【摘要】：本文采用生物力學(xué)、細(xì)胞生物學(xué)、分子生物學(xué)等方法,研究機(jī)械拉伸刺激對成骨細(xì)胞生理活性和生長因子表達(dá)的影響,重點(diǎn)就力學(xué)信號——基因表達(dá)關(guān)系進(jìn)行了初步探索。主要工作和結(jié)論如下: ①采用消化法從初生大鼠的顱蓋骨中分離、純化、培養(yǎng)和鑒定成骨細(xì)胞。 ②使用周期性拉伸裝置和靜態(tài)拉伸裝置對大鼠顱蓋骨成骨細(xì)胞施加機(jī)械拉伸應(yīng)變,應(yīng)變水平均為15%,受載時(shí)間為6、12、24、36h,研究同一應(yīng)變水平的機(jī)械拉伸刺激對成骨細(xì)胞在不同加載時(shí)間生化響應(yīng)以及基因表達(dá)的影響。 1) 應(yīng)用流式細(xì)胞技術(shù)研究機(jī)械拉伸應(yīng)變對成骨細(xì)胞增殖的影響,探討此類細(xì)胞在力學(xué)生理背景下的生長行為。結(jié)果表明:同一應(yīng)變水平(15%)的周期性拉伸和靜態(tài)拉伸下,隨著受載時(shí)間的增加,成骨細(xì)胞的相對增殖指數(shù)增加,在各加載時(shí)間點(diǎn)周期性拉伸下成骨細(xì)胞相對增殖指數(shù)都大于靜態(tài)拉伸下成骨細(xì)胞的相對增殖指數(shù),但變化趨勢相似。在受載6h 后,成骨細(xì)胞的相對增殖指數(shù)最大,而在受載12h、24h、36h 后,成骨細(xì)胞相對增殖指數(shù)下降,特別是在受載36h 后,細(xì)胞的相對增殖指數(shù)趨向于1。說明機(jī)械拉伸能促進(jìn)細(xì)胞增殖,并且這種促進(jìn)作用與受載時(shí)間相關(guān)。而且周期性機(jī)械拉伸的促進(jìn)作用比靜態(tài)機(jī)械拉伸的作用更強(qiáng),就相對增殖指數(shù)而言,受載的初期(6h),對成骨細(xì)胞增殖的影響最明顯。 2) 同一應(yīng)變水平(15%)周期性拉伸與靜態(tài)拉伸作用下,受載成骨細(xì)胞堿性磷酸酶相對活力和胞外無機(jī)鈣質(zhì)的相對分泌量顯示,靜態(tài)拉伸下成骨細(xì)胞在受載初期(6h),與對照組相比,相對堿性磷酸酶活力和胞外無機(jī)鈣質(zhì)的相對分泌量均略小于1,隨受載時(shí)間的增加逐漸升高且均大于1。相對堿性磷酸酶活力36h 達(dá)最高;相對胞外鈣分泌量24h 達(dá)最高,36h 后略有下降,但仍大于1。而周期性拉伸下的成骨細(xì)胞,各受載時(shí)間點(diǎn)相對堿性磷酸酶活力與相對胞外鈣分泌量都小于1。說明短期靜態(tài)拉伸刺激能抑制成骨細(xì)胞的分化、礦化,較長時(shí)間的靜態(tài)拉伸刺激能促進(jìn)成骨細(xì)胞的分化、礦化。而周期性拉伸刺激對成骨細(xì)胞的分化、礦化起抑制作用。 3) 通過半定量RT-PCR 方法分析了機(jī)械拉伸對成骨細(xì)胞IGF-1、MGF mRNA表達(dá)的影響。結(jié)果顯示:在周期性拉伸組、靜態(tài)拉伸組和對照組中IGF-1 都有一定水平的表達(dá),相比較而言,周期性拉伸組細(xì)胞表達(dá)最強(qiáng)。MGF 在周期性拉伸組細(xì)胞中的表達(dá)明顯高于靜態(tài)拉伸組細(xì)胞的表達(dá),對照組幾乎沒有表達(dá)。而在各受載時(shí)間組中IGF-1 mRNA 、MGF mRNA 的表達(dá)量都是周期性拉伸組細(xì)胞明顯強(qiáng)于靜態(tài)拉伸組細(xì)胞。MGF mRNA 在6h 后達(dá)到最大值,IGF-1 mRNA 在12h 后達(dá)到最
[Abstract]:In this paper, biomechanics, cell biology and molecular biology were used to study the effects of mechanical tensile stimulation on the physiological activity and the expression of growth factors in osteoblasts, with emphasis on the relationship between mechanical signal-gene expression. The main work and conclusions are as follows: 1 digested and purified from the skull of newborn rats. Culture and identification of osteoblasts. (2) Mechanical tensile strain was applied to osteoblasts of rat calvaria with cyclic and static stretching devices. The effects of mechanical tensile stimulation at the same strain level on the biochemical response and gene expression of osteoblasts at different loading times were studied. 1) the effects of mechanical tensile strain on proliferation of osteoblasts were studied by flow cytometry. To study the growth behavior of this kind of cells under the background of mechanics and physiology. The results showed that the relative proliferation index of osteoblasts increased with the increase of loading time at the same strain level (15%). The relative proliferation index of osteoblasts under cyclic stretching was higher