模拟失重条件下对骨髓体外成骨能力及承重骨影响的研究
发布时间：2018-08-07 19:54
【摘要】： 空间飞行过程中，宇航人员暴露于失重环境下，将导致骨骼系统发生结构和功能的变化，主要表现为骨量丢失，骨骼脱矿，骨力学性能下降，出现负钙平衡。这种失重导致的骨骼系统变化，在重力生理学中称为失重性骨质稀少。失重性骨质稀少是人类长期太空停留和探索其它星球的主要障碍之一。虽然经过大量研究，但目前对失重性骨丧失的机理仍缺乏深刻认识，所以，深入探讨失重性骨丧失的发生机理，发展行之有效的对抗措施是目前重力生理学的重要任务。已有的研究表明，失重性骨丧失主要发生在承重骨。其机制主要是抑制了局部成骨作用，而对破骨细胞活性可能影响不大。在此过程中，由骨的细胞分泌并在局部发挥作用的一些调节蛋白和细胞因子被认为起着重要作用，但这类物质在失重条件下的确切作用及其机制并不是十分清楚。 本研究采用后肢去负荷大鼠模型，通过细胞培养观察了4w模拟失重后对骨基质细胞数及骨髓体外成骨能力的影响；比较2w和4w模拟失重后对骨局部调节蛋白和细胞因子的影响作用，以及骨形态学的变化。主要结果与发现如下： 1．4w尾吊对大鼠股骨基质细胞数的影响 本实验通过细胞培养进行原代细胞培养计数，细胞计数法和MTT法绘制生长曲线，，结果显示：S组培养系的股骨基质细胞数明显减少，约是C组细胞数的50％。S组和C组细胞生长曲 第四军医大学硕士学位论文 一 线和倍增时问相似，表明S组和C组细胞增咱速度相似。提示。lw模拟失重大 鼠股骨基质细胞数减少。 2．4W 尾吊对大鼠股骨骨髓体外成骨能力的影响 本实验通过细胞培养 检测原代和传代培养细胞，ALP活性及体外矿化小结形成量。结果显示：原代 培养中S组ALP活性和小结形成量同C组相比较显著下降（50％），说明骨髓成 骨祖细胞减少，导致分化的成骨细胞数减少。传代培养中细胞接种密度相同， S组竹P活性和小结形成数较C明显降低（约50们，表明去负荷对骨髓基质中 的成骨祖细胞有抑制作用。 3．OPNr，OC，BMP和 TGF－日在 ZW，4W尾吊大鼠承重骨的变化 本实验通过 原位杂交的方法检测了股骨和骨髓 OPN，OC，BMP和 TGF－6。结果发现：S组 OPN，OC，BMP和 TGF－p在骨细胞和骨髓中含量显著低于 C组，S组 4W低于 ZW。 提示模拟失重可能抑制了成骨细胞分泌OPN，影响细胞在骨面的粘附。同样可 能抑制了 OC的产生，影响骨质的矿化。另外抑制了 BMP和 TGF－p的产生及促 成骨作用的发挥。 4．ALP和 ACP在 ZW，4W尾吊大鼠承重骨的变化 本实验通过特殊染色的 方法检测了股骨ALP和 ACP。结果显示尾吊大鼠股骨中 ALP显著降低，ACP显 著增高。提示模拟失重后，大鼠成骨作用降低，破骨作用增强。 5．承重骨形态学变化 本实验通过三色染色的方法观察股骨形态学变 化，显示骨基质钙化障碍，原有沉积钙盐脱钙明显，提示模拟失重使大鼠钙 盐代谢障碍。 总之，本工作利用细胞培养的方法，观察到模拟失重对骨基质细胞数减 少和骨髓体外成骨能力下降的影响；通过原位杂交的方法观察BMP、TGF－p、 OPN和 OC在模拟失重中承重骨分泌量减少的变化；通过特殊染色方法观察骨 质中 ALP降低和 ACP增高及承重骨中钙盐代谢的形态学变化，对进一步探讨 失重性骨丧失的机理具有参考意义。
[Abstract]:During space flight, the space flight personnel exposed to the weightlessness environment will lead to changes in the structure and function of the skeletal system, which mainly include bone loss, bone demineralization, bone mechanical performance decline, and negative calcium balance. This kind of weightlessness results in skeletal system change, which is called weightless bone scarcity in heavy stress physiology. Scarce is one of the main obstacles for human long space stay and exploration of other planets. Although a lot of research has been done, the mechanism of weightless bone loss is still lack of deep understanding. Therefore, it is an important task for gravity physiology to explore the mechanism of weightless bone loss and to develop effective countermeasures. Studies have shown that weightless bone loss mainly occurs in the load-bearing bone. Its mechanism is mainly to inhibit the local osteogenesis and may not affect the activity of osteoclast. In this process, some regulatory proteins and cytokines secreted by bone cells and part of the role play an important role in the process, but these substances are in weightless strips. The exact role and mechanism of this component are not very clear.
