肿瘤坏死因子在小鼠药理性孕酮撤退月经样模型中的定位和表达
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摘要
目的在药理性撤退小鼠月经样模型中,观察孕酮撤退后,肿瘤坏死因子(TNF-α)的定位和表达情况。方法卵巢切除的小鼠在诱导蜕膜化之后给予米非司酮即通过拮抗孕酮受体的方法(药理性撤退),导致孕酮撤退,并用免疫组化的方法观察米非司酮处理的不同时相,TNF-α在小鼠子宫内膜的定位;用实时PCR的方法观察TNF-α表达情况。结果免疫组化结果显示米非司酮处理后0～8 h,TNF-α主要存在于腔周围的血管内皮细胞;12～16 h,主要存在于内流的白细胞;24～48 h,主要存在于崩解组织边缘的间质细胞区域和修复好腺上皮细胞周围的间质细胞。实时PCR结果显示TNF-αmRNA在孕酮撤退后24 h内都有表达,并且TNF-α表达量逐渐增高,在12 h、16 h达到峰值。结论在此小鼠月经样模型中,米非司酮处理即孕酮撤退后TNF-α在此模型子宫中定位呈区域性分布,并诱导TNF-α表达升高,可能参与了子宫内膜的崩解。
Objective:To investigate the localization and expression of tumor necrosis factor-alfa(TNF-α)in mouse menstrual-like model created by pharmacologic progesterone withdrawal. Methods:The menstrual-like model was prepared by administration of mifepristone(RU486 ) after decidualization of endometrium in ovariectomized mice.The location of TNF-αprotein in uterus was detected by immunohistochemistry.The mRNA expression of TNF-αwas analyzed by real-time PCR at different time within 24 hours after RU486 administration. Results:The immunohistochemical results showed that TNF-αimmunopositive cells were present in the focal subluminal vascular endothelial cells 0-8 hours after administration of mifepristone.The staining appeared in leukocytes in the myometrium after 12-16 hours and the stroma in the basal zone and the decidualized zone,as well as in the extracellular matrix(ECM)across the border of dissociating tissue after 24-48 hours.In addition,TNF-αwas also present in repaired stromal cells around glandular epithelium.RU486 induced a remarkable increase in expression of TNF-αmRNA with maximal level at 12 and 16 hours. Conclusion:The localization of TNF-αin the model displays a regional character.Progesterone withdrawal can induce expression of TNF-α,which may be involved in the process of tissue breakdown in the model.
Objective:To investigate the localization and expression of tumor necrosis factor-alfa(TNF-α)in mouse menstrual-like model created by pharmacologic progesterone withdrawal. Methods:The menstrual-like model was prepared by administration of mifepristone(RU486 ) after decidualization of endometrium in ovariectomized mice.The location of TNF-αprotein in uterus was detected by immunohistochemistry.The mRNA expression of TNF-αwas analyzed by real-time PCR at different time within 24 hours after RU486 administration. Results:The immunohistochemical results showed that TNF-αimmunopositive cells were present in the focal subluminal vascular endothelial cells 0-8 hours after administration of mifepristone.The staining appeared in leukocytes in the myometrium after 12-16 hours and the stroma in the basal zone and the decidualized zone,as well as in the extracellular matrix(ECM)across the border of dissociating tissue after 24-48 hours.In addition,TNF-αwas also present in repaired stromal cells around glandular epithelium.RU486 induced a remarkable increase in expression of TNF-αmRNA with maximal level at 12 and 16 hours. Conclusion:The localization of TNF-αin the model displays a regional character.Progesterone withdrawal can induce expression of TNF-α,which may be involved in the process of tissue breakdown in the model.
引文
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[15]Harada T,Iwabe T,Terakawa N.Role of cytokines in endometriosis[J].Fertil Steril,2001,76:1-10..
[2]Ross JW,Ashworth MD,Mathew D,et al.Activation of the transcription factor,nuclear factor kappa-B.during the estrous cycle and early pregnancy in the pig[J].Reprod Biol Endocrinol,2010,8:39.
[3]Peluffo MC,Young KA,Hennebold JD,et al.Expression and regulation of tumor necrosis factor(TNF) and TNF-receptor family members in the macaque corpus luteum during the menstrual cycle[J].Mol Reprod Dev,2009,76:367-378.
[4]Catalano RD,Wilson MR,Boddy SC,et al.Comprehensive expression analysis of prostanoid enzymes and receptors in the human endometrium across the menstrual cycle[J].Mol Hum Reprod,2011,17:182-192.
[5]Salamonsen LA,Zhang J,Brasted M.Leukocyte networks and human endometrial remodelling[J].J Reprod Immunol, 2002,57:95-108.
[6]Li YF,Xu XB,Chen XH,et al.The nuclear factor-κB pathway is involved in matrix metalloproteinase-9 expression in RU486-induced endometrium breakdown in mice[J].Hum Reprod,2012,27:2096-106.
[7]Chegini N.Dou Q,Williams RS.An inverse relation between the expression of tumor necrosis factor alpha(TNF-α) and TNF-αreceptor in human endometrium[J],Am J Reprod Immunol,1999,42:297-302.
[8]Brasted M,White CA,Kennedy TG,et al.Mimicking the eventsof menstruation in the murine uterus[J].Biol Reprod, 2003,69:1273-1280.
[9]Xu XB,He B,Wang JD.Menstrual-like changes in mice are provoked through the pharmacologic withdrawal of ??progesterone using mifepristone following induction of ecidualization[J].Hum Reprod,2007,22:3184-3191.
[10]Brannstrom M,Friden BE,Jasper M,et al.Variations in peripheral blood levels of immunoreactive tumor necrosis factor alpha(TNFalpha) throughout the menstrual cycle and secretion of TNFalpha from the human corpus luteum[J]. Eur J Obstet Gynecol Reprod Biol,1999,83:213-217.
[11]Markee JE.Menstruation in intraocular endometrial transplants in the Rhesus monkey[J].Am J Obstet Gynecol, 1978,131:558-559.
[12]Lumsden M A,Brown A,Baird D T.Prostaglandin production from homogenales of separated glandular epithelium and stroma from human endometrium[J].Prostaglandins,1984,28:485-496.
[13]Kim YA.Kim J Y.Kim MR.et al.Tumor necrosis factor-alpha-induced cyclooxygenase-2 overexpression in eutopic endometrium of women with endomet riosis by stromal cell culture through nuclear factor-kappaB activation[J].J Reprod Med,2009,54: 625-630.
[14]Braun D,Gogacz M,Dmowski W,et al.Regulation of endometrial cell apoptosis by TNFa is abnormal in women with endometriosis[J].Fertil Steril,2002,77.1-2.
[15]Harada T,Iwabe T,Terakawa N.Role of cytokines in endometriosis[J].Fertil Steril,2001,76:1-10..