本文選題：翻譯控制腫瘤蛋白TCTP + 胚胎著床　； 參考：《重慶醫(yī)科大學》2011年碩士論文

【摘要】：背景和目的 翻譯控制腫瘤蛋白(The translationally controlled tumor protein,TCTP)最初發(fā)現(xiàn)于腫瘤組織,后來又從其他組織中分離出來。TCTP廣泛表達于動物、植物及酵母中,其基因序列高度保守。最初認為TCTP僅是一類生長相關蛋白,近年來有研究表明,TCTP具有多種生物學功能:可作為一種組織胺釋放因子,參與細胞的生長、分裂和分化等;也可作為一種小分子熱休克蛋白,以分子伴侶的形式保護細胞蛋白免受熱休克損傷;同時還可作為一種鈣結合蛋白,在人類滋養(yǎng)層細胞中起鈣運輸?shù)淖饔�。另�?課題組在小鼠胚胎著床相關基因文庫研究中發(fā)現(xiàn)TCTP基因在小鼠胚胎著床窗口期子宮內膜組織中呈現(xiàn)高表達。結合文獻和前期研究工作,我們推測TCTP可能參與了胚胎著床和早期胚胎發(fā)育過程。鑒于TCTP在小鼠胚胎圍著床期的研究還未見報道,本實驗運用qRT-PCR、Western blot和免疫組化等方法對小鼠胚胎圍著床期子宮內膜TCTP基因表達進行了分析,并采用子宮角注射實驗初步探討了TCTP基因的功能。這不僅有助于深入理解TCTP在胚胎著床中的作用,而且為探索胚胎著床的分子機制提供相關實驗依據(jù)。 方法 分別采用qRT-PCR法、免疫組化和Western blot法檢測分析孕D1, D2, D3, D4, D5, D6和D7小鼠子宮內膜中TCTP mRNA和蛋白的表達情況。然后以假孕D4和D6的小鼠子宮內膜為對照,分別采用qRT-PCR、免疫組化和Western blot法檢測TCTP在著床前(孕D4)和著床后(孕D6)子宮內膜中的表達。最后對孕D3.5小鼠實施子宮角注射TCTP反義寡聚脫氧核苷酸,24 h后取出子宮內膜組織,再分別用qRT-PCR、Western blot和免疫組化法檢測TCTP mRNA和蛋白表達的差異,同時觀察術后孕D8小鼠子宮中胚胎數(shù)目和形態(tài)的變化以期探討TCTP基因的功能。 結果 qRT-PCR結果顯示TCTP mRNA在孕D1~D5天小鼠子宮內膜中隨孕天數(shù)的遞增其表達量逐漸升高,孕D5(著床窗口期)達到頂峰,隨后孕D6和D7表達下降。Western blot和免疫組化檢測結果與qRT-PCR一致。TCTP主要表達于子宮內膜的腔上皮、腺上皮和基質細胞中。TCTP mRNA和蛋白在正常妊娠小鼠著床前(孕D4)和著床后(孕D6)子宮內膜中的表達均高于假孕D4和D6小鼠(*P0.05)。在小鼠子宮角一側注射TCTP反義寡聚脫氧核苷酸后,與另一側注射TCTP正義寡聚脫氧核苷酸和蒸餾水相比,TCTP mRNA和蛋白的表達均受到抑制,并且小鼠胚胎著床數(shù)目明顯減少(*P0.05)。 結論 TCTP mRNA和蛋白在早孕小鼠子宮內膜圍著床期的表達規(guī)律與胚胎著床時間和部位高度一致,且在著床窗口期(孕D5)呈現(xiàn)出高表達現(xiàn)象;對小鼠實施宮角注射TCTP反義寡聚脫氧核苷酸后,TCTP的表達受到抑制,且小鼠胚胎著床數(shù)目明顯下降,提示TCTP與胚胎著床密切相關,在小鼠胚胎著床過程中可能發(fā)揮著重要的作用。
[Abstract]:Background and purpose The translationally controlled tumor protein (TCTP) was first found in the tumor tissue, then isolated from other tissues. TCTP was widely expressed in animals, plants and yeast, and its gene sequence was highly conserved. It was initially thought that TCTP is only a kind of growth-related protein. In recent years, it has been shown that TCTP has many biological functions: it can be used as a histamine releasing factor to participate in cell growth, division and differentiation. It can also be used as a small molecular heat shock protein to protect cell proteins from heat shock damage in the form of molecular chaperones and to act as a calcium binding protein in the transport of calcium in human trophoblast cells. In addition, the study of mouse embryo implantation related gene library showed that TCTP gene was highly expressed in the endometrium of mouse embryo implantation window. We speculate that TCTP may be involved in embryo implantation and early embryonic development. In view of the fact that the study of TCTP in the period of mouse embryo surrounding bed has not been reported, the expression of TCTP gene in the endometrium of mouse embryo around the bed was analyzed by using qRT-PCRG Western blot and immunohistochemical method. The function of TCTP gene was preliminarily studied by the experiment of uterine horn injection. This will not only help to understand the role of TCTP in embryo implantation, but also provide experimental basis for exploring the molecular mechanism of embryo implantation. Method The expressions of TCTP mRNA and protein in endometrium of pregnant D1, D2, D3, D4, D5, D6 and D7 mice were detected by qRT-PCR, immunohistochemistry and Western blot. Then the expression of TCTP in preimplantation (D4) and postimplantation (D6) endometrium was detected by qRT-PCR, immunohistochemistry and Western blot method. TCTP antisense oligodeoxynucleotides (TCTP antisense oligodeoxynucleotides) were injected into the uterus of pregnant D3.5 mice for 24 hours. The expression of TCTP mRNA and protein was detected by qRT-PCRG Western blot and immunohistochemistry. At the same time, the number and morphology of embryo in the uterus of pregnant D8 mice after operation were observed in order to investigate the function of TCTP gene. Result QRT-PCR results showed that the expression of TCTP mRNA in endometrium of D1~D5 mice increased gradually with the increase of pregnant days, and the peak of D5 (implantation window period) was observed. Subsequently, the expression of D6 and D7 decreased. Western blot and immunohistochemistry showed that D6 and D7 were mainly expressed in the luminal epithelium of endometrium. The expression of. TCTP mRNA and protein in glandular epithelium and stromal cells in normal pregnant mice before and after implantation (D4) and postimplantation (D6) were higher than those in pseudo-pregnant D4 and D6 mice. After injection of TCTP antisense oligodeoxynucleotides into one side of mouse uterus horn, the expression of mRNA and protein of TCTP sense oligodeoxynucleotides and distilled water were inhibited, and the number of embryo implantation in mice was significantly decreased than that of TCTP sense oligodeoxynucleotides and distilled water. Conclusion The expression of TCTP mRNA and protein in endometrium of early pregnant mice was highly consistent with the time and location of embryo implantation, and the expression of TCTP mRNA and protein was high in the implantation window (D5). The expression of TCTP antisense oligodeoxynucleotides was inhibited after injection of antisense oligodeoxynucleotides into mouse uterus horn, and the number of mouse embryo implantation decreased significantly, suggesting that TCTP is closely related to embryo implantation and may play an important role in mouse embryo implantation.
【學位授予單位】：重慶醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2011
【分類號】：R346;R714.1

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