Martínez P, Esbrit P, Rodrigo A, Alvarez-Arroyo M V, Martínez M E
Biochemistry Division, Hospital La Paz, Madrid, Spain.
Osteoporos Int. 2002 Nov;13(11):874-81. doi: 10.1007/s001980200120.
Osteogenesis and angiogenesis occur in a coordinated manner in skeletal tissue, so that impaired angiogenesis is associated with decreased bone formation in aged subjects. However, the interaction between bone endothelium and osteoblastic cells is poorly understood. Parathyroid hormone-related protein (PTHrP), a bone factor which modulates osteoblastic cell growth and/or differentiation, stimulates vascular endothelial growth factor (VEGF), a potent angiogenic factor, in primary cultures of human osteoblastic (hOB) cells. In the present study, we examined the age-related changes of both factors in these cells. Human OB cells were isolated from trabecular bone samples from knee or hip explants obtained from 45 osteoarthritic patients: 12 <60 years (21-59 years), 5 women and 7 men, and 33 >60 years (61-82 years), 20 women and 13 men. Cell total RNA was isolated, and mRNA analysis was performed by reverse transcription-polymerase chain reaction. Relative ratios of amplified products with respect to glyceraldehyde-3-phosphate dehydrogenase were then calculated. PTHrP and VEGF were measured in the cell-conditioned medium, after stimulation with (or without) 10 nM 1,25(OH)(2)D(3) for 72 h, using specific immunoradiometric assay and a competitive immunoassay, respectively. A positive correlation was found between PTHrP and VEGF (both mRNA and secreted protein), and also between PTHrP mRNA and the secreted protein levels, in these cells. PTHrP, both mRNA and protein secretion levels, and VEGF secreted values were higher in knee hOB cells than in hip hOB cells only in the younger group. In addition, a decrease in the secreted levels of these factors occurs with aging only in hOB cells from knee. Treatment with 10 nM 1,25(OH)(2)D(3) induced a lower inhibitory response of PTHrP secretion, and a higher stimulatory response of secreted VEGF, in hOB cells with age. These findings indicate that age-related bone loss in humans is associated with a decrease in the osteoblastic secretion of both PTHrP and VEGF in the knee, a predominantly trabecular bone. These data might provide a rationale to explain the impaired angiogenesis associated with trabecular bone loss in aging.
骨生成和血管生成在骨骼组织中以协调的方式发生，因此血管生成受损与老年受试者骨形成减少有关。然而，骨内皮细胞与成骨细胞之间的相互作用尚不清楚。甲状旁腺激素相关蛋白（PTHrP）是一种调节成骨细胞生长和/或分化的骨因子，在人成骨（hOB）细胞原代培养物中可刺激血管内皮生长因子（VEGF），一种强效血管生成因子。在本研究中，我们检测了这些细胞中这两种因子的年龄相关变化。从45例骨关节炎患者的膝关节或髋关节外植体的小梁骨样本中分离出人OB细胞：12例年龄<60岁（21 - 59岁），5名女性和7名男性，以及33例年龄>60岁（61 - 82岁），20名女性和13名男性。分离细胞总RNA，并通过逆转录 - 聚合酶链反应进行mRNA分析。然后计算相对于甘油醛 - 3 - 磷酸脱氢酶的扩增产物的相对比率。在用（或不用）10 nM 1,25(OH)₂D₃刺激72小时后，分别使用特异性免疫放射分析和竞争性免疫分析在细胞条件培养基中测量PTHrP和VEGF。在这些细胞中，发现PTHrP与VEGF（mRNA和分泌蛋白两者）之间以及PTHrP mRNA与分泌蛋白水平之间呈正相关。仅在较年轻组中，膝关节hOB细胞中的PTHrP、mRNA和蛋白分泌水平以及VEGF分泌值均高于髋关节hOB细胞。此外，仅在膝关节的hOB细胞中，这些因子的分泌水平随衰老而降低。用10 nM 1,25(OH)₂D₃处理后，hOB细胞中PTHrP分泌的抑制反应随年龄增长而降低，而分泌的VEGF的刺激反应随年龄增长而升高。这些发现表明，人类与年龄相关的骨质流失与膝关节（主要是小梁骨）中PTHrP和成骨细胞分泌的VEGF减少有关。这些数据可能为解释衰老过程中小梁骨丢失相关的血管生成受损提供理论依据。