The effect of 4W simulated weightlessness on the number of bone stromal cells and bone marrow in vitro after weightlessness was observed by cell culture, and the effects of 2W and 4W on bone local regulatory proteins and cytokines after weightlessness were compared, and the changes of bone morphology were compared. The main results were as follows:
The effect of 1.4W tail suspension on the number of stromal cells in the femur of rats was carried out by cell culture to count the primary cell culture, cell count method and MTT method to draw the growth curve. The results showed that the number of femur stroma cells in the S group was significantly reduced, which was about the 50%.S and C cell growth of the C group.
Master's degree thesis of The Fourth Military Medical University
One
The similarity between the line and doubling time indicated that the cells in group S and group C were similar in speed, suggesting that.Lw simulation was not significant.
The number of stromal cells in the femur of the rat was reduced.
Effects of 2.4W tail suspension on osteogenic potential of femoral bone in rats in vitro
The ALP activity and mineralization nodule formation in primary and passage cells were detected.
ALP activity and nodule formation in S group decreased significantly compared with C group (50%), indicating bone marrow formation.
The number of osteoblasts decreased as the number of bone progenitor cells decreased.
In group S, P activity and nodule formation were significantly lower than those in C (about 50), indicating that the load was in bone marrow stroma.
The osteoblast progenitor cells have inhibitory effect.
3.OPNr, OC, BMP and TGF - day in ZW, 4W tail suspended rats bearing bone changes in this experiment
OPN, OC, BMP and transforming growth factor - 6 were detected by in situ hybridization. The results showed that: S group
The contents of OPN, OC, BMP and transforming growth factor - P in bone cells and bone marrow were significantly lower than those in group C, while 4W in group S was lower than ZW..
It is suggested that simulated weightlessness may inhibit the secretion of OPN from osteoblasts and affect the adhesion of cells to the bone surface.
It can inhibit the production of OC, affect the mineralization of bone, and inhibit the production and promotion of BMP and TGF P.
The exertion of osteogenesis.
4.ALP and ACP changes in the weight bearing bone of ZW and 4W tail suspended rats.
Methods detection of ALP and ACP. in the femur showed that ALP in the femur of tail suspended rats was significantly lower than that in ACP rats.
The results showed that after simulated weightlessness, the osteogenesis of rats decreased and the osteoclast increased.
5. the morphological changes of the weight-bearing bone were observed by trichrome staining.
It showed that the calcification of bone matrix was impaired and the calcium deposition was decalcified.
Salt metabolism disorder.
In conclusion, cell culture was used to observe the reduction of bone marrow stromal cells by simulated weightlessness.
The effects of BMP and TGF P on bone marrow in vitro were observed by in situ hybridization.
OPN and OC changes in the weight loss of the bearing bone in simulated weightlessness; bone is observed by special staining.
Morphological changes of ALP and ACP in the bone and calcium metabolism in the bearing bone were further explored.
The mechanism of weightless bone loss is of reference significance.
