热休克蛋白25在大鼠牙囊细胞中的表达及生物学意义

Expression and biological roles of heat shock protein 2S in rat dental follicle cells

目的检测不同出生时间段以及体外培养大鼠牙囊细胞中热休克蛋白25（heatshock protein25，HSP-25）的表达，探讨其对牙囊细胞增殖和碱性磷酸酶（alkaline phosphatase，ALP）活性的影响，进一步了解HSP-25在牙齿发育中的作用。方法免疫组化法检测HSP-25在出生后第1、3、5、7、9、11天的SD大鼠下颌第一磨牙牙囊中的表达情况；间接免疫荧光法检测HSP-25在体外培养的大鼠牙囊细胞中的表达；甲基噻唑基四唑（MTT）法和流式细胞术分别检测不同质量浓度的重组鼠HSP-25刺激牙囊细胞增殖及细胞周期的变化，并检测ALP活性的变化。结果HSP-25在出生早期大鼠牙囊细胞内不表达或弱表达，第5天时表达增强并维持到第11天。HSP-25在牙囊细胞的胞质中呈阳性表达。MTF法结果显示不同质量浓度的HSP-25刺激牙囊细胞1、2、3d后细胞增殖无明显差异，流式细胞仪结果显示0和100μg/L HSP-25刺激牙囊细胞3d后细胞周期无明显差异。50和100μg／L HSP-25刺激牙囊细胞9d后ALP活性达到最高[A值分别为（1．6±0．3）、（1．8±0．2）]。结论HSP-25在出生早期的大鼠牙囊细胞中无表达，之后随时间变化表达逐渐增强；HSP-25在体外培养的大鼠牙囊细胞胞质中表达；不同质量浓度HSP-25对大鼠牙囊细胞增殖无影响，高质量浓度HSP-25可刺激牙囊细胞ALP活性增高。

Objective To examine the expression of heat shock protein 25 （HSP-25） in dental rat follicles in vivo and in vitro in order to investigate the possible effect of HSP-25 on cell proliferation and alkaline phosphatase （ALP） activity. Methods The expression of HSP-25 in mandibles of postnatal rats from day 1,3, 5, 7, 9, 11 was examined by immunohistochemistry in vitro, the expression of HSP-25 in the dental follicle ceils was detected by the indirect immunofluorescence method. Methyl thiazolyl tetrazolium （MTI＂） assay, flowcytometry and ALP assay were used to detect the effect of HSP-25 on rat dental follicles. Results HSP-25 expression was absent or weak in rat dental follicle cells at early postnatal stage and present from day 5 till day 11. HSP-25 was detected in the cytoplasm of cultured dental follicle ceils. MTT results showed no effect could be detected on dental follicle cell proliferation after stimulation of different concentrations of HSP-25 on day 1, 2, 3. Flowcytometry results also exhibited no difference in cell cycles after stimulation of HSP-25 at 0 μg/L and 100 μg/L. HSP-25 at a concentration of 50 μg/L and 100μg/L could up-regulate the ALP activity on day 9. Conclusions Expression of HSP-25 increases chronologically in the rat dental follicle cells. HSP-25 locates in the cytoplasm of cultured rat dental follicle cells. HSP-25 has no effect on the proliferation of dental follicle cells, however it can up-regulate the ALP activity.