脂多糖诱导成牙本质细胞凋亡模型的建立与评价

Apoptosis model of odontoblasts induced by lipopolysaccharide

目的：建立并评价脂多糖（LPS）诱导的成牙本质细胞（OB）凋亡模型。方法：采用梯度LPS（0、0．1、1、10、100tzg／mL）刺激法诱导小鼠OB细胞系MDPC-23细胞凋亡；分别于诱导后24、48、72、96h各时间点，以MTT法检测细胞增殖活力，流式细胞术检测细胞凋亡比例，确定合适的诱导浓度和时间后，建立OB细胞凋亡模型，并通过Hoechst染色观察细胞形态和凋亡比例。结果：0．1μg／mL和1μg／mL的LPS作用24h可明显促进MDPC-23细胞增殖，72h后则明显抑制细胞增殖（P〈0．05）；当LPS浓度大于1μg／mL时，对细胞的影响以毒性作用为主。在各个时间点不同浓度的LPS均可诱导MDPC-23细胞凋亡，其中以0．1μg／mL的LPS诱导MDPC-23细胞凋亡的效果较理想。Hoechst染色显示0．1μg／mL LPS可诱导细胞出现典型的凋亡形态学特征，且随着LPS作用时间的延长，凋亡细胞比例逐渐增高（P〈0．05），但96h时有大量细胞死亡。结论：LPS诱导MDPC-23细胞凋亡的合适浓度为0．1μg／mL，合适作用时间为24—72h。

AIM： To establish and assess cell apoptosis model of odontoblastic MDPC-23 cells induced by lipopolysaccharide （LPS） in vitro. METHODS： E. eoli LPS at 0 μg/mL, 0.1 μg/mL, 1 μg/mL, 10 μg/mL and 100 μg/mL was used to induce apopotosis of MDPC-23 cells for 24 h, 48 h, 72 h and 96 h respectively. MTT assay was employed to observe the cell proliferation. Flow cytometry （FCM） and Hoechst staining were used to detect cell apoptosis ratio. RESULTS ： 0.1 μg/mL and 1 μg/mL LPS promoted cell proliferation in 24 h exposure, but after 72 h exposure, cell proliferation was significantly inhibited （P 〈 0.05）. LPS with higher concentration （ 〉 1 Izg/mL） mainly exhibited toxic effects on MDPC-23 cells. At various time points （24, 48, 72 and 96 h） , all concentrations of LPS induced apoptosis of MDPC-23 cell cells. However, 0.1 μg/mL LPS showed appropriate apoptosis rate. Hoechst33258 fluorescent staining confirmed that MDPC-23 cell induced by LPS exhibited specific morphological features of apopto- sis, such as pyknosis, nuclear fragmentation and apoptotie bodies. LPS-induced MDPC-23 cell apoptosis showed a time-dependent manner. However, a great number of MDPC-23 cell died after 96 h exposure. CONCLUSION： Ex- posure of 0.1 μg/mL LPS to MDPC-23 cells for 24-72 h is appropriate for the apoptosis induction of the cells.