【学位授予单位】：第四军医大学
【学位级别】：硕士
【学位授予年份】：2002
【分类号】：R852.22
本文编号：2171180
[Abstract]:During space flight, the space flight personnel exposed to the weightlessness environment will lead to changes in the structure and function of the skeletal system, which mainly include bone loss, bone demineralization, bone mechanical performance decline, and negative calcium balance. This kind of weightlessness results in skeletal system change, which is called weightless bone scarcity in heavy stress physiology. Scarce is one of the main obstacles for human long space stay and exploration of other planets. Although a lot of research has been done, the mechanism of weightless bone loss is still lack of deep understanding. Therefore, it is an important task for gravity physiology to explore the mechanism of weightless bone loss and to develop effective countermeasures. Studies have shown that weightless bone loss mainly occurs in the load-bearing bone. Its mechanism is mainly to inhibit the local osteogenesis and may not affect the activity of osteoclast. In this process, some regulatory proteins and cytokines secreted by bone cells and part of the role play an important role in the process, but these substances are in weightless strips. The exact role and mechanism of this component are not very clear.
The effect of 4W simulated weightlessness on the number of bone stromal cells and bone marrow in vitro after weightlessness was observed by cell culture, and the effects of 2W and 4W on bone local regulatory proteins and cytokines after weightlessness were compared, and the changes of bone morphology were compared. The main results were as follows:
The effect of 1.4W tail suspension on the number of stromal cells in the femur of rats was carried out by cell culture to count the primary cell culture, cell count method and MTT method to draw the growth curve. The results showed that the number of femur stroma cells in the S group was significantly reduced, which was about the 50%.S and C cell growth of the C group.
Master's degree thesis of The Fourth Military Medical University
One
The similarity between the line and doubling time indicated that the cells in group S and group C were similar in speed, suggesting that.Lw simulation was not significant.
The number of stromal cells in the femur of the rat was reduced.
Effects of 2.4W tail suspension on osteogenic potential of femoral bone in rats in vitro
The ALP activity and mineralization nodule formation in primary and passage cells were detected.
ALP activity and nodule formation in S group decreased significantly compared with C group (50%), indicating bone marrow formation.
The number of osteoblasts decreased as the number of bone progenitor cells decreased.
In group S, P activity and nodule formation were significantly lower than those in C (about 50), indicating that the load was in bone marrow stroma.
The osteoblast progenitor cells have inhibitory effect.
3.OPNr, OC, BMP and TGF - day in ZW, 4W tail suspended rats bearing bone changes in this experiment
OPN, OC, BMP and transforming growth factor - 6 were detected by in situ hybridization. The results showed that: S group
The contents of OPN, OC, BMP and transforming growth factor - P in bone cells and bone marrow were significantly lower than those in group C, while 4W in group S was lower than ZW..
It is suggested that simulated weightlessness may inhibit the secretion of OPN from osteoblasts and affect the adhesion of cells to the bone surface.
It can inhibit the production of OC, affect the mineralization of bone, and inhibit the production and promotion of BMP and TGF P.
The exertion of osteogenesis.
4.ALP and ACP changes in the weight bearing bone of ZW and 4W tail suspended rats.
Methods detection of ALP and ACP. in the femur showed that ALP in the femur of tail suspended rats was significantly lower than that in ACP rats.
The results showed that after simulated weightlessness, the osteogenesis of rats decreased and the osteoclast increased.
5. the morphological changes of the weight-bearing bone were observed by trichrome staining.
It showed that the calcification of bone matrix was impaired and the calcium deposition was decalcified.
Salt metabolism disorder.
In conclusion, cell culture was used to observe the reduction of bone marrow stromal cells by simulated weightlessness.
The effects of BMP and TGF P on bone marrow in vitro were observed by in situ hybridization.
OPN and OC changes in the weight loss of the bearing bone in simulated weightlessness; bone is observed by special staining.
Morphological changes of ALP and ACP in the bone and calcium metabolism in the bearing bone were further explored.
The mechanism of weightless bone loss is of reference significance.
【学位授予单位】：第四军医大学
【学位级别】：硕士
【学位授予年份】：2002
【分类号】：R852.22
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本文编号：2171180
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