Ki67 and MMP2 were significantly increased(P<0.05).(2)20 ?M,50 ?M PA stimulated Ishikawa cells for 24 and 48 hours,compared with the control group,the invasion and migration ability of the cells were significantly increased(P<0.05).However,there was no significant effect on cell proliferation ability(P>0.05).2.KLF4 can affect the expression of downstream related genes and cause changes in the biological behavior of endometrial cancer cells(1)After up-regulated KLF4 for 24 h,the expression levels of Ki67 and MMP2 mRNA and protein were significantly lower than those in the control group(P<0.05).After up-regulation of KLF4 for 24 and 48 h,the proliferation,invasion and migration of Ishikawa cells were significantly lower than those of the control group(P<0.05).(2)After down-regulated KLF4 for 24 h,compared with the control group,the expression of P53 m RNA and protein was significantly decreased(P<0.05),which the expression levels of Ki67 and MMP2 mRNA and protein were significantly increased(P<0.05).After downregulating KLF4 for 24 and 48 h,compared with the control group,the invasion and migration ability of Ishikawa cells were significantly increased(P<0.05).3.PA inhibits endometrial cancer cell KLF4 and affects downstream gene expression,leading to changes in cell biological behavior(1)50 ?M PA stimulates endometrial cancer cell Ishikawa,while up-regulating KLF4 for 24 h,compared with the control group,the expression level of P53 m RNA was significantly increased(P<0.05),which the expression levels of Ki67 and MMP2 m RNA and protein were significantly decreased(P<0.05).(2)50 ?M PA stimulates endometrial cancer cell Ishikawa,while up-regulating KLF4 for 24 h,compared with the control group,the proliferation,invasion and migration ability of Ishikawa cells were significantly decreased(P<0.05).4.PA inhibits KLF4 expression by up-regulating methyltransferase(DNMT1/3b)(1)The m RNA and protein expression levels of DNMT1 and DNMT3 b in cancer tissues of patients with endometrial cancer were significantly higher than those in non-cancer individuals(P<0.05).(2)20 ?M,50 ?M PA stimulated Ishikawa cells for 24 hours,compared with the control group,the m RNA and protein expression levels of DNMT1 and DNMT3 b were significantly increased(P<0.05).(3)After down-regulating the expression of Ishikawa DNMT1/3b in endometrial cancer cells for 24 h,compared with the control group,the expression of KLF4 m RNA and protein was significantly increased(P<0.05).(4)50 ?M PA stimulated endometrial cancer cell Ishikawa,while down-regulating DNMT1/3b for 24 h,compared with the control group,the expression of KLF4 m RNA and protein was significantly increased(P<0.05).Conclusions:1.PA may inhibit the expression of downstream related oncogenes and tumor suppressor genes by inhibiting the expression of KLF4,which finally promote the proliferation,invasion and migration of endometrial cancer cells Ishikawa.2.PA may inhibit the expression of endometrial cancer cell Ishikawa KLF4 by promoting the expression of DNA methyltransferase DNMT1/3b.