Expression Of Fragile Histidine Triad(FHIT) And Cell Proliferation Associated Nuclear Antigen Ki-67 In Endometriosis

【Backgroud】In 1885,Von Recklinghausen raised and named endometriosis(EM) first time. EM is a condition where tissue similar to the lining of the uterus(the endometrial stroma and glands,which should only be located inside the uterus) is found elsewhere in the body.Now EM has become a kind of common and frequently-occurring desease in reproductive-age women.It has year-by-year-increased morbidity and high recurrence rate after medication.Though EM is benign,but it has some similarity with malignant tumor,such as infiltrative growth,implantation,distant metastasis. The difference is that EM ectopic endometrium maintains normal and controllable proliferation and growth.The cause of endometriosis is unknown.One theory is that the endometrial tissue is deposited in unusual locations by the backing up of menstrual flow into the fallopian tubes and the pelvic and abdominal cavity during menstruation(termed retrograde menstruation).The cause of retrograde menstruation is not clearly understood.But retrograde menstruation is not the entire story.Many women have retrograde menstruation in varying degrees,yet not all of them develop endometriosis.Another possibility is that areas lining the pelvic organs possess primitive cells that are able to grow into other forms of tissue,such as endometrial cells.It is also likely that direct transfer of endometrial tissues during surgery may be responsible for the endometriosis implants sometimes seen in surgical scars(for example,episiotomy or Cesarean section scars).Transfer of endometrial cells via the bloodstream or lymphatic system is the most likely explanation for the rare cases of endometriosis that develop in the brain and other organs distant from the pelvis. Finally,some studies have shown alternations in the immune response in women with endometriosis,which may affect the body's natural ability to recognize and destroy any misdirected growth of endometrial tissue.Recently,domestic scholar proposed "eutopic endometrium determinism": pathological changes of EM eutopic endometrium is the key factor of EM cause and determines if the endometrial tissue in menstrual blood can implant in unusual locations.However,there isn't any theory can explain the cause of all kinds of EMs.So the common viewpoint is that EM is maybe caused by multiple mechanism and multiple factors together.【Objective】Our research is to observe the expression of Fhit and ki-67 in the tissue of normal endometria,in ectopic and eutopic endometria of endometriosis,as well their expression defference in different phase of menstrual cycle and in different r-AFS phase,to study the correlation between the expression of Fhit and endometrial cell proliferation in the occurence and progress of endometriosis,so as to explore the pathogenesis of endometriosis and to provided the new path in diagnose and therapies of endometriosis.【Materials and Methods】1 To select 48 cases of ovarian endometriosis(OEM group) who were operated at the Department of Obstetrics and Gynecology at the Nanfang Hospital of Southern Medical University from 2006.3～2007.9,including 25 cased in the proliferative phase and 23 cases in the secretory phase;13 cases in r-AFS stageⅠ-Ⅱ,21 cases in r-AFS stageⅢand 14 cases in r-AFS stage -Ⅳ.22 cases adenomyosis(AM group) were selected in the same period,including 15 cases in the proliferative phase and 7 cases in the secretory phase.Ectopic and eutopic endometrial tissues of OEM group and AM group were obtained to detect.20 cases hysteromyoma were selected as control group at the same time.Including 10 cases in the proliferative phase and 10 cases in the secretory phase.Their normal endometria were obtained to detect.The mean age of all the patients is 32.6±5.7(range 23-48).All of them are without other gynecologic diseases and had not received either chemotherapy or radiation therapy or hormones or IUD 6 months before surgery.2 The pathological change and menstrual cycle phase of all the specimen were observed through HE staining.3 The expression of Fhit and ki-67 was detected by immunohistochemistrical SP staining method.