than that under static tension, but the trend was similar. The relative proliferation index of osteoblasts was the highest after loading for 6 h, while that of osteoblasts decreased after being loaded for 12 h or 24 h or 36 h, especially after being loaded for 36 h, the relative proliferation index of osteoblasts tended to be 1. The results showed that mechanical stretching could promote cell proliferation, and the effect was related to loading time. And the effect of periodic mechanical stretching is stronger than that of static mechanical stretching, in terms of relative proliferation index, At the early stage of loading (6h), the proliferation of osteoblasts was affected most obviously. 2) under the same strain level (15%), the cells were subjected to cyclic and static stretching. The relative activity of alkaline phosphatase (ALP) and the relative secretion of extracellular calcium in osteoblasts were compared with those in the control group. The activity of relative alkaline phosphatase and the relative secretion of extracellular inorganic calcium were slightly less than 1, and gradually increased with the increase of loading time. The relative alkaline phosphatase activity reached the highest level at 36h, and the relative extracellular calcium secretion reached the highest level at 24h, and decreased slightly after 36h, but still more than 1. The relative alkaline phosphatase activity and extracellular calcium secretion of osteoblasts under cyclic stretching were less than 1. The results showed that the short-term static tensile stimulation could inhibit the differentiation of osteoblasts and mineralization, and the long time static tension stimulation could promote the differentiation and mineralization of osteoblasts. Periodic tensile stimulation inhibited the differentiation and mineralization of osteoblasts. 3) the effect of mechanical stretching on IGF-1,MGF mRNA expression in osteoblasts was analyzed by semi-quantitative RT-PCR. The results showed that there was a certain level of IGF-1 expression in the periodic stretching group, the static stretching group and the control group. The expression of MGF was significantly higher in the periodic stretching group than in the static stretching group, but almost no expression in the control group. However, the expression of IGF-1 mRNA, MGF mRNA was significantly stronger in the cyclically stretched group than in the static stretch group at 6 h, and the maximum expression of IGF-1 mRNA was reached at 12 h after each loading time.
【學(xué)位授予單位】：重慶大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2005
【分類號】：R329

【引證文獻(xiàn)】

相關(guān)碩士學(xué)位論文 前1條

1 王峰;不同負(fù)荷運(yùn)動(dòng)對老年大鼠IGF-I表達(dá)及骨骼生物力學(xué)性能的影響[D];河北師范大學(xué);2011年

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本文編號：2278306