miR-34a對(duì)健康及感染狀態(tài)下MG63細(xì)胞成骨分化的影響
發(fā)布時(shí)間：2019-04-13 14:20
【摘要】：牙周炎是由牙菌斑生物膜引起的牙周組織感染性疾病,以牙齦卟啉單胞菌(Porphyromonas gingivalis,Pg)等為代表的牙周致病菌可以分泌多種毒力因子,激發(fā)宿主固有免疫應(yīng)答,引發(fā)牙槽骨吸收,進(jìn)而導(dǎo)致牙周組織破壞。研究顯示約47%的美國(guó)人患有不同程度的牙周炎,在65歲以上的成年人中,發(fā)病率更是高達(dá)64%。牙周炎導(dǎo)致的牙槽骨吸收現(xiàn)已成為成人失牙的首要原因,嚴(yán)重影響患者咀嚼、發(fā)音,并影響患者生存質(zhì)量。研究牙周炎所致的牙槽骨吸收機(jī)制,抑制炎性牙槽骨吸收并促進(jìn)骨再生具有重要意義。近期研究發(fā)現(xiàn),微小分子RNA(microRNAs,miRNAs)可能是牙周炎所致牙槽骨吸收的生物學(xué)標(biāo)記及治療靶點(diǎn)。研究者認(rèn)為一些miRNAs可以通過(guò)調(diào)控破骨細(xì)胞及成骨細(xì)胞功能,影響牙周炎導(dǎo)致的牙槽骨吸收進(jìn)程。在這些miRNAs中,miR-34a備受關(guān)注。miR-34a是一種具有調(diào)控功能的內(nèi)源性非編碼RNA,其編碼基因位于染色體1p36區(qū)域。近年來(lái)一些研究發(fā)現(xiàn)其可以影響包括炎癥反應(yīng)及骨改建在內(nèi)的多種病理生理進(jìn)程。實(shí)驗(yàn)證實(shí)miR-34a可以抑制LPS誘導(dǎo)的巨噬細(xì)胞炎癥應(yīng)答,抑制破骨細(xì)胞的形成從而抑制骨質(zhì)疏松,并且可以促進(jìn)骨髓間充質(zhì)干細(xì)胞及脂肪干細(xì)胞成骨分化。鑒于miR-34a可以促進(jìn)成骨分化,抑制LPS誘導(dǎo)的炎癥應(yīng)答及破骨細(xì)胞功能,我們推測(cè)miR-34a可能具有促進(jìn)骨形成、抑制炎性牙槽骨吸收的作用。因此我們?cè)O(shè)計(jì)本實(shí)驗(yàn)檢測(cè)miR-34a對(duì)健康及炎癥狀態(tài)下成骨樣細(xì)胞成骨分化能力的影響,為進(jìn)一步探討miR-34a能否抑制牙周炎所致的牙槽骨吸收奠定實(shí)驗(yàn)基礎(chǔ)。方法:1選取人成骨樣細(xì)胞MG63為實(shí)驗(yàn)細(xì)胞,使用Lipofectamine2000將miR-34a擬態(tài)物轉(zhuǎn)染至細(xì)胞。2熒光顯微鏡及實(shí)時(shí)定量PCR法檢測(cè)miR-34a轉(zhuǎn)染效率。3使Pg感染MG63細(xì)胞,實(shí)時(shí)定量PCR法檢測(cè)細(xì)胞內(nèi)miR-34a表達(dá)量變化。4實(shí)時(shí)定量PCR法檢測(cè)轉(zhuǎn)染miR-34a擬態(tài)物后健康或感染狀態(tài)下MG63細(xì)胞中成骨相關(guān)因子Runx2、SP7及COLⅠ基因表達(dá)變化。結(jié)果:Lipofectamine2000可有效將miR-34a擬態(tài)物轉(zhuǎn)染至MG63細(xì)胞內(nèi);miR-34a可以上調(diào)健康MG63細(xì)胞內(nèi)Runx2、SP7及COLⅠ基因表達(dá)水平;感染狀態(tài)下MG63細(xì)胞內(nèi)miR-34a表達(dá)量降低;miR-34a下調(diào)感染狀態(tài)下MG63細(xì)胞內(nèi)Runx2、SP7及COLⅠ基因表達(dá)水平。結(jié)論:miR-34a促進(jìn)健康狀態(tài)下成骨細(xì)胞成骨分化,抑制感染狀態(tài)下成骨細(xì)胞成骨分化。
[Abstract]:Periodontitis is an infectious disease of periodontal tissue caused by dental plaque biofilm. Periodontal pathogenic bacteria, such as Porphyromonas gingivalis (Porphyromonas gingivalis,Pg), can secrete many virulence factors to stimulate host innate immune response. The alveolar bone resorption is induced, which leads to the destruction of periodontal tissue. The study found that about 47 percent of Americans suffer from periodontitis of varying degrees, and that 64 percent of adults over 65 suffer from periodontitis. Alveolar bone absorption caused by periodontitis has become the primary cause of adult tooth loss, which seriously affects the mastication, pronunciation and quality of life of the patients. It is of great significance to study the mechanism of alveolar bone resorption induced by periodontitis, inhibit the inflammatory alveolar bone resorption and promote bone regeneration. Recent studies have found that RNA (microRNAs,miRNAs) may be a biological marker and therapeutic target of alveolar bone resorption induced by periodontitis. The researchers believe that some miRNAs may affect alveolar bone resorption by regulating osteoclast and osteoblast function. Among these miRNAs, miR-34a has attracted much attention. MiR-34a is an endogenous non-coding RNA, with regulatory function, and its coding gene is located in the 1p36 region of chromosome. In recent years, some studies have found that it can affect a variety of pathophysiological processes, including inflammatory reaction and bone remodeling. It was proved that miR-34a could inhibit the inflammatory response of macrophages induced by LPS, inhibit the formation of osteoclasts and inhibit osteoporosis, and promote the osteogenic differentiation of bone marrow mesenchymal stem cells and adipose-derived stem cells. Since miR-34a can promote osteogenic differentiation, inhibit LPS-induced inflammatory response and osteoclast function, we speculate that miR-34a may promote bone formation and inhibit inflammatory alveolar bone resorption. Therefore, we designed this experiment to detect the effect of miR-34a on osteoblast-like osteoblast differentiation in healthy and inflammatory condition, which laid the experimental foundation for further exploring whether miR-34a can inhibit alveolar bone resorption induced by periodontitis. Methods: 1 Human osteoblast-like cells (MG63) were selected as experimental cells, and miR-34a mimicry was transfected into cells by Lipofectamine2000. 