Progesterone stimulates osteoprogenitor proliferation and differentiation in osteoblast-like cell populations derived from adult female rat vertebrae

黄体酮刺激源自成年雌性大鼠椎骨的成骨细胞样细胞群中的骨祖细胞增殖和分化

• 批准号: 15591579
• 负责人: ISHIDA Yoichiro
• 金额: $ 2.18万
• 依托单位: Yamaguchi University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15591579/
• 关键词: Osteoprogenitor; Progesterone; Dexamethasone; Bone formation; Aging; Estrogen; 骨芽細胞; 骨形成能

A significant contribution to the bone loss associated with aging is likely to be a decline in bone formation. We have characterized and compared the number, capacity for proliferation and differentiation and the self-renewal ability of osteoprogenitors of aged (24-week-old) and young (6-week-old) female Wistar rats using limiting dilution analyses and continuous subculture experiments. Cells were obtained from outgrowths of explants of lumbar vertebrae (L1-L6) and grown in α-minimal essential medium (α-MEM), 10% FBS and 50mg/ml ascorbic acid with or without dexamethasone (Dex) or progesterone (Prog). Growth curves for cell populations of both age groups were similar with population doubling times of 28.3 and 27.5 h for the aged and young animals, respectively. Osteoprogenitors from both age groups formed bone nodules when cultured in the presence of either Dex or Prog. Limiting dilution analysis in the presence of 10 nM Dex showed no difference between the aged and young rats in the n … More umber of colony forming units-fibroblast (CFU-F), alkaline phosphatase-positive colony forming units-fibroblast (AP+CFU-F) or colony forming units-osteoblast (CFU-O). Limiting dilution analysis in the presence of 3 mM Prog showed no differences in the numbers of CFU-F, AP+CFU-F or CFU-O between the aged and young groups. Frequencies and numbers of both progenitor types were determined for up to six subcultures using continuous subculturing, limiting dilution analysis, and colony assays. In Dex-containing medium, subculturing resulted in a significant increase in the total number of Dex-responsive osteoprogenitors in second subculture cells over first subculture cells without a significant increase in the frequency of these progenitors. From the third subculture onward, the frequency of osteoprogenitors decreased in a linear manner and none were observed after six subcultures. Similar results were obtained in Prog-containing medium. Results indicate that the cell population doubling times, growth characteristics, and the number of CFU-F and osteoprogenitors in vertebral bone cell populations from aged rats and young rats are similar. This suggests that the bone loss associated with aging is not caused by a decrease in osteoprogenitor cell number. However, cell populations from the aged rats showed a reduced capacity for self-renewal in vitro, which would ultimately translate into a reduced number of osteoblasts and might be partly responsible for a decrease in bone formation in aged animals.Previous experiments have demonstrated that bone cell populations derived from explants of lumbar vertebral bone of adult female rats contain osteoprogenitors that require Dex or Prog to proliferate and differentiate into fully differentiated bone-forming osteoblasts. We have shown that the Prog-dependent population can not be detected in male rats after sexual maturation but is present in prepubertal rats of both sexes, and can be induced in adult male-derived populations by culturing the explants in media containing 17 β-estradiol (10^<-9>-10^<-8>M). This suggested that the Frog- and Dex- dependent osteoprogenitors in adult female-derived populations were likely to be distinct populations and that the survival of the Prog-dependent osteoprogenitors and/or their ability to proliferate is dependent on the presence of estrogen. The results imply that Prog plays an important role in maintaining bone mass through regulating the class of osteoprogenitors responsive to Prog. Less

