ADULT STEM CELL SUPPRESSION DUE TO ALCOHOL INDUCED MICROENVIRONMENT ALTERATIONS

描述（由申请人提供）：酒精中毒和酒精滥用影响至少1400万美国人。长期饮酒与骨质疏松症和腰椎骨折率增加有关。后外侧脊柱融合术是酒精性骨质疏松症常见腰椎压缩性骨折的标准治疗方法。长期酒精暴露会抑制骨折愈合，导致脊柱融合失败的发生率显著升高。2003年，与脊柱融合翻修手术相关的费用高达72 000美元。再生医学是一种新的治疗方式，采用干细胞来促进组织愈合。脂肪组织来源的基质细胞（ASCs）在体内和体外均能促进成骨。干细胞和微环境之间的复杂相互作用是增强骨生成所必需的，而慢性酒精暴露的改变会限制它们的综合功效。基于这一认识，我们假设：（A）慢性酒精摄入降低了ASC的数量、扩张速度和胸膜潜能;（B）接受者的慢性酒精暴露抑制或阻止了在合适的生物材料载体中应用正常ASC加速脊柱融合;和（C）从慢性酒精暴露的受试者中收获并应用于合适的生物材料载体中的ASC不会加速正常受试者的脊柱融合。这些假设将通过以下具体目标进行检验：目标1。在大鼠模型中确定慢性酒精摄入对ASCs的影响。将对从正常大鼠和暴露于已建立的慢性酒精摄入模型的大鼠中采集的细胞进行研究。将使用标准程序定量体外扩增和传代的干细胞的数量和行为。目标二。确定正常ASCs对慢性酒精暴露和非酒精暴露大鼠脊柱融合的影响。将从无酒精暴露的受试者中收获的ASC植入有和无慢性酒精暴露的受试者中进行研究。结果测量的射线照相，显微CT，RT-PCR，成分分析，组织学和免疫组织化学将提供周围的正常干细胞的能力，以诱导成骨在正常微环境与慢性酒精暴露改变微环境的信息。目标3。确定从慢性酒精暴露大鼠中收获的ASCs对有和无慢性酒精暴露大鼠脊柱融合的影响。将从长期酒精暴露的受试者中收获的ASC植入有和无长期酒精暴露的受试者中进行研究。与目标2相同的结果测量将提供关于慢性酒精暴露改变的干细胞在正常微环境中诱导成骨的能力与慢性酒精暴露改变的微环境的信息。这项研究的结果将大大推进酒精诱导的干细胞和基质改变抑制骨形成的知识。这项研究的结果将大大有助于目前的知识酒精诱导的干细胞和基质的变化，抑制骨形成。公共卫生相关性：阐明慢性酒精滥用导致脊柱融合失败的病理生理学将为解决这一重要的医学问题提供机制。此外，ASC应用是一种有前途的治疗方法，可以加速与酒精暴露无关的天然成骨细胞前体数量减少的患者的骨折愈合。

DESCRIPTION (provided by applicant): Alcoholism and alcohol abuse affect at least 14 million Americans. Chronic alcohol consumption is associated with osteoporosis and an increased rate of lumbar spinal fractures. Posterolateral spinal fusion is the standard treatment for lumbar compression fractures common in ethanol-induced osteoporosis. Chronic alcohol exposure inhibits fracture healing and results in a significantly higher incidence of spinal fusion failures. Costs associated with revision spinal fusion surgery were as high as $72,000 each in 2003. Regenerative medicine is a novel therapeutic modality that employs stem cells to promote tissue healing. Adipose tissue-derived stromal cells (ASCs) promote osteogenesis both in vivo and in vitro. Complex interactions between stem cells and the microenvironment are necessary to enhance osteogenesis, and alterations in either from chronic alcohol exposure can limit their combined efficacy. Based on this knowledge we postulate: (A) That chronic alcohol ingestion reduces the number, rate of expansion, and pleuripotential capacity of ASCs; (B) That chronic alcohol exposure in the recipient inhibits or prevents the acceleration of spinal fusion by application of normal ASCs in a suitable biomaterial carrier; and (C) That ASCs harvested from subjects with chronic alcohol exposure and applied in a suitable biomaterial carrier do not accelerate spinal fusion in normal recipients. These hypotheses will be tested with the following Specific Aims: Aim 1. To determine the effect of chronic alcohol ingestion on ASCs in a rat model. Studies will be performed on cells harvested from normal rats and rats exposed to an established model of chronic alcohol ingestion. The number and behavior of stem cells expanded and passaged in vitro will be quantified with standard procedures. Aim 2. To determine the effect of normal ASCs on spinal fusion in rats with and without chronic alcohol exposure. Studies will be performed with ASCs harvested from subjects with no alcohol exposure implanted into subjects with and without chronic alcohol exposure. Outcome measures of radiographs, micro-CT, RT-PCR, compositional analysis, histology, and immunohistochemistry will provide information surrounding the ability of normal stem cells to induce osteogenesis in normal microenvironments versus microenvironments altered by chronic alcohol exposure. Aim 3. To determine the effect of ASCs harvested from rats with chronic alcohol exposure on spinal fusion in rats with and without chronic alcohol exposure. Studies will be performed with ASCs harvested from subjects with chronic alcohol exposure implanted into subjects with and without chronic alcohol exposure. Outcome measures identical to those of Aim 2 will provide information surrounding the ability of stem cells altered by chronic alcohol exposure to induce osteogenesis in normal versus microenvironments altered by chronic alcohol exposure. Results from this investigation will substantially advance knowledge surrounding alcohol-induced stem cell and matrix alterations that inhibit bone formation. Results from this investigation will contribute substantially to current knowledge surrounding alcohol-induced stem cell and matrix alterations that inhibit bone formation. PUBLIC HEALTH RELEVANCE: Elucidation of the pathophysiology contributing to failure of spinal fusion in the face of chronic alcohol abuse will provide mechanisms to address this important medical problem. Additionally, ASC application is a promising treatment that may accelerate fracture healing in patients with reduced numbers of native osteoblast precursors unrelated to alcohol exposure.