INHIBITED INTRAMEMBRANEOUS BONE HEALING IN DIABETES

描述（由申请人提供）：糖尿病并发症的部分原因是血糖水平升高。这导致蛋白质的非酶糖基化形成晚期糖基化终产物（AGE's）。AGE在糖尿病的许多并发症中起着重要作用。骨质减少、低矿物质密度和骨骼脆弱是1型糖尿病的并发症，被称为“糖尿病性骨病”。令人惊讶的是，我们对AGE在调节骨愈合、骨形成和成骨细胞功能中的作用知之甚少。例如，晚期糖基化终产物受体（RAGE）的存在和活性尚未在成骨细胞中报道。提出的研究的主要假设是AGE通过结合和激活成骨细胞中的RAGE来抑制骨愈合和形成。我们认为这导致NFKB激活，转录抑制和下调关键成骨细胞生长因子和细胞外基质基因。

DESCRIPTION (provided by applicant): Complications of diabetes result in part from elevated serum glucose levels. This leads to non-enzymatic glycation of proteins to form advanced glycation end products (AGE's). AGE's play a significant role in many complications of diabetes. Osteopenia and low mineral density and weak bones is a complication of Type 1 diabetes and is known as "diabetic bone disease". Surprisingly little is known regarding the role of AGE's in modulating bone healing, bone formation, and osteoblast function. For example, the presence and activity of the receptor for advanced glycation end products (RAGE) has not been reported in osteoblasts. The principal hypothesis of the proposed research is that AGE's inhibit bone healing and formation by binding and activating RAGE in osteoblasts. We propose that this results in NFKB activation and transcriptional repression and down-regulation of key osteoblast growth factors and extracellular matrix genes. Aim 1 will measure in vivo the expression of selected growth factors and extracellular matrix products (BMP-1, BMP- 2, BMP-4, and type I collagen) in healing calvaria defects made in diabetic and non-diabetic mice. Diabetes will be induced by the multiple low dose streptozotocin protocol in Balb/c mice; selected studies will be performed in the nonchemically-induced murine diabetic model (NOD strain). The degree of inhibition of bone healing in diabetic animals and expression patterns of RAGE will be determined by quantitative histomorphometric and quantitative immunohistochemical measurements. Studies will directly determine the role of AGE's in diminished diabetic bone formation by local application of AGE's to calvaria defects in non-diabetic mice. The degree to which this mimics diabetic bone will be determined by measuring inhibition of healing and regulation of the same growth factors and extracellular matrix products. Aim 2 will determine in vitro in primary rat osteoblast cell cultures that AGE's inhibit production of osteoblast growth factors and type I collagen via RAGE activated NF-KB. RAGE function blocking antibody studies will identify the AGE/RAGE-dependent NF-KB activation mechanism in the regulation of the specified osteoblast genes. The role of NF-KB activation in down-regulating target osteoblast genes will be directly determined by transfection with the super-repressor 32A/36A IKB-alpha, a potent and specific inhibitor of NF-KB activation. These studies will identify a new mechanism that contributes to diabetic bone disease; and should lead to the identification of new therapeutic treatment targets for this increasingly prevalent clinical condition.