The Expression And Effects Of Osteopontin, BAFF At The Maternal-Fetal Interface

PARTâ… THE EXPRESSION OF OSTEOPONTIN AT THE MATERNAL-FETAL INTERFACE AND IT'S EFFECT ON THE RECRUITMENT OF uNK CELLOBJECTIVE:To study the expression of osteopontin(OPN)in trophoblasts and deciduas of normal early pregnant women and unexplained recurrent spontaneous abortion(URSA)patients and the relationship between the expression of OPN and the the number of CD56⁺NK cell in deciduas.To demonstrate the effect of OPN on migration of CD56⁺NK cell to the uterine.To explore the effect of progesterone on the expression of OPN in early pregnancy decidual stromal cells in vitro.METHODS:(1)45 clinically elective normal pregnancy termination and 45 unexplained recurrent spontaneous abortion patients(gestational age,4-10w)were included at the Department of Obstetrics and Gynecology,Qi Lu Hospital of Shandong University from Jan 2005 to Oct 2006.The 45 patients were allotted to three groups according to gestational weeks(group1:4-5^6/7w;group2:6-7^6/7w,group3: 8-10w;n=15 each group),respectively.The first-trimester human trophoblasts and deciduas were obtained from the above patients.(2)Expression of OPN in the trophoblasts and deciduas of clinically elective normal pregnancy termination and unexplained recurrent spontaneous abortion patients was studied by reverse transcriptase-polymerase chain reaction(RT-PCR)and immunohistochemistry.(3)The number of CD56⁺NK cell was detected in URSA deciduas and artificial abortion deciduas by immunohistochemistry.(4)Correlations between the expression of OPN and the number of CD56⁺NK cell in decidual tissues were analyzed by Spearman's correlation coefficient by rank test.(5)Early pregnancy decidual stromal cell was isolated by enzymolysis method and cultured in vitro.Decidual stromal cell was investigated under the inverted microscope and identified by immunohistochemical analysis.(6)Passaged decidual stromal cells in 6-well plates and plastic flasks were cultured in serum-free media supplemented with 5×10^-7mol/L of progesterone dissolved in ethanol for 48,72 and 96h.Ethanol was also used as a vehicle control. Expression of OPN in cells was studied by RT-PCR and immunohistochemistry.RESULTS:(1)OPN were clearly detected in total RNA acquired from first-trimester trophoblasts and deciduas of normal early pregnant women and unexplained recurrent spontaneous abortion patients by RT-PCR.There was significantly higher expression of OPN in the trophoblasts and deciduas of normal early pregnant women compared to that of unexplained recurrent spontaneous abortion patients under the same gestational weeks(p＜0.05).The expression of OPN was increaded as gestation progressed(p＜0.05).(2)OPN protein were expressed in the cytoplasm of epithelia and stroma cell of human deciduas.In trophoblasts,OPN protein was localized to the cytotrophoblast,and almost no staining in the syncytiotrophoblast cells.The expression of OPN was decreased in the trophoblasts and deciduas of unexplained recurrent spontaneous abortion patients compared to that of normal early pregnant women(p＜0.05).This was consistent with the results of RT-PCR.(3)CD56 was expressed in the cytoplasm of NK cells.The location of CD56⁺NK cells in the decidual stroma was focally and diffusely.There was dramatic increase of CD56⁺ NK cell accompanied with first-trimester gestation progression (P＜0.05).The number of CD56⁺ NK cell in the deciduas of normal early pregnant women was more than that of unexplained recurrent spontaneous abortion patients (P＜0.05).(4)Expression of OPN in deciduas has positive-correlation with number of CD56⁺ cells(r_s=0.87,P＜0.05).(5)The isolated decidual stromal cells attached after overnight and were positive for vimentin staining.(6)We tested the effect of 5×10^-7mol/L progesterone on the expression of OPN in decidual stromal cells for 48, 72,96h and found that the stimulating effect was hormone time-dependent.The expressions of OPN in the control cells didn't changed with the culture time prolonged.CONCLUSIONS:(1)OPN probably is orchestrated to ensure establishment and maintenance of a successful pregnancy.The lower expression of OPN may be involved in the pathogenesis of URSA.(2)OPN expressed at the maternal-fetus interface and may relate with the recruitment of the CD56⁺ NK cell to the uterus.(3) Progesterone may affect CD56⁺ NK cell recruitment through influencing the expression of OPN. PARTâ…¡THE EXPRESSION AND FUNCTION OF BAFF AT THE MATERNAL-FETAL INTERFACEOBJECTIVE:To study the expression and the effect of B cell-activating factor of TNF family(BAFF)in trophoblasts and deciduas of normal early pregnant women and unexplained recurrent spontaneous abortion(URSA)patients.METHODS:(1)45 clinically elective normal pregnancy termination and 45 unexplained recurrent spontaneous abortionpatients(gestational age,4-10w)were included at the Department of Obstetrics and Gynecology,Qi Lu Hospital of Shandong University from Jan 2005 to Oct 2006.The 45 patients were allotted to three groups according to gestational weeks(group1:4-5^6/7w;group2:6-7^6/7w;group3: 8-10w;n=15 each group),respectively.The trophoblasts and deciduas were gotten in the asepsis operations.(2)The expression of BAFF and BAFF-R in trophoblasts and deciduas of normal early pregnant women and URSA patients was detected by reverse transcriptase-polymerase chain reaction(RT-PCR)and Western blotting.(3)The expression and distribution of BAFF protein at the maternal-fetal interface of normal early pregnant women and URSA patients was observed with technique of immunohistochemical staining.RESULTS:(1)The expression of BAFF were clearly detected at RNA and protein level in first-trimester trophoblasts and deciduas of normal early pregnant women and unexplained spontaneous abortion patients by RT-PCR and Western blotting.There was significantly higher expression of BAFF in the trophoblasts and deciduas of normal early pregnant women compared to that of unexplained spontaneous abortion patients under the same gestational weeks(p＜0.05).(2)The expression of BAFF-R was not observed in the trophoblasts and deciduas of all the samples.(3)BAFF protein was localized to the cytotrophoblast and syncytiotrophoblast cells.In the same cells of unexplained spontaneous abortion patients,immunostaining was weaker than that of normal early pregnant women. BAFF in deciduas of normal early pregnant women was localized to stromal cells and blood vessel endothelium cells.As in first--trimester trophoblasts,BAFF declined as immunohistochemistry was performed in decidual cells of unexplained spontaneous abortion patients.We failed to find the localization of BAFF protein in decidual blood vessel endothelium cells of unexplained spontaneous abortion patients.CONCLUSIONS:BAFF,a member of the placenta-derived, nonapoptosis-inducing TNF superfamily members,is an important factor in the implantation of blastocyst and maintenance of pregnancy.The lower expression of BAFF may be involved in the pathogenesis of URSA.