Effects Of Estrogen And Estrogen Receptor β On Colorectal Tumorigenesis And The Biological Behaviors Of Colorectal Cancer

Epidemical studies show that postmenopausal women who had never used hormonereplacement therapy(HRT) were at increased risk of colorectal cancer(CRC) comparedwith premenopausal women.On the other hand,data from prospective randomized trialsshowed that HRT reduced the risk of CRC in postmenopausal women greatly.The effectsof estrogens are mediated by estrogen receptor(ER).Existed researches indicated that therewere two types of estrogen receptors,estrogen receptor a(ERa) estrogen receptorβ(ERβ).The two estrogen receptors have been found to exert some different biological effects.Andthe ratio of ERa and ERβdiffers with tissue type.ERβis the predominant ER in the colonicepithelium,while the expression of ERa is very low.Many clinical studies revealed that theexpression of ERβwas obviously reduced in colon cancer compared with normal colon.The researches involved of breast cancer or prostate cancer indicated that ERβcould inhibitthe growth and invasion of cancer cells efficiently.These findings suggest that estrogen andestrogen receptorβplayed a protective role against the tumorigenesis of CRC.The presentstudy investiged how and by which mechanism estrogen and ERβinfluenced colorectaltumorigenesis and the biological behaviors of colorectal cancer in vivo and in vitro. PartⅠEffects of ovariectomy on colorectal tumorigenesis and microsatelliteinstability(MSI) in rat colorectal tumors induced by 1,2-dimethylhydrazineObjectives To study the effects of ovariectomy on colorectal tumorigenesis andmicrosatellite instability(MSI) in rat colorectal tumors induced by 1,2-dimethylhydrazine,to investigate the role of MSI in tumorigenesis of postmenopausal colorectal cancer.Methods Forty female Wistar rats were randomly divided into two groups:Ovariectomized(OVX) group and Sham-ovariectomized(Sham-OVX) group.All rats wereinjected intraperitoneally with 1,2-dimethylhydrazine(DMH)(20mg/kg b.w) once a weekfor 20 weeks.Ten weeks after the final DMH injection,all the rats were sacrificed tocollect tumors.Microsatellite instability of six microsatellite loci was detected usingfluorescent PCR followed by fragment analysis on automatic DNA sequencer withGeneScan 3.7 software.Results The tumor multiplicity in the OVX group was significantlyhigher than that in the Sham-OVX group(3.6±1.4 vs 2.4±1.6,P＜0.05).The incidence ofMSI-positive tumors in OVX group was higher than that in Sham-OVX group(32.1% vs10.8%,P＜0.05).The incidence of tumors showing MSI at multiple loci in OVX group wasalso higher than that in Sham-OVX group(18.9% vs 2.7%,P＜0.05).ConclusionOvariectomy increased tumor formation and the frequency of MSI in DMH-induced colontumors.It implied that MSI played an important role in the colorectal tumorigenesis ofpostmenopausal women.Keywords:Microsatellite instability;Colorectal cancer;Postmenopausal;1,2-dimethylhydrazinePartⅡEffects of estrogen and estrogen receptorβon proliferation and apoptosis ofcolon cancer cellsObjectives To evaluate the effects of estrogen and ERβgene over-expression onproliferation and apoptosis of colon cancer cells,and to investigate the mechanismsinvolved in these effects. Methods Liposome-mediated gene transfection method was used to transfer ERβgeneinto SW480 cells.The positive clone cells(SW480-C1- ERβ) were selected by G418 andconfirmed by RT-PCR and Western Blot.Normal SW480 cells and emptyplasmid-transfected SW480 cells(SW480-pEGFP-C1) were set as control.Under theconditions with or without estrogen,proliferation of these cells was measured by MTT,andapoptosis was assessed by Flow Cytometer(FCM).The expression of Survivin and BaxmRNA were quantitated by Real-time RT-PCR.Results The ERB mRNA and protein levels in SW480 cells and SW480-pEGFP-C1cells were low,while the ERβmRNA and protein levels in SW480-C1- ERβcells increasedmarkedly.Under the condition with or without the treatment of estrogen,proliferation andapoptosis of SW480-C1-ERβcells were significantly lower and higher than for SW480cells(or SW480-pEGFP-C1 cells),respectively;while the survivin mRNA and Bax mRNAlevels in SW480-C1-ERβcells were significantly lower and higher than that in SW480cells(or SW480-pEGFP-C1 cells),respectively.For SW480-C1-ERβcells,proliferationand apoptosis of estrogen-treated cells were significantly lower and higher than fornon-estrogen-treated cells,respectively;while the survivin mRNA and Bax mRNA levelsin estrogen-treated cells were significantly lower and higher than that innon-estrogen-treated cells,respectively.For SW480 cells(or SW480-pEGFP-C1 cells);there was no significant difference between proliferation,apoptosis and the expression ofsurvivin and Bax mRNA in estrogen-treated cells and non-estrogen-treated cells.Conclusion Over-expression of ERβcan inhibit proliferation and induce apoptosis ofcolon cells both in a ligand-independent manner and ligand-dependent manner,and theseeffects might be associated with modulation of survivin and Bax. PartⅢEffects of estrogen and estrogen receptorβon invasion and metastasis ofcolon cancer cellsObjectives To evaluate the effects of estrogen and ERβgene over-expression oninvasion of colon cancer cells,the expression of E-cadherin and the expression of CXCR4.Methods Under the conditions with or without estrogen,the invasion capability ofSW480 cells,SW480-pEGFP-C1 cells and SW480-C1-ERβcells was examined using atranswell chamber assay.E-cadherin and CXCR4 expression of these cells was examinedby Real-time RT-PCR and Western Blot.Results Under the conditions without estrogen,the invasion capability and CXCR4expression of SW480-C1-ERβcells decreased significantly compared with SW480 cells(orSW480-pEGFP-C1 cells).Under the conditions with estrogen,the invasion capability andE-cadherin expression of SW480-C1-ERβcells decreased and increased significantlycompared with SW480 cells(or SW480-pEGFP-C1 cells),respectively.ForSW480-C1-ERβcells,the invasion capability and E-cadherin expression ofestrogen-treated cells were significantly lower and higher than for non-estrogen-treatedcells,respectively;while CXCR4 expression did not change significantly.For SW480 cells(or SW480-pEGFP-C1 cells);there was no significant difference between the invasioncapability and the expression of E-cadherin and CXCR4 in estrogen-treated cells andnon-estrogen-treated cells.Conclusion Over-expression of ERβcan inhibit the invasion capability of colon cellsboth in a ligand-independent manner and ligand-dependent cancer,and these effects mightbe associated with modulation of E-cadherin and CXCR4.