Overexpression Of MiR-21 Promotes The Progress Of Gestational Trophoblastic Disease By Downregulating PDCD4

Objectives: micro RNAs are small noncoding RNAs implicated in post-transcriptional gene regulation.As one of miRNAs,miRNA-21(miR-21)is brought into focus by its role in the carcigenesis of many tumors.PDCD4(programmed cell death 4)and PTEN are two tumor suppressor genes.Their down-regulation or loss of function can promote the occurrence and development of tumor.The mechanism of gestational trophoblastic disease(GTD)is still not so clear.The expressions of miR-21、PDCD4 and PTEN were detected and the correlation of these three were investigated in this study.The downstream effectors and effects on trophoblastic cells were also explored.Our study aims to clarify the mechanism for development of GTD and progress of gestational trophoblastic neoplasm(GTN),to look for new molecular treatment target for chemoresistent GTN and new follow up biomarker for hydatidifrom mole(HM).Methods:1)The expression of miR-21 was detected in 20 cases of normal first trimester(1st)placentas and 16 cases of HM frozen tissues by quantitative real-time reverse-transcription polymerase chain reaction(q RT-PCR).2)The expression and distribution of miR-21 were detected in 23 cases of normal first trimester(1st)placentas and 42 cases of HM paraffin tissue sections by in situ hybridization(ISH)and fluorescent ISH(FISH).3)The expression of PDCD4 in 53 cases of 1st placentas,98 cases of HMs and 10 cases of invasive HMs were detected by immunohistochemistry(IHC).4)The m RNA and protein expressions of PDCD4 and PTEN in 10 cases of 1st placentas and HM frozen tissues were detected by q RT-PCR and Western blot respectively.5)The effect on choriocarcinoma JAR and JEG3 cells after transfected with miR-21 mimic/inhibitor were evaluated by CCK assay,transwell chamber and flowcytometric analyses.6)The luciferase reporter assay was performed to determine whether PDCD4 is the target gene of miR-21.The relationship was further confirmed by q RT-PCR and Western blot in choriocarcinoma cell lines.7)The q PCR-array technology was used to screen the PDCD4 targeted downstream effectors which are resposible for dysregulted cell function in choriocarcinoma cells.Result:1)The expression of miR-21 in HM tissues is significantly higher than in 1st placentas(P<0.05).The expression of miR-21 in choriocarcinoma cells is significantly higher than in primary cultured normal trophoblast cells(P<0.05).2)The expression of PDCD4 in HM tissues is significantly lower than in 1st placentas(P<0.05),nearly loss of expression in invasice HM.The expression of PDCD4 in choriocarcinoma cells is lower than in primary cultured normal trophoblast cells(P<0.05).3)PDCD4 and PTEN are negatively regulated by miR-21.4)Overexpression of miR-21 promoted the proliferation,invasion,migration and suppressed the apopotosis in choriocarcinoma cells whereas inhibition of miR-21 reversed those functional effects on choriocarcinoma cells.5)MMP3,MMP7 and their inhibitor TIMP1 were screened and verified as the downstream effectors of PDCD4 in choriocarcinoma cells.Conclusion:Overexpression of miR-21 in GTD endow trophoblastic cells with high aggressive phenotype by inhibiting the expression of PDCD4 and PTEN.PDCD4 effects the ability of MMPs,which eventually promotes the progress of GTN.The miR-21/PDCD4 or miR-21/PTEN axis could be served as molecular treatment target.The miR-21 could be used as biomarker for HM prognosis.