①Fhit expression was quantificated with mean optical density(MOD) though image analysis software- image-pro plus v6.0. MOD=integerated optical density/scanning area;②ki-67 expression was indicated with proliferation index(PI).PI=positive cells number/total cells number×100%.4 Statistic software SPSS13.0 was used to treat the data.T test and F test was used to compare the difference between or among different groups.Size of test is P＜0.05.【Results】1 Fhit expression1.1 Location of Fhit expression:Fhit mainly expressed in the endochylema and membrane of eutopic or ectopic endometrium glandual epithelial.The stroma of eutopic endometrium had expression too,but the stroma of EM(OEM+AM) ectopic endometrium hadn't expression.1.2 Difference of Fhit expression in different menstrual phase:Fhit expression of the eutopic and ectopic endometrium in control group and EM group showed no difference between proliferative phase and secretory phase in the menstrual cycle(P＞0.05),which indicated Fhit expression didn't affect menstrual cycle.1.3 Difference of Fhit expression in eutopic endometrium of control group,OEM group and AM group:In proliferative phase,the difference of Fhit expression among the three groups were such as follow:glandual epithelial-0.311±0.030 vs 0.295±0.016, 0.287±0.017(P＜0.05),stroma-0.037±0.006 vs 0.032±0.007,0.031±0.006(P＞0.05).In secretory phase,glandual epithelial-0.326±0.031 vs 0.301±0.020, 0.298±0.018(P＜0.05),stroma-0.040±0.006 vs 0.035±0.003,0.034±0.004(P＜0.05). Fhit expression of EM group was lower than that of control group,which showed eutopic endometrium of EM group had different bionomics with control group.1.4 Difference of Fhit expression between eutopic and ectopic endometrium:In proliferative phase,the difference of Fhit expression between OEM eutopic endometrium and ectopic endometrium was significant-0.295±0.016 vs 0.192±0.020(t=20.719,P=0.000),In secretory phase,the difference of Fhit expression between OEM eutopic endometrium and ectopic endometrium was significant too-0.301±0.020 vs 0.196±0.018(t=19.010,P=0.000).In proliferative phase,the difference of Fhit expression between AM eutopic endometrium and ectopic endometrium was significant-0.287±0.017 vs 0.191±0.016(t=15.926,P=0.000),In secretory phase,the difference of Fhit expression between AM eutopic endometrium and ectopic endometrium was significant too-0.298±0.018 vs 0.192±0.017(t=13.803, P=0.000).It is evident that the bionomics of EM ectopic changed further.1.5 Difference of Fhit expression in different r-AFS stages:Fhit of OEM ectopic endometrium showed significant difference in different r-AFS stages(F=6.299, P=0.004).But Fhit of OEM eutopic endometrium showed no difference in different r-AFS stages(F=0.170,P=0.844).Fhit inⅠ～Ⅱstage was higher than that ofⅢstage(0.296±0.024 vs 0.297±0.016,P=0.009) andⅣstage(0.296±0.024 vs 0.300±0.016,P=0.001).It is clear that bionomics change of eutopic endometrium wasn't in relation with the development of EM.The development of EM was determined by the bionomics of ectopic endometrium.2 Ki-67 expression2.1 Location of ki-67 expression:Ki-67 mainly expressed in the cell nucleus and cell membrane of endometrium glandual epithelial.The stroma of eutopic endometrium had weak expression,but the stroma of EM ectopic endometrium hadn't expression.2.2 Difference of ki-67 expression in different menstrual phase:Ki-67 expression of glandular epithelium and stromal cells in control group endometrium and EM group eutopic endometrium in proliferative phase were all higher than that in secretory phase(P＜0.05),ki-67 of ectopic endometrium in proliferative phase showed no difference with that in secretory phase(P＞0.05).It was evident that proliferation of ectopic endometrium lost cyclical viariation and keeped continual activated condition.2.3 Difference of ki-67 expression in eutopic endometrium of control group,OEM group and AM group:Ki-67 of EM eutopic endometrium showed no difference with normal endometrium(P＞0.05),which indicated that the proliferative ability of EM eutopic endometrium was equal to that of normal endometrium.2.4 Difference of ki-67 expression between eutopic and ectopic endometrium:Ki-67 of ectopic endometrium was lower than that of eutopic endometrium in proliferative