锌激活体外培养大鼠成骨细胞肌醇磷脂信号转导系统

目的：　研究锌对体外培养大鼠成骨细胞肌醇磷脂信号转导系统的影响。方法：　从大鼠颅骨分离出成骨细胞，于 Ｄ Ｍ Ｅ Ｍ 培养基中进行传代培养。实验分为对照组、１０μｍｏｌ／ Ｌ、２５μｍｏｌ／ Ｌ 及５０μｍｏｌ／ Ｌ Ｚｎ２ ＋ 剂量组：分别测定了 Ｚｎ２ ＋ 对成骨细胞长期或短期作用后蛋白激酶 Ｃ（ Ｐ Ｋ Ｃ） 活性及三磷酸肌醇（ Ｉ Ｐ３） 含量的变化。根据被磷酸化多肽底物的量测定 Ｐ Ｋ Ｃ 活性，按照３ Ｈ ｍ ｙｏ 肌醇掺入及层析分离法测定 Ｉ Ｐ３ 含量。结果：　（１） 三个不同剂量的 Ｚｎ２ ＋ 作用于大鼠成骨细胞３ 天后， Ｉ Ｐ３ 含量及 Ｐ Ｋ Ｃ 活性均显著高于对照组；且随着 Ｚｎ２ ＋ 剂量的增加， Ｉ Ｐ３ 含量及 Ｐ Ｋ Ｃ 活性有相应增加的趋势；（２） 在短期实验中，２５μｍｏｌ／ Ｌ Ｚｎ２ ＋ 对成骨细胞的作用比较明显，仅作用１５ 分钟， Ｉ Ｐ３ 含量即明显提高，且在以后的各个作用时点均明显高于对照组，但时间效应曲线平坦；５０μｍ ｏｌ／ Ｌ Ｚｎ２ ＋ 组 Ｉ Ｐ３ 含量在各个时点略低于２５μｍｏｌ／ Ｌ Ｚｎ２ ＋组；１０μｍ ｏｌ／ Ｌ Ｚｎ２ ＋ 组略高于对照组。２５μｍｏｌ／ Ｌ Ｚｎ２ ＋ 、５０μｍ ｏｌ／ Ｌ Ｚｎ２ ＋ 组 Ｐ Ｋ

ZINC ACTIVATES PHOSPHOINOSITOL SIGNAL TRANSDUCTION SYSTEM IN RAT OSTEOBLASTS IN VITRO

Objective: The effect of zinc on phosphoinositol signal transduction system in rat osteoblasts in vitro was studied. Methods: The osteoblasts were isolated from the rat calvaria, and were subcultured in DMEM medium. Four groups were designed in the study, and they were control, 10μmol/L, 25μmol/L and 50μmol/L Zn 2+ groups respectively. Cellular PKC activity and IP 3 content were measured after treatment of Zn 2+ for a long period or a short period. PKC activity was measured according to the level of phosphorylated peptide. IP 3 content was measured according to the methods of 3H myo inositol incorporation and chromatography separation. Results: (1)In long term test cellular IP 3 content and PKC activity in three Zn 2+ groups were significantly elevated as compared with the control group after treatment of Zn 2+ for three days. IP 3 content and PKC activity increased with Zn 2+ levels. (2)In short term test the effect of 25μmol/L Zn 2+ was obvious. Cellular IP 3 content increased significantly just after treatment of 25μmol/L Zn 2+ for 15 minutes, and remained at a high level in two hours. The time response curve kept stable. IP 3 content in 50μmol/L Zn 2+ group was a little lower than that of 25μmol/L Zn 2+ group. IP 3 content in 10μmol/L Zn 2+ group was just a little more than that of control group. PKC activities in 25μmol/L Zn 2+ group and 50μmol/L Zn 2+ group were higher than that of the control group at each time point in 2 hours. The time response curve of PKC was just stable and similar to that of IP 3. Conclusion: Zinc can activate the phosphoinositol signal transduction system in rat osteoblasts in vitro, and elevate IP 3 content and PKC activity. The mechanism includes not only the membrane receptor mediated pathway but also possibly the direct Zn 2+ action.