Extracts of Actinobacillus actinomycetemcomitans induce apoptotic cell death in human osteoblastic MG63 cells
Extracts of Actinobacillus actinomycetemcomitans induce apoptotic cell death in human osteoblastic MG63 cells
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DOI：
10.1177/00220345990780030501
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发表时间：
1999-03-01
影响因子：
7.6
通讯作者：
Haneji, T
中科院分区：
文献类型：
--
作者：
Morimoto, Y;Morimoto, H;Haneji, T
牙周病致病细菌的细胞外成分是否会诱导骨相关细胞凋亡尚不清楚。为了研究从超声处理的伴放线放线杆菌和中间普雷沃氏菌中获得的提取物对成骨细胞的影响，我们在这些提取物的存在下培养人成骨细胞系 MG63 和 Saos-2 细胞以及小鼠成骨细胞系 MC3T3-E1 细胞。添加来自伴放线放线杆菌的提取物在0.1至10μg/mL的浓度范围内以剂量和时间依赖性方式诱导MG63细胞中的细胞死亡。相比之下，即使存在 10 μg/mL 蛋白质，中间普雷沃氏菌提取物也不会诱导这些细胞死亡。通过使用Hoechst 33342染色技术，我们观察到用Actinobacillus actinomycetemcomitans提取物处理的MG63细胞中明显的核浓缩和染色质断裂。在用放线放线杆菌伴生提取物处理的 MG63 细胞中也检测到了 DNA 梯的形成，这是细胞凋亡的标志。在 MG63 细胞中，DNA 梯的形成具有剂量依赖性，浓度为 10 μg/mL 时效果最大，并且具有时间依赖性（12 至 48 小时）。然而，中间普雷沃氏菌提取物不会在 MG63、Saos-2 或 MC3T3-E1 细胞中诱导 DNA 断裂。 Actinobacillus actinomycetemcomitans 的提取物不会诱导 Saos-2 和 MC3T3-E1 细胞的细胞死亡和 DNA 断裂。经过热和胰蛋白酶处理的伴放线放线杆菌的超声提取物不会诱导 MG63 细胞中 DNA 梯的形成，表明凋亡诱导因子是蛋白质。放线菌酰亚胺以剂量依赖性方式阻止放线杆菌伴放线菌诱导的 MG63 细胞中 DNA 阶梯的形成，表明新基因转录和蛋白质合成在放线杆菌伴放线菌诱导的 MG63 细胞凋亡中受到调节。我们的结果表明，伴放线放线杆菌诱导的牙槽骨细胞凋亡在牙周疾病中发挥重要作用。
Whether an extracellular component of periodontal-disease-causing bacteria induces apoptotic cell death in bone-related cells is unknown. To study the effects on osteoblasts of extracts obtained from sonicated Actinobacillus actinomycetemcomitans and Prevotella intermedia, we cultured human osteoblastic cell lines MG63 and Saos-2 cells and mouse osteoblastic cell line MC3T3-E1 cells in the presence of such extracts. The addition of the extracts from Actinobacillus actinomycetemcomitans induced cell death in MG63 cells in a dose- and time-dependent fashion over the concentration range of 0.1 to 10 mu g/mL. By contrast, the extracts from Prevotella intermedin did not induce cell death in these cells, even in the presence of 10 mu g/mL protein. By using the Hoechst 33342 staining technique, we observed marked nuclear condensation and fragmentation of chromatin in MG63 cells treated with the extracts of Actinobacillus actinomycetemcomitans. DNA ladder formation, a hallmark of apoptosis, also was detected in MG63 cells treated with extracts from Actinobacillus actinomycetemcomitans. In MG63 cells, DNA ladder formation was dose-dependent with a maximal effect at a concentration of 10 mu g/mL, and time-dependent, from 12 to 48 hrs. However, the extracts from Prevotella intermedia did not induce DNA fragmentation in MG63, Saos-2, or MC3T3-E1 cells. The extracts from Actinobacillus actinomycetemcomitans did not induce cell death and DNA fragmentation in Saos-2 and MC3T3-E1 cells. Sonicated extracts of Actinobacillus actinomycetemcomitans that had been treated with heat and trypsin did not induce DNA ladder formation in MG63 cells, suggesting that the apoptosis-inducing factors are proteinaceous. Cycloheximide prevented the Actinobacillus actinomycetemcomitans-induced DNA ladder formation in MG63 cells in a dose-dependent fashion, suggesting that new gene transcription and protein synthesis are regulated for Actinobacillus actinomycetemcomitans-induced apoptosis in MG63 cells. Our results indicate that apoptosis in alveolar bone cells induced by Actinobacillus actinomycetemcomitans plays an important role in periodontal diseases.