Institute of Nano Biomedicine and Engineering, Shanghai Engineering Research Center for Intelligent Diagnosis and Treatment Instrument, Department of Instrument Science and Engineering, School of Electