Intracellular Movement of Cholesterol in Rat Adrenal Cells KINETICS AND EFFECTS OF INHIBITORS
Intracellular Movement of Cholesterol in Rat Adrenal Cells KINETICS AND EFFECTS OF INHIBITORS
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发表时间：
2001
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通讯作者：
F.;R.
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作者：
F.;R.
促肾上腺皮质激素（ACTH）刺激的胆固醇转运到线粒体已定量在大鼠肾上腺在体内和肾上腺细胞悬液。在体内和细胞悬液中，初始转运速率（1.25 pg/mg线粒体/ min）相似，但线粒体胆固醇水平在uivo中达到更高水平。当胆固醇侧链裂解被氨鲁米特（AMG）抑制时，ACTH刺激胆固醇转运进入线粒体（3.1 nmol/107细胞/min）大大超过不存在AMG时ACTH刺激的类固醇生成（0.5 nmol/107细胞/min），并暗示不存在AMG时类固醇生成期间胆固醇转运的速率。肾上腺细胞内的胆固醇转运和类固醇生成均受到细胞松弛素B（抗微丝剂）和长春碱（抗微管剂）的抑制，而放线菌酮（蛋白质合成抑制剂）仅抑制类固醇生成。细胞松弛素抑制胆固醇转运到线粒体更有效地（IDSO = 6至7 SM）比总类固醇在大鼠肾上腺。这些抑制作用在体外细胞悬浮液和体内均发生。这两种抑制剂迅速采取行动uitm：细胞松弛素，tl/2 = 4%min;长春碱，tl/f = 2%min。释放AMG抑制在灌流细胞产生的“过冲”类固醇流出的细胞中，类固醇生成达到160%的正常最大ACTH刺激率。一个直接的对应关系已经建立在过量类固醇激素在这个过冲期和线粒体胆固醇的积累。胆固醇蓄积和过冲均在暴露于AMG 7至8分钟后达到峰值，随后下降。细胞松弛素B和长春碱可阻止线粒体胆固醇水平的晚期下降。它的结论是，微丝和微管参与运输的胆固醇和从线粒体，而蛋白质合成和类固醇生成的耦合最终完全表达在限速转移的胆固醇细胞色素P-450内的线粒体。在血清脂蛋白存在的情况下，细胞松弛素B对肾上腺细胞悬浮液中的类固醇生成没有明显的刺激作用，而这在Y-1肾上腺肿瘤细胞中已有报道。细胞松弛素对大鼠肾上腺的体内作用表明，在短期刺激大鼠肾上腺期间，胆固醇的细胞内转移而不是从血浆脂蛋白转移对皮质酮合成速率至关重要。
Adrenocorticotropin (ACTH)-stimulated cholesterol transport to mitochondria has been quantitated in rat adrenals in vivo and in adrenal cell suspensions. The initial rate of transport (1.25 pg/mg of mitochondria/ min) is similar in vivo and in cell suspensions, but the mitochondrial cholesterol level reaches higher levels in uivo. ACTH stimulation of cholesterol transport into mitochondria when cholesterol side chain cleavage is inhibited by aminoglutethimide (AMG) (3.1 nmol/107 cells/min) greatly exceeds both ACTH-stimulated steroidogenesis (0.5 nmol/107 cells/min) in absence of AMG and, by implication, the rate of cholesterol transport during steroidogenesis in absence of AMG. Both cholesterol transport within adrenal cells and steroidogenesis are inhibited by cytochalasin B (antimicrofilament agent) and vinblastine (antimicrotubule agent) while only steroidogenesis is inhibited by cycloheximide (protein synthesis inhibitor). Cytochalasin inhibits cholesterol transport to mitochondria more effectively ( I D S O = 6 to 7 SM) than total steroidogenesis in rat adrenals. These inhibitory effects occur both with in vitro cell suspensions and in vivo. Both inhibitors acted rapidly in uitm: cytochalasin, tl/2 = 4% min; vinblastine, t l I f = 2% min. Release from AMG inhibition in superfused cells generated an “overshoot” in steroid efflux from the cells in which steroidogenesis reached 160% of the normal maximum ACTH-stimulated rate. A direct correspondence has been established between excess steroidogenesis in this overshoot period and the accumulation of mitochondrial cholesterol. Both cholesterol accumulation and overshoot reach peak values after 7 to 8 min of exposure to AMG, followed by a decline. This late decline in mitochondrial cholesterol levels was prevented by both cytochalasin B and vinblastine. It is concluded that microfilaments and microtubules are involved in the transport of cholesterol both to and from the mitochondria while coupling of protein synthesis and steroidogenesis is expressed ultimately entirely on the rate-limiting transference of cholesterol to cytochrome P-450 within the mitochondria. R a t adrenal cell suspensions did not show the marked stimulation of steroidogenesis by cytochalasin B in the presence of serum lipoproteins which has been reported for Y-1 adrenal tumor cells. The in vivo effects of cytochalasin on rat adrenals suggest that intracellular transfer of cholesterol rather than transfer from plasma lipoproteins is critical to the rate of corticosterone synthesis during short term stimulation of rat adrenals.