本文選題：寡脫氧核苷酸 + MT01��； 參考：《吉林大學(xué)》2014年碩士論文

【摘要】：MT01是本課題組成員依據(jù)人線粒體DNA序列設(shè)計(jì)的單鏈寡脫氧核苷酸，可以靶向阻斷TLR9的激活，減輕組織的炎癥性損傷，抑制由TLR9激活引起的機(jī)體固有免疫應(yīng)答，其功能和結(jié)構(gòu)已獲得國(guó)家專利保護(hù)[一種具有免疫抑制功能的寡核苷酸，國(guó)家發(fā)明專利，專利號(hào)CN101643496]。本課題組在前期工作中發(fā)現(xiàn)，MT01能夠促進(jìn)骨髓間充質(zhì)干細(xì)胞向成骨細(xì)胞分化，增強(qiáng)成骨細(xì)胞成骨功能；同時(shí)，體內(nèi)實(shí)驗(yàn)證實(shí)，在大鼠牙周炎模型中局部應(yīng)用MT01，可以有效抑制牙周炎大鼠牙槽骨吸收。以上研究成果提示，MT01有可能開發(fā)為一種減緩牙槽骨吸收的新型制劑。因此，有必要對(duì)局部應(yīng)用時(shí)，MT01在體內(nèi)重要臟器內(nèi)的分布、代謝、清除率以及毒理反應(yīng)等進(jìn)行系統(tǒng)的研究，從而為進(jìn)一步深入研究開發(fā)MT01奠定實(shí)驗(yàn)基礎(chǔ)。 目的：通過(guò)局部注射Cy5標(biāo)記寡脫氧核苷酸MT01，采用激光共聚焦顯微鏡觀察MT01在大鼠體內(nèi)重要臟器組織中不同時(shí)間點(diǎn)的分布情況，初步探討其代謝分布的規(guī)律性。 方法：選取60只SPF級(jí)健康雄性Wistar大鼠，體重200g±10g，隨機(jī)數(shù)字表法分為實(shí)驗(yàn)組和對(duì)照組，每組30只。實(shí)驗(yàn)組于右側(cè)上頜第一恒磨牙頰側(cè)牙齦粘膜注射Cy5標(biāo)記的MT012μg，對(duì)照組注射未經(jīng)標(biāo)記的MT012μg，于注射后15min、1h、4h、8h、16h、1d、2d、3d、4d和5d全麻下取大鼠肺、肝、脾、腎、心和腦組織，制備快速冰凍切片并染色。激光共聚焦顯微鏡下觀測(cè)MT01熒光分布，以熒光陽(yáng)性細(xì)胞率表示各臟器組織內(nèi)MT01進(jìn)入的量。 結(jié)果：在對(duì)照組中未觀察到熒光陽(yáng)性細(xì)胞。在實(shí)驗(yàn)組中除心、腦組織外，肺、肝、脾和腎組織細(xì)胞中均可觀察到熒光陽(yáng)性細(xì)胞；腎組織中熒光陽(yáng)性細(xì)胞呈片狀分布，熒光主要集中于腎小管上皮細(xì)胞的胞質(zhì)中；肺、肝、脾組織中熒光陽(yáng)性細(xì)胞呈點(diǎn)狀散在分布；肝、脾和腎組織的熒光陽(yáng)性細(xì)胞率隨時(shí)間呈規(guī)律性變化，峰值分別出現(xiàn)在注射后4、3和4d時(shí)，而肺組織的熒光陽(yáng)性細(xì)胞率隨時(shí)間未見明顯規(guī)律性。腎組織的熒光陽(yáng)性細(xì)胞率明顯高于其他組織。 結(jié)論：MT01經(jīng)大鼠牙齦局部注射后可進(jìn)入肺、肝、脾和腎組織，分布具有一定規(guī)律性，，MT01主要集中在腎臟組織中，提示腎臟可能為MT01主要代謝器官。
[Abstract]:MT01 is a single-stranded oligodeoxynucleotide designed according to the sequence of human mitochondrial DNA. It can block the activation of TLR9, alleviate the inflammatory injury of tissues and inhibit the innate immune response induced by TLR9 activation.Its function and structure have been protected by national patent [an immunosuppressive oligonucleotide, national invention patent, patent number CN101643496].In our previous work, we found that MT01 could promote the differentiation of bone marrow mesenchymal stem cells into osteoblasts and enhance the osteogenic function of bone marrow mesenchymal stem cells.Local application of MT01 in rat periodontitis model can effectively inhibit alveolar bone resorption in rats with periodontitis.These results suggest that MT01 may be a novel preparation for reducing alveolar bone resorption.Therefore, it is necessary to systematically study the distribution, metabolism, clearance rate and toxicological reaction of TMT 01 in important organs during local application, so as to lay an experimental foundation for further research and development of MT01.Aim: to observe the distribution of MT01 at different time points in important organs of rats by local injection of Cy5 labeled oligodeoxynucleotides (MT01) and to explore the regularity of metabolic distribution of MT01 at different time points.Methods: 60 SPF grade healthy male Wistar rats, weighing 200 g 鹵10 g, were randomly divided into experimental group and control group with 30 rats in each group.Cy5 labeled MT012 渭 g was injected into the buccal mucosa of the right maxillary first permanent molar in the experimental group, and the unlabeled MT012 渭 g was injected into the control group. The lung, liver, spleen, kidney, heart and brain tissues of the rats were harvested under general anesthesia for 4 days, liver, spleen, kidney, heart and brain under general anesthesia for 4 days and 5 days.The fluorescence distribution of MT01 was observed under laser confocal microscope, and the amount of MT01 entering into the tissues was expressed by fluorescence positive cell rate.Results: no fluorescent positive cells were observed in the control group.Fluorescence positive cells were observed in the lung, liver, spleen and kidney except heart and brain tissues in the experimental group, and the fluorescent positive cells in the renal tissue were flake distribution, fluorescence mainly concentrated in the cytoplasm of renal tubular epithelial cells, lung, liver, liver, lung, liver, kidney, liver, lung, liver, lung, liver, lung, liver, lung, liver, lung, liver and liver.The percentage of fluorescent positive cells in liver, spleen and kidney showed regular changes with time, and the peak value appeared at 4d and 4d after injection, respectively.However, the rate of fluorescent positive cells in lung tissues did not show obvious regularity with time.The rate of fluorescent positive cells in renal tissue was significantly higher than that in other tissues.Conclusion the lung, liver, spleen and kidney can be injected into the lung, liver, spleen and kidney by local injection of the rat gingival. The distribution of MT01 is mainly concentrated in the kidney tissue, suggesting that the kidney may be the main metabolic organ of MT01.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R783.1

【參考文獻(xiàn)】

相關(guān)期刊論文 前3條

1 劉紅,魏東芝;反義硫代磷酸酯寡聚核苷酸及其應(yīng)用研究進(jìn)展[J];國(guó)外醫(yī)學(xué).輸血及血液學(xué)分冊(cè);1997年03期

2 王舉;王磊;王有德;;反義核酸與疾病治療研究進(jìn)展[J];生物醫(yī)學(xué)工程學(xué)雜志;2006年01期

3 柳yN,孫宗全;細(xì)胞間粘附分子-1反義寡核苷酸在小鼠體內(nèi)的分布及其作用[J];華中科技大學(xué)學(xué)報(bào)(醫(yī)學(xué)版);2005年02期

本文編號(hào)：1751706