Effects of Advanced Glycation Endproducts on Type 2 Diabetic and Fragility Fractures

摘要 骨折极大地增加了医疗费用，影响了生活质量。临床上，骨折风险 可通过双X射线骨密度仪(DXA)或骨折风险评估(FRAX)工具进行预测。因为 2型糖尿病(T2D)患者表现出高骨密度(BMD)，这两种工具都无法正确预测 骨折风险，导致糖尿病受试者脆性骨折显著增加。因此，有一个 需要研究骨骼中胶原和其他有机成分的修饰如何预测 糖尿病骨折。戊苷(PEN)，一种荧光高级糖基化终产物(AGE)，形成于 骨骼通过糖和蛋白质之间的反应，是唯一确定的骨骼脆性的标志。然而，它 不能始终如一地预测T2D和脆性骨折。在这里，我们第一次在骨骼中演示 存在羧甲基赖氨酸(CML)，这是一种非荧光糖氧化年龄，并提出了一种技术来 量一量。我们发现，它在骨骼中大量形成，并与骨骼韧性的丧失高度相关。 我们证明，与其他年龄相比，CML在T2D人骨中上调了60%。 与他们的年龄匹配的对照组。然后我们提供证据证明慢性粒细胞白血病促进了 额外的羟基磷灰石(HA)晶体，类似于人类的T2D条件，形成了一个分子链接 骨骼的有机成分和无机成分(胶原-HA界面)之间的相互作用会损害骨骼质量。我们的 总体假设是CML是一种新的和相关的T2D骨折的生物标志物，它捕捉到了 高血糖和氧化应激对骨骼的影响，并解释了骨骼对 T2D中的骨折。使用肥胖和非肥胖的T2D小鼠模型，模拟因果关系和 人类T2D对骨骼的影响，我们提供了T2D增加年龄的证据，CML解释了骨骼 脆弱。同样，使用健康、老龄化和身体成分(ABC)研究的数据，我们表明 较高的血清CML水平与T2D患者发生临床骨折的风险增加相关， 不受BMD影响。因此，我们的总体目标是将CML建立为一种新的和相关的骨生物标志物 并确定其对T2D患者骨脆性的影响。使用体外模型，体外人类 身体组织，肥胖和非肥胖的体内小鼠模型，以及来自Health的现有数据 ABC研究我们将追求的目标有：目标1：为促进CML的形成创造NEG条件 并确定其降低骨骼能量消耗的机制(S)；目的2： 确定慢性粒细胞白血病和其他年龄在骨基质改变和能量消耗中的作用 来自髋部骨折患者的人类T2D椎骨、皮质和松质骨；以及目标3：验证CML 作为T2D骨骼脆性的生物标志物，并建立其与高血糖和氧化应激的关联。 我们的发现将对慢性粒细胞白血病和其他AGE的作用机制和影响提供新的理解。 骨折导致了预测、管理和缓解T2D和脆性骨折的新策略。

Abstract Bone fractures contribute significantly to healthcare cost affecting the quality of life. Clinically, fracture risk can be predicted by dual x-ray absorptiometry (DXA) or the fracture risk assessment (FRAX) tool. Because type 2 diabetes (T2D) patients exhibit high bone mineral density (BMD), both tools fail to correctly predict fracture risk, leading to a significant increase of fragility fractures in diabetic subjects. Therefore, there is a need to investigate how modifications in collagen and other organic components in bone can predict diabetic fractures. Pentosidine (PEN), a fluorescent Advanced Glycation Endproduct (AGE) that forms in bone by reaction between sugars and proteins, is the only established marker of bone fragility. However, it does not consistently predict T2D and fragility fractures. Here, for the first time in bone, we demonstrate the presence of carboxymethyl-lysine (CML), a non-fluorescent glycoxidative AGE, and present a technique to measure it. We show that it forms in abundance in bone and is highly correlated to loss of bone toughness. We demonstrate that, in contrast to other AGEs, CML is upregulated >60% in T2D human bone compared to their age-matched controls. We then provide evidence that CML promotes formation and growth of additional hydroxyapatite (HA) crystals, similar to human T2D condition, and forms a ‘molecular link’ between the organic and inorganic components of bone (collagen-HA interface) impairing bone quality. Our overall hypothesis is that CML is a ‘new and relevant’ biomarker of T2D fracture that captures the effects of both hyperglycemia and oxidative stress in bone and explains the susceptibility of bone to fracture in T2D. Using an obese and a non-obese mouse model of T2D, that mimic both causality and impact of human T2D on bone, we provide evidence that T2D increases AGEs, with CML explaining bone fragility. Similarly, using data from the Health, Aging and Body Composition (ABC) study we show that higher serum CML levels are associated with increased risk of incident clinical fractures in T2D, independent of BMD. Thus, our overall goal is to establish CML as a new and relevant biomarker of bone fragility and determine how it contributes to bone fragility in T2D. Using in vitro models, ex vivo human cadaveric tissue, in vivo mouse models of obese and non-obese T2D, and existing data from the Health ABC study we will pursue there aims: Aim 1: Establish NEG conditions for enhanced formation of CML over other AGEs and determine the mechanism(s) by which it reduces energy dissipation in bone; Aim 2: Determine the contribution of CML and other AGEs to alterations in bone matrix and energy dissipation in human T2D vertebrae and cortical and cancellous bone from hip fracture patients; and Aim 3: Validate CML as a biomarker of T2D bone fragility and establish its association with hyperglycemia and oxidative stress. Our findings will provide a new understanding of the mechanism and the effects of CML and other AGEs on bone fractures leading to new strategies to predict, manage and mitigate T2D and fragility fractures.