与衰老相关的骨丢失的一个重要原因可能是骨形成的减少。采用极限稀释法和连续传代实验，对老年(24周龄)和年轻(6周龄)雌性Wistar大鼠的骨祖细胞的数量、增殖分化能力和自我更新能力进行了表征和比较。取L1~L6腰椎外植体，在α-MEM+10%胎牛血清+50 mg/ml抗坏血酸+地塞米松(Dex)或孕酮(Prog)的条件下培养。两个年龄组的细胞群体生长曲线相似，老年和幼龄动物的群体倍增时间分别为28.3h和27.5h。两个年龄组的骨祖细胞在地塞米松或Prog存在下培养时形成骨结节。在10 nM地塞米松存在下的极限稀释分析表明，老年大鼠和青年大鼠的n…没有差异更多的集落形成单位为成纤维细胞(CFU-F)、碱性磷酸酶阳性集落形成单位(AP+CFU-F)或集落形成单位-成骨细胞(CFU-O)。3 mM Prog对CFU-F、AP+CFU-F和CFU-O的极限稀释分析显示，老年组和青年组CFU-F、AP+CFU-F和CFU-O的数量无差异。通过连续继代培养、有限稀释分析和菌落分析，确定了最多六种继代培养的两种祖细胞类型的频率和数量。在含地塞米松的培养液中，继代培养的第二代细胞中对地塞米松敏感的成骨细胞总数比第一代细胞显著增加，但这些祖细胞的出现频率并没有显著增加。从第3次传代开始，成骨细胞的频率呈线性下降，传代6次后均未观察到成骨细胞。在含有Prog的介质中也得到了类似的结果。结果表明，老年大鼠和青年大鼠椎骨细胞群体的细胞倍增时间、生长特性、CFU-F和骨祖细胞的数量相似。这表明与衰老相关的骨丢失不是由骨祖细胞数量的减少引起的。然而，老年大鼠的细胞群体在体外自我更新的能力下降，最终转化为成骨细胞的数量减少，这可能是老年动物成骨减少的部分原因。先前的实验证明，成年雌性大鼠腰椎外植体来源的骨细胞群体含有成骨细胞，需要Dex或Prog增殖并分化为完全分化的成骨细胞。我们发现在性成熟后的雄性大鼠性成熟后不能检测到Prog依赖的群体，但在青春期前的大鼠中存在这种群体，并且通过在含有17β-雌二醇的培养基中培养成年雄性来源的群体可以诱导出依赖于Prog的群体。这表明，在成年雌性来源的群体中，依赖于青蛙和地塞米松的骨祖细胞可能是不同的群体，依赖于Prog的骨祖细胞的存活和/或其增殖能力依赖于雌激素的存在。结果提示，Prog通过调节Prog应答的骨祖细胞种类，在维持骨量方面发挥重要作用。较少

项目成果

Comparative efficacy of therapies for postmenopausal osteoporosis : results of the Yamaguchi Osteoporosis Prevention Study (YOPS).

绝经后骨质疏松症治疗方法的比较疗效：山口骨质疏松症预防研究 (YOPS) 的结果。

• 发表时间: 2004
• 期刊: Calcified Tissue International 74
• 作者: Yoichiro Ishida;Yoichiro Ishida;Yoichiro Ishida
• 通讯作者: Yoichiro Ishida

Affecting ambulatory status and survival prognosis of patients 90 years and older with hip fractures

影响90岁及以上髋部骨折患者的卧床状态和生存预后

• 发表时间: 2005
• 期刊: Clinical Orthopaedics (in press)
• 作者: Yoichiro Ishida

Comparative efficacy of hormone replacement therapy, etidronate, calcitonin, alfacalcidol, and vitamin K in postmenopausal women with osteoporosis : The Yamaguchi Osteoporosis Prevention Study.

激素替代疗法、依替膦酸、降钙素、阿法骨化醇和维生素 K 对患有骨质疏松症的绝经后妇女的比较疗效：山口骨质疏松症预防研究。

• 发表时间: 2004
• 期刊: American Journal of Medicine 117
• 作者: Yoichiro Ishida;Yoichiro Ishida
• 通讯作者: Yoichiro Ishida

Comparative efficacy of hormone replacement therapy, etidronate, calcitonin, alfacalcidol, and vitamin K in postmenopausal women with osteoporosis : The Yamaguchi Osteoporosis Prevention Study

激素替代疗法、依替膦酸、降钙素、阿法骨化醇和维生素 K 对患有骨质疏松症的绝经后妇女的疗效比较：山口骨质疏松症预防研究

• 发表时间: 2004
• 期刊: American Journal of Medicine 117
• 作者: Yoichiro Ishida;Yoichiro Ishida;Yoichiro Ishida;Yoichiro Ishida
• 通讯作者: Yoichiro Ishida

Comparative efficacy of therapies for postmenopausal osteoporosis : results of the Yamaguchi Osteoporosis Prevention Study (YOPS)

绝经后骨质疏松症治疗方法的比较疗效：山口骨质疏松症预防研究 (YOPS) 的结果

• 发表时间: 2004
• 期刊: Calcified Tissue International 74
• 作者: Yoichiro Ishida;Yoichiro Ishida;Yoichiro Ishida;Yoichiro Ishida;Yoichiro Ishida
• 通讯作者: Yoichiro Ishida