碱性微环境成骨开关效应：动态环境中可降解生物活性材料表面微碱性梯度形成及成骨机制研究

The imbalance of bone homeostasis is the root cause of osteoporotic fracture. At present, there is still no bone repairing material for bone homeostasis reconstruction clinically. Under the premise of satisfying the biological safety, the material can not effectively restore the balance of bone metabolism, and the factors that regulate the balance between bone cells are still unclear. Therefore, on the basis of previous studies, our team found that the pH value plays an important role in the regulation of the balance between osteoclast and osteoblast, Our preliminary investigation indicated that the degradation of the material will generate a pH value that apparently different from the physiological condition. Thus, in this study, using non-invasive scanning ion selective electrode technique, the influence of surface flow on the establishment of a pH gradient from degradable material in the liquid under the dynamic environment will be studied. The pH’s effect on the adhesion, proliferation and differentiation of the osteoclasts, bone marrow stromal stem cells and osteoblasts during the "material - cell" interaction process will be evaluated. In addition, the movement direction and the flow rate of hydrogen ions on the cell surface will be detected. The present study will establish a pH “switch on/off” theory to better understand the regulation effect of acid-base equilibrium on the bone homeostasis, and will further define the boundary of the material microenvironment. By using osteoporosis bone defect animal model, the present study will evaluate the regeneration efficiency of that influenced by the implantation of materials with different microenvironment pHs. Therefore, this study will provide an important theoretical basis for the formation of a new generation of biodegradable biomaterials in vivo.

骨稳态失衡是引起骨质疏松性骨折的根本原因。目前，临床上尚无以骨稳态重建为目标的骨修复材料。材料在满足生物安全性的前提下，并不能有效的恢复骨代谢平衡，调控骨细胞间平衡的因素尚不明朗。因此，本团队在前期研究的基础上发现pH值在调控破骨与成骨细胞间平衡具有重要作用，并采用微电极初步探明材料降解有一个不同生理环境下pH值。进而，在本研究中将采用非损伤扫描离子选择电极技术，模拟生理动态液流下，考察多种可降解材料表面在液流动态环境下，离子及pH的梯度效应；明晰“材料-细胞”交互作用下，破骨细胞、骨髓基质干细胞和成骨细胞随pH值变化产生的黏附、增殖、及细胞间相互作用的能力，并进一步观测氢离子跨膜运输规律；建立pH开关效应调控骨稳态重建的理论，研究微环境实控边界效应，定义成骨微环境的区域范围，并在体研究材料对骨质疏松骨缺损的修复效果。因此，本课题将为新一代可降解生物材料在体内诱导成骨提供重要的理论依据。

骨稳态失衡是引起骨质疏松性骨折的根本原因。目前，临床上尚无以骨稳态重建为目标的骨修复材料。材料在满足生物安全性的前提下，并不能有效的恢复骨代谢平衡，调控骨细胞间平衡的因素尚不明朗。研究发现，通过设计细胞培养基中碳酸氢钠的浓度，实现了碱性pH微环境的调控。而碱性微环境一方面能抑制破骨细胞的活性，同时促进骨髓间充质干细胞的成骨分化。深入的研究表明，碱性微环境促进BMSC向成骨方向分化的机制可能在于，通过上调PYCR1与P5CS基因和蛋白的表达，促进了P5C与脯氨酸的合成，进而引起COL1A1的表达升高，实现hBMSCs细胞的成骨分化的调控。基于上述碱性微环境调控骨稳态的研究，项目团队基于玻璃结构网络设计技术，开发了组成和降解可控的生物活性玻璃。非损伤微测技术（NMT）表明，硼/硅比的设计可调控生物活性玻璃的化学活性和碱性微环境形成，从而实现理想成骨微环境的构建。基于骨质疏松大鼠骨折微环境的修复结果表明，材料在体内降解形成的碱性pH微环境，能有效抑制破骨细胞活性和破骨相关基因的表达，从而有效促进早期成骨。基于生物活性玻璃的碱性和离子微环境促进成骨的研究，项目团队进一步构建了多孔生物活性支架，实现了如兔子股骨缺损与兔子胫骨节段性等不同骨缺损微环境的修复重建。最后，项目团队选取了能营造碱性pH微环境（生物活性玻璃），中性pH微环境（PMMA）和酸性pH微环境（PLGA）等材料，并植入骨质疏松大鼠股骨中段的临界尺寸骨缺损。结果表明研究，碱性微环境材料（生物活性玻璃）能显著促进缺损部位的体内矿化和新骨形成。基于上述研究，项目执行期间，共发表SCI论文6篇，中文期刊1篇；同时申请专利2项。因此项目执行过程中，完成了立项所设定的研究内容和研究目标。