Chen C J, Anast C S, Brown E M
Endocrine Division, Children's Hospital, Boston, MA.
J Bone Miner Res. 1988 Jun;3(3):279-88. doi: 10.1002/jbmr.5650030306.
Fluoride ion (F-) alone or in conjunction with aluminum (Al3+) has been shown to stimulate the activity of guanine nucleotide-binding proteins (G proteins) in cell membrane preparations from a variety of cell types and in intact hepatic cells. Several studies have indicated that G proteins are involved in the regulation of parathyroid hormone (PTH) secretion. Intracellular second messengers which modulate PTH secretion (e.g., cAMP) have also been found to be regulated by G proteins. We have, therefore, employed F- as a probe to investigate the possible role of G proteins in the modulation of PTH release and the intracellular second messengers that have been implicated in the control of PTH secretion. F- produces a dose-dependent inhibition of PTH release with a maximal inhibitory effect (67%) at 5 mM. F- exerts its inhibitory effect within 5 min and the degree of suppression of PTH secretion gradually increases over 1 hr. F- (5 mM) inhibits PTH secretion at 0.5 mM Ca2+ to the level observed with 2 mM Ca2+ alone; moreover, the effects of F- and high Ca2+ are not additive. While 1 mM F- suppresses PTH secretion by only 21%, and 10 microM Al3+ has virtually no effect at all, together they inhibit PTH release approximately to the level (63% inhibition) observed with 5 mM NaF alone. In the presence of 10(-5) M dopamine, F- produces a concentration-dependent inhibition of cAMP accumulation (0.684 +/- 0.033 pmoles/10(5) cells at 0 mM F- vs. 0.256 +/- 0.048 at 5 mM F-). However, the F- -induced decrease in cAMP cannot account for the inhibition of PTH release by this agent, since addition of methylisobutylxanthine (10(-4) M) by F- -treated cells raises intracellular cAMP content above that of control cells but fails to reverse the inhibition of PTH release. The cytosolic calcium concentration in Fura-2-loaded cells increases from 210 +/- 20 nM to 340 +/- 44 nM after 5 mM F- was added to incubation media. Prior removal of extracellular Ca2+ by EGTA totally blocks the F- -induced rise in cytosolic Ca2+ without preventing the inhibition of PTH release by NaF. F- also produces a time- and dose-dependent increase in the accumulation of IP, IP2, and IP3 in cells prelabeled with [3H]inositol and incubated with 10 mM Li+, consistent with activation of phospholipase C. We conclude that F- is a potent inhibitor of PTH secretion.(ABSTRACT TRUNCATED AT 400 WORDS)
已表明，氟离子（F-）单独或与铝离子（Al3+）结合，能刺激多种细胞类型的细胞膜制剂以及完整肝细胞中鸟嘌呤核苷酸结合蛋白（G蛋白）的活性。多项研究表明，G蛋白参与甲状旁腺激素（PTH）分泌的调节。调节PTH分泌的细胞内第二信使（如cAMP）也已发现受G蛋白调控。因此，我们用F-作为探针，研究G蛋白在调节PTH释放以及参与PTH分泌控制的细胞内第二信使方面可能发挥的作用。F-对PTH释放产生剂量依赖性抑制，在5 mM时具有最大抑制作用（67%）。F-在5分钟内发挥其抑制作用，PTH分泌的抑制程度在1小时内逐渐增加。F-（5 mM）在0.5 mM Ca2+时抑制PTH分泌至单独使用2 mM Ca2+时观察到的水平；此外，F-和高钙的作用并非相加。虽然1 mM F-仅抑制PTH分泌21%，而10 microM Al3+几乎没有作用，但它们共同抑制PTH释放的程度约为单独使用5 mM NaF时观察到的水平（63%抑制）。在存在10(-5) M多巴胺的情况下，F-对cAMP积累产生浓度依赖性抑制（0 mM F-时为0.684±0.033 pmoles/10(5)细胞，5 mM F-时为0.256±0.048）。然而，F-诱导的cAMP减少并不能解释该试剂对PTH释放的抑制作用，因为用F-处理的细胞添加甲基异丁基黄嘌呤（10(-4) M）后，细胞内cAMP含量升高至对照细胞之上，但未能逆转对PTH释放的抑制。在孵育培养基中加入5 mM F-后，用Fura-2负载的细胞的胞质钙浓度从210±20 nM增加到340±44 nM。用EGTA预先去除细胞外Ca2+完全阻断了F-诱导的胞质钙升高，但并未阻止NaF对PTH释放的抑制。F-还使预先用[3H]肌醇标记并与10 mM Li+孵育的细胞中IP、IP2和IP3的积累产生时间和剂量依赖性增加，这与磷脂酶C的激活一致。我们得出结论，F-是PTH分泌的有效抑制剂。（摘要截短至400字）
