Cholesterol internalization and metabolism in insect prothoracic gland, a steroidogenic organ, via lipoproteins
Cholesterol internalization and metabolism in insect prothoracic gland, a steroidogenic organ, via lipoproteins
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DOI：
10.1016/j.steroids.2018.01.012
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发表时间：
2018-06-01
期刊：
影响因子：
2.7
通讯作者：
Kataoka, Hiroshi
中科院分区：
文献类型：
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作者：
Igarashi, Fumihiko;Ogihara, Mad H.;Kataoka, Hiroshi
膳食甾醇包括胆固醇和植物甾醇，是昆虫前胸腺合成类固醇激素（蜕皮甾体）的重要底物。在昆虫模式物种之一的家蚕中，胆固醇在PG细胞中生物转化为蜕皮激素，从而很好地证明了类固醇的合成。由于昆虫缺乏从头合成细胞固醇的能力，脂蛋白，载脂蛋白（Lp），一直被认为是主要的胆固醇供应源;然而，胆固醇从Lp到PG细胞的行为的细节至今尚未分析。本研究采用胆固醇标记物22-NBD-胆固醇和胆固醇-25，26，26，26，27，27，27-d 7（cholesterol-d 7）建立了胆固醇掺入法，并以家蚕为实验材料，研究了胆固醇在体外PGs中的内化和代谢。使用22-NBD-胆固醇可视化胆固醇的内化。PGs细胞内22-NBD-胆固醇信号富集，与脂蛋白解离，定位于细胞膜近端。在PG中观察到的分布模式与其他组织如脑、脂肪体和马氏管不同，表明PG中胆固醇的内化与其他组织不同。使用LC-MS/MS方法追踪胆固醇的代谢，以检测胆固醇-d 7、7-脱氢胆固醇-d 7（预期的中间代谢物）和最终产物蜕皮激素-d 6。在与标记的Lp孵育的PG培养物中检测到7-脱氢胆固醇-d 7和蜕皮激素-d 6，表明Lp的胆固醇在PG中用于蜕皮激素的合成。我们的研究结果揭示了胆固醇在PGs中的独特行为，首次直接证明了脂蛋白胆固醇在昆虫类固醇生成器官中的生化命运。这将有助于了解脂蛋白胆固醇参与昆虫类固醇激素合成。
Dietary sterols including cholesterol and phytosterols are essential substrates for insect steroid hormone (ecdysteroid) synthesis in the prothoracic glands (PGs). In the silkworm Bombyx mori, one of the model species of insects, the steroidogenesis has been well demonstrated that cholesterol biotransformation into ecdysone in the PG cells. Because insects lack the ability to synthesize cellular sterol de novo, lipoprotein, lipophorin (Lp), has been thought to be the major cholesterol supply source; however, details of cholesterol behavior from Lp to the PG cells has not been analyzed till date. In this report, we developed Lp incorporation method using labeled cholesterols such as 22-NBD-cholesterol and cholesterol-25,26,26,26,27,27,27-d7 (cholesterol-d7), and analyzed the internalization and metabolism of cholesterol in PGs in vitro using the silkworm Bombyx mori. The internalization of cholesterol was visualized using 22-NBD-cholesterol. PGs showed an enriched cellular 22-NBD-cholesterol signal, which dissociated from the Lp localizing at the close area of cell membrane. The distribution pattern observed in the PGs was different from other tissues such as the brain, fat body, and Malpighian tubules, suggesting that the internalization of cholesterol in the PGs was distinct from other tissues. The metabolism of cholesterol was traced using LC-MS/MS methods to detect cholesterol-d7, 7-dehydrocholesterol-d7 (an expected intermediate metabolite), and the final product ecdysone-d6. 7-Dehydrocholesterol-d7 and ecdysone-d6 were detected in the PG culture incubated with labeled Lp, showing that the cholesterol of Lp was utilized for ecdysone synthesis in the PGs. Our results reveal the distinct behavior of cholesterol in the PGs, with the first direct evidence of biochemical fate of lipoprotein cholesterol in insect steroidogenic organ. This will aid in the understanding of the involvement of lipoprotein cholesterol in steroid hormone synthesis in insects.