2 fluorescent microscope and real-time quantitative PCR were used to detect the transfection efficiency of miR-34a. 3. Pg was used to infect MG63 cells. The expression of miR-34a in MG63 cells was detected by real-time quantitative PCR. (4) the expression of osteogenic factors Runx2,SP7 and COL 鈪，
本文編號(hào)：2457648
[Abstract]:Periodontitis is an infectious disease of periodontal tissue caused by dental plaque biofilm. Periodontal pathogenic bacteria, such as Porphyromonas gingivalis (Porphyromonas gingivalis,Pg), can secrete many virulence factors to stimulate host innate immune response. The alveolar bone resorption is induced, which leads to the destruction of periodontal tissue. The study found that about 47 percent of Americans suffer from periodontitis of varying degrees, and that 64 percent of adults over 65 suffer from periodontitis. Alveolar bone absorption caused by periodontitis has become the primary cause of adult tooth loss, which seriously affects the mastication, pronunciation and quality of life of the patients. It is of great significance to study the mechanism of alveolar bone resorption induced by periodontitis, inhibit the inflammatory alveolar bone resorption and promote bone regeneration. Recent studies have found that RNA (microRNAs,miRNAs) may be a biological marker and therapeutic target of alveolar bone resorption induced by periodontitis. The researchers believe that some miRNAs may affect alveolar bone resorption by regulating osteoclast and osteoblast function. Among these miRNAs, miR-34a has attracted much attention. MiR-34a is an endogenous non-coding RNA, with regulatory function, and its coding gene is located in the 1p36 region of chromosome. In recent years, some studies have found that it can affect a variety of pathophysiological processes, including inflammatory reaction and bone remodeling. It was proved that miR-34a could inhibit the inflammatory response of macrophages induced by LPS, inhibit the formation of osteoclasts and inhibit osteoporosis, and promote the osteogenic differentiation of bone marrow mesenchymal stem cells and adipose-derived stem cells. Since miR-34a can promote osteogenic differentiation, inhibit LPS-induced inflammatory response and osteoclast function, we speculate that miR-34a may promote bone formation and inhibit inflammatory alveolar bone resorption. Therefore, we designed this experiment to detect the effect of miR-34a on osteoblast-like osteoblast differentiation in healthy and inflammatory condition, which laid the experimental foundation for further exploring whether miR-34a can inhibit alveolar bone resorption induced by periodontitis. Methods: 1 Human osteoblast-like cells (MG63) were selected as experimental cells, and miR-34a mimicry was transfected into cells by Lipofectamine2000. 2 fluorescent microscope and real-time quantitative PCR were used to detect the transfection efficiency of miR-34a. 3. Pg was used to infect MG63 cells. The expression of miR-34a in MG63 cells was detected by real-time quantitative PCR. (4) the expression of osteogenic factors Runx2,SP7 and COL 鈪，
本文編號(hào)：2457648
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