Vitamin D regulation of metalloproteinase activity in matrix vesicles
Vitamin D regulation of metalloproteinase activity in matrix vesicles
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DOI：
10.3109/03008209609029208
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发表时间：
1996-01-01
影响因子：
2.9
通讯作者：
Boyan, BD
中科院分区：
文献类型：
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作者：
Dean, DD;Schwartz, Z;Boyan, BD
基质囊泡（MVs）富含能够降解蛋白聚糖的基质金属蛋白酶（MMPs）。本研究的目的是确定哪些MMPs存在于mv中，并确定这些MMPs是否受1,25-(OH)(2)D-3[1,25]和24,25-(OH)(2)D-3[24,25]的调节。为此，从大鼠肋软骨中分离生长区（GC）和休止区（RC）软骨细胞并置于培养中。合流时，GCs用1、25治疗，RCs用24、25治疗24小时。然后从培养物中收集mv，质膜（pm）和条件培养基。RT-PCR结果显示，RCs和GCs中均存在基质溶素-1和72 kDa明胶酶mRNA。酪蛋白酶谱分析显示，在MV中，酪蛋白活性为M(r) 48和28 kDa，而在PM和条件培养基中没有活性；Western分析证实该活性为stromelysin-1。在mv中也发现了低水平的明胶溶解活性，但在pm或条件培养基中没有发现。当使用蛋白多糖球测定酶活性时，发现GCs和RCs都产生含有中性金属蛋白酶的mv和pm。这两个细胞也产生含有纤溶酶原激活剂的mv和pm。在GCs中添加1,25可使MVs的金属蛋白酶活性显著增加4- 5倍，而pm则没有。相比之下，24、25处理的RCs培养物的mv含有降低的金属蛋白酶活性；PMs酶活性不受24,25的影响。24、25处理后，RC的MV酶活性升高，而1、25处理后，GC的MV酶活性降低。这项研究表明，RCs和GCs都产生基质溶素-1和72 kDa明胶酶，这些酶优先定位于MVs。此外，MMP和纤溶酶原激活剂在MVs和pm中的活性受维生素D代谢产物的调节。
Matrix vesicles (MVs) are enriched in matrix metalloproteinases (MMPs) capable of degrading proteoglycans. The aim of the present study was to identify which MMPs are present in MVs and determine whether these MMPs are regulated by 1,25-(OH)(2)D-3 [1,25] and 24,25-(OH)(2)D-3 [24,25]. To do this, growth zone (GC) and resting zone (RC) chondrocytes were isolated from rat costochondral cartilage and placed into culture. At confluence, GCs were treated with 1,25 and RCs with 24,25 for 24 hours. MVs, plasma membranes (PMs), and conditioned media were then collected from the cultures. RT-PCR demonstrated the presence of mRNA for stromelysin-1 and 72 kDa gelatinase in both RCs and GCs. Casein zymography revealed activity at M(r) 48 and 28 kDa in MV, but not PM or conditioned media; Western analysis confirmed that this activity was stromelysin-1. Gelatinolytic activity, at low levels, was also found in MVs, but not PMs or conditioned media. When enzyme activity was measured using a proteoglycan bead assay, it was found that both GCs and RCs produced MVs and PMs containing neutral metalloproteinase. Both cells also produced MVs and PMs containing plasminogen activator. The addition of 1,25 to GCs caused a significant 4- to 5-fold increase in metalloproteinase activity in MVs, but not PMs. In contrast, MVs from cultures of RCs treated with 24,25 contained decreased metalloproteinase activity; enzyme activity in PMs was unaffected by 24,25. Plasminogen activator in MVs from RC was increased by treatment with 24,25, while MV enzyme activity was decreased after treatment of GC cultures with 1,25. This study shows that both RCs and GCs produce stromelysin-1 and 72 kDa gelatinase and that these enzymes are preferentially localized in MVs. Further, MMP and plasminogen activator activities in MVs and PMs are regulated by vitamin D metabolites.