The Apoptosis Role Of TRAIL And Its Receptors In The Cervical Cancer Cell

Objective: Resently,the incidence of cervical cancer was higher in youngwomen than before. Despite intensive multimodality treatment includingsurgical resection followed by chemotherapy and radiotherapy were important incervical cancer,the effect of chemotherapy and radiotherapy remained poorbecause of intrinsic or acquired drug resistance,and the toxin effect in normalcell,These made the patient stop treating. In addition, the treatment influencednormal physical functions, The life quality of the female was decreased, and themortality of cervical cancer remained high. The etiopathogenisis of cervicalcancer is clear ,the role of human papilloma viruses were major in thedevelopment of cervical cancer.many vaccines were developed in immunologyaccording to the human papilloma viruses types,and entered the clinicalexperiment,the effects were satisfied,but the types of the human papillomaviruses became more and more ,the application of the vaccines was limited inpreventing and treating the cervical cancer.Resently TNF-related apoptosis-inducing ligand(TRAIL) was identified, this protein had beared exciting promiseas a potential cancer therapeutic agent. the protein is innocuous in normalcell.TRAIL initiates apoptosis of tumor cell from different organism uponbinding to its cognate death recepors(DR4,DR5),The other two receptors appearto act as "decoys",decoy receptor 1 (DcR1) and decoy receptor 2(DcR2).DcR1and DcR2 lack the functional cytoplasmic death domain .Both receptors aretherefore incapable of transmitting an apoptosis signal. Overexpression of thesereceptos protect cells from apoptosis induced by TRAIL.These characteristicshave attracted more attention.After many research , people found one possiblenew strategy in the treatment of cancer is to combine chemotherapeutic agentswith TRAIL,this way could trigger apoptosis in different levels ,therebyenhancing apoptosis induction and resulting in a better outcome.So we wereinterested in investigating the synergistic role of apoptosis by combining withTRAIL and cisplatin.The experient collect specimens from cervicitis ,cervical intraepithelialneopasia (CIN)grade Ⅰ,gradeⅡandⅢ,squamous cell cervical cancer.TRAILand its receptors were detected .In this study,we cultured human cervical cancerHeLa cells in vitro and examined the apoptosis,proliferation of HeLa cells. Andexamined mRNA expression of DR4,DR5 of the cell line after cispatin treatedHela cells for 24 hours. In order to investigate the association between TRAILand cervical lesions ,and the apoptosis role of TRAIL,and to investigatesynergistic mechanisms of TRAIL in combination with cisplatin for clinicalapplication.Material and Methods: In the experient,immunohistochemical assayswere adopted to detect the levels of TRAIL,DR4 and DcR1 in cervicitis,cervicalintraepithelial neopasia (CIN)grade Ⅰ,gradeⅡandⅢ,squamous cell cervicalcancer.A human cervical cancer cell line HeLa was cultured in vitro and TRAILwas divided into 50ng/ml,100ng/ml,200ng/ml at concentration and 100ng/mlTRAIL combined with 10 uM cisplatin . (1) The viability and cell growth curvewere observed by MTT assay .(2) Apoptosis rate was investigated by flowcytometry (FCM).(3) The mRNA expression of DR4,DR5 were measured byreserse transcription-polymerase chain reaction (RT-PCR).Results:(1)The expression of TRAIL was decreased gradually following thecervical lesion ,the expression of TRAIL in cervical cancer was the lowest ,butno remarkable difference in different lesions. The expression of DR4 wasdecreased in cervical cancer than in cervicitis,CINⅠ, CINⅡandⅢ,thedifference was remarkable,no difference in cervicitis,CINⅠ, CINⅡandⅢ, Theexpression of DcR1 was decreased gradually following the cervical lesion, Theexpression of DcR1 is lowest in cervical cancer,there was remarkable differencebetween cervicitis and cervical cancer,but no difference in cervicitis,CINⅠ,CINⅡandⅢ.The expression of DR4 in well differentiated and moderatelydifferentiated cervical cancer was significant higher than poorly differentiatedcervical cancer, the difference was remarkable,no differences in other twogroups.(2)The cell growth curve showed TRAIL at various concentration inhibitedthe growth of HeLa in a time –and dose-dependent manner.combination with100ng/mlTRAIL and 10uM cisplatin showed lower viability than 100ng/mlTRAIL or 10uM cisplatin alone.(3)The apoptosis rate were detected by flow cytometry , The HeLa weretreated with different concentration (50ng/ml, 100ng/ml, 200ng/ml) and thecombination of 100ng/ml TRAIL and 10uM cisplatin for 24 hours, Theapoptosis rate represented a inceased trend , The apoptosis rate was higher incombination of 100ng/mlTRAIL and 10uM cisplatin than 100ng/mlTRAIL or10uM cisplatin alone.(4) The expression of DR4,DR5mRNA in HeLa cells were examined byRT-PCR after 10uM Cisplatin treatment for 24 hours,cisplatin upregulated theexpression of DR4,DR5 and were significant compared with the control groups.Conclusion:(1) The expression of TRAIL was gradually decreased following degree ofcervical lesions,but there was not association between the expression of TRAILand cervical lesions;DR4 was associated with the initiation and prognosis ofcervical cancer , the increasing expression of DcR1 in cervicitis showed theprotection role in normal cells from apoptosis induced by TRAIL .the resultsconfirmed the important role of death receptors and their ligands in apoptosis.(2)The research in vitro showed TRAIL can inhibited the growth of humancervical cancer HeLa cell in time-and dose-dependent manner,there was lowerviability and higher apoptosis in combination with TRAIL and cisplatin thanTRAIL or cisplatin alone.There was a synergistic role in combination withTRAIL and cisplatin.(3) The cisplatin played a synergistic role by upregulating the expression ofthe death receptors .