Effects of mechanical loading on energy production of intervertebral disc cells

描述（由申请人提供）： 夏季椎间盘退变（IVD）与腰痛密切相关，腰痛在美国折磨着5200万人[21]。由于IVD是人体中最大的无血管组织，因此重要的营养物质（例如，氧气和葡萄糖）通过扩散和对流在长距离上通过致密的细胞外基质递送到IVD细胞。因此，营养供应不足被认为是椎间盘退变的潜在机制[3，5，35]。本研究的长期目标是：（1）了解椎间盘的机械生物学和椎间盘退变的机制;（2）开发避免或延缓椎间盘退变的策略。细胞消耗氧气和葡萄糖来产生能量，这是细胞基质合成的重要组成部分[61]，用于维持组织的完整性和防止组织变性。三磷酸腺苷（ATP）是IVD中主要通过糖酵解产生的主要能量形式[4]。在身体运动期间，IVD在骨性椎体之间传递大载荷。我们最近的理论研究表明，动态压缩通过增强氧和乳酸盐的运输促进IVD内的糖酵解[42，43]，这表明动态负荷可以促进IVD细胞的能量产生。此外，以前的研究表明，机械负荷改变了葡萄糖的膜转运和一氧化氮的产生[8，16，17，62]，这可能会内在地影响细胞能量的产生。本提案的主要目的是检查（1）动态和静态压缩对全椎间盘培养物中IVD细胞ATP产生的总体影响和（2）机械负荷对IVD细胞ATP产生的内在影响。提出以下假设：（1）动态负荷促进IVD细胞ATP生成、乳酸和氧的转运，而静态压缩则相反;（2）动静压对IVD细胞ATP生成有内在影响。为了检验这些假设，我们将（1）确定静态和动态压缩下IVD中ATP、氧气和乳酸盐的浓度（具体目标1）和（2）确定动态和静态压缩对IVD细胞ATP产生的内在影响（具体目标2）。为了实现这些特定的目标，我们将构建一个器官培养系统，为IVD提供体外培养环境和机械刺激。将进行生化测定以评估IVD中的乳酸盐和ATP浓度，同时将通过光学传感系统测定氧浓度。拟议研究的结果将提高我们对机械负荷下IVD中细胞能量代谢和营养转运的理解，并促进预防椎间盘退变的新策略的开发。 公共卫生相关性： 椎间盘退变（IVD）与腰痛密切相关，在美国有5200万人患有腰痛。营养供应不足被认为是椎间盘退变的潜在机制。拟议研究的结果将提高我们对机械负荷下IVD中细胞能量代谢和营养转运的理解，并促进预防椎间盘退变的新策略的开发。

DESCRIPTION (provided by applicant): SUMMERY Degeneration of intervertebral disc (IVD) is closely associated with low back pain which afflicts 52 million individuals in the United States [21]. Since IVD is the largest avascular tissue in the human body, vital nutrients (e.g., oxygen and glucose) are delivered by diffusion and convection over a long distance through dense extracellular matrix to IVD cells. Therefore, poor nutrient supply has been suggested as a potential mechanism for disc degeneration [3,5,35]. The long term goals of our research are: (1) to understand mechanobiology of intervertebral disc and the mechanisms of disc degeneration and (2) to develop strategies to avoid or retard disc degeneration. Cells consume oxygen and glucose to produce energy that is an essential component in cellular matrix synthesis [61] for maintaining the integrity of tissue and preventing tissue degeneration. Adenosine triphosphate (ATP) is the major energy form which is mainly generated through glycolysis in the IVD [4]. During body motion, IVDs transmit large loads between bony vertebral bodies. Our recent theoretical study demonstrated that dynamic compression promoted glycolysis within the IVD by enhancing the transport of oxygen and lactate [42,43], suggesting that energy production of IVD cells can be promoted extrinsically by dynamic loading. Furthermore, previous studies showed that mechanical loading altered membrane transport of glucose and production of nitric oxide [8,16,17,62] which may affect cellular energy production intrinsically. The major objectives of this proposal are to examine (1) the overall effects of dynamic and static compression on ATP production of the IVD cells in the whole disc culture and (2) the intrinsic effect of mechanical loading on ATP production of the IVD cells. The hypotheses are proposed as the followings: (1) Cellular ATP production and transport of lactate and oxygen are promoted in the IVD under dynamic loading whereas static compression exhibits the reverse effect; and (2) Dynamic and static compression influence ATP production of IVD cells intrinsically. To test these hypotheses, we will (1) determine the concentrations of ATP, oxygen, and lactate in the IVD under static and dynamic compression (Specific Aim 1) and (2) determine the intrinsic effects of dynamic and static compression on the ATP production of IVD cells (Specific Aim 2). To achieve these specific aims, we will construct an organ culture system to provide an in-vitro culture environment and mechanical stimuli for IVD. Biochemical assays will be performed to assess concentrations of lactate and ATP in the IVD while oxygen concentration will be determined by an optic sensing system. The outcomes of the proposed studies will improve our understanding of cellular energy metabolism and nutrient transport in the IVD under mechanical loading and facilitate the development of new strategies to prevent disc degeneration. PUBLIC HEALTH RELEVANCE: Degeneration of intervertebral disc (IVD) is closely associated with low back pain which afflicts 52 million individuals in the United States. Poor nutrient supply has been suggested as a potential mechanism for disc degeneration. The outcomes of the proposed studies will improve our understanding of cellular energy metabolism and nutrient transport in the IVD under mechanical loading and facilitate the development of new strategies to prevent disc degeneration.