Combined Metabonomic and Quantitative Real-Time PCR Analyses Reveal Systems Metabolic Changes in Jurkat T-Cells Treated with HIV-1 Tat Protein
Combined Metabonomic and Quantitative Real-Time PCR Analyses Reveal Systems Metabolic Changes in Jurkat T-Cells Treated with HIV-1 Tat Protein
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结合代谢组学和定量实时 PCR 分析揭示了用 HIV-1 Tat 蛋白处理的 Jurkat T 细胞的系统代谢变化
DOI：
10.1021/pr300173c
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发表时间：
2012-11-01
影响因子：
4.4
通讯作者：
Chai, Yifeng
中科院分区：
文献类型：
--
作者：
Liao, Wenting;Tan, Guangguo;Chai, Yifeng
HIV-1达特蛋白由感染细胞释放，可影响旁观者未感染的T细胞，并诱导许多有助于其发病的生物学应答。为了阐明复杂的致病机制，我们进行了全面的调查达特蛋白相关的细胞外和细胞内代谢的变化，Jurkat T细胞使用组合气相色谱-质谱（GC-MS），反相液相色谱-质谱（RPLC-MS）和亲水相互作用液相色谱-质谱（HILIC-MS）为基础的代谢组学方法。进一步采用定量实时PCR（qRT-PCR）分析来测量几种相关酶的表达以及受干扰的代谢途径。结合代谢组学和qRT-PCR分析显示，HIV-1达特引起显著和全面的代谢变化，表现为HIV-1 Tat处理的细胞中37种代谢物和10种相关酶的显著变化。使用MetaboAnalyst 2.0，发现受调节的途径中有11条途径（Impact-value >0.10）受到剧烈干扰，包括鞘脂代谢、甘氨酸、丝氨酸和苏氨酸代谢、丙酮酸代谢、磷酸肌醇代谢、精氨酸和脯氨酸代谢、柠檬酸循环、苯丙氨酸代谢、色氨酸代谢、磷酸戊糖途径、甘油磷脂代谢、糖酵解或糖异生。这些结果为HIV-1达特蛋白作为“病毒毒素”的复杂致病机制提供了代谢证据，并将有助于使达特蛋白成为治疗干预和疫苗开发的“重要靶点”。
HIV-1 Tat protein is released by infected cells and can affect bystander uninfected T cells and induce numerous biological responses which contribute to its pathogenesis. To elucidate the complex pathogenic mechanism, we conducted a comprehensive investigation on Tat protein-related extracellular and intracellular metabolic changes in Jurkat T-cells using combined gas chromatography-mass spectrometry (GC-MS), reversed-phase liquid chromatography-mass spectrometry (RPLC-MS) and a hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-MS)-based metabonomics approach. Quantitative real-time PCR (qRT-PCR) analyses were further employed to measure expressions of several relevant enzymes together with perturbed metabolic pathways. Combined metabonomic and qRT-PCR analyses revealed that HIV-1 Tat caused significant and comprehensive metabolic changes, as represented by significant changes of 37 metabolites and 10 relevant enzymes in HIV-1 Tat-treated cells. Using MetaboAnalyst 2.0, it was found that 11 pathways (Impact-value >0.10) among the regulated pathways were acutely perturbed, including sphingolipid metabolism, glycine, serine and threonine metabolism, pyruvate metabolism, inositol phosphate metabolism, arginine and proline metabolism, citrate cycle, phenylalanine metabolism, tryptophan metabolism, pentose phosphate pathway, glycerophospholipid metabolism, glycolysis or gluconeogenesis. These results provide metabolic evidence of the complex pathogenic mechanism of HIV-1 Tat protein as a "viral toxin", and would help obligate Tat protein as "an important target" for therapeutic intervention and vaccine development.