phase[OEM-66.03±5.60 vs 9.35±1.26(t=49.149,P=0.000);AM-63.50±5.22 vs 9.26±1.51(t=38.659,P=0.000)],but higher than eutopic endometrium in secretory phase[OEM-5.01±0.94 vs 8.92±1.11(t=12.327,P=0.000);AM-5.13±0.87 vs 8.75±1.24(t=6.323,P=0.000)].Expression of OEM showed no difference with that of AM(P＞0.05).It was clear that the ectopic endometrium still keeped activated condition in secretory phase.2.5 Difference of ki-67 expression in different r-AFS stages:Ki-67 of OEM ectopic endometrium glandular epithelium inⅠ～Ⅱphase was higher than that inⅢ,Ⅳphase(P=0.020 in proliferative phase,P=0.031 in secretory phase),but ki-67 of OEM eutopic endometrium showed no difference in different r-AFS phases(P＞0.05).It indicated that the ectopic endometrium in different r-AFS stage has different proliferative ability.3 The correlation of Fhit and ki-67:Fhit and ki-67 expression of EM eutopic endometrium showed no correlation(r=-0.171,P=0.245),but Fhit and ki-67 expression of EM ectopic endometrium showed significant positive correlation(r=0.301,P=0.038).It is evident that Fhit of ectopic endometrium had significant influence on proliferative ability.【Conclusions】 1 Expression of Fhit and ki-67 of EM endometrium had changed as compared with normal endometrium.Fhit and ki-67 may play important roles in the the occurence and progress of EM.2 The sequence of Fhit expression,normal endometrium＞EMS eutopic endometrium＞ectopic endometrium,indicated that EM eutopic endometrium had different bionomics with normal endometrium,which supports "eutopic endometrium determinism".3 In menstrual cycle,the proliferative ability of endometrium had cyclical variation. Proliferation of ectopic endometrium lost cycling and keeped continual activated condition,which probably was the cause of EM.4 Fhit and ki-67 of OEM ectopic endometrium showed significant difference in different r-AFS phases,which manifests Fhit and ki-67 are related with the severity of EM.The development of EM was determined by the bionomics of ectopic endometrium.However,bionomics change of eutopic endometrium wasn't in relation with the development of EM.5 Fhit and ki-67 expression of EM eutopic endometrium showed no correlation,but Fhit and ki-67 expression of EM ectopic endometrium showed significant positive correlation,which indicated that Fhit had not significant affection on the proliferation of EM eutopic endometrium cells but on the proliferation of EM ectopic endometrium cells.【SUMMARY】In this study,immunohistochemistrical SP staining method was used to determined Fhit and ki-67 expression in normal and EM endometrium.The differences of Fhit and ki-67 expression among normal endometrium,OEM endometrium and AM endometrium,between eutopic endometrium and ectopic endometrium,between proliferative phase and secretory phase,among different r-AFS stage,were compared respectively.The results were as follow:Fhit expression in normal,OEM and AM endometrium hadn't cyclical viariation in menstrual cycle. The sequence of Fhit expression was:normal endometrium＞EMS eutopic endometrium＞ectopic endometrium.Fhit of OEM ectopic endometrium showed significant difference in different r-AFS stages.But Fhit of OEM eutopic endometrium showed no difference in different r-AFS stages.Ki-67 expression in control endometrium and EM eutopic endometrium in proliferative phase were all higher than that in secretory phase.Ki-67 of ectopic endometrium lost cyclical viariation.Ki-67 of EM eutopic endometrium showed no difference with normal endometrium.Ki-67 of ectopic endometrium was lower than that of eutopic endometrium in proliferative phase,but higher than eutopic endometrium in secretory phase.Ki-67 of OEM eutopic endometrium showed no difference in different r-AFS phases,but ki-67 of OEM ectopic endometrium showed significant difference in different r-AFS phases.Fhit and ki-67 expression of EM eutopic endometrium showed no correlation,but Fhit and ki-67 expression of EM ectopic endometrium showed significant positive correlation.The upper results showed that EM eutopic endometrium had different bionomics with normal endometrium,the bionomics of EM ectopic changed further.It is evident that Fhit and ki-67 may play important roles in the the occurence and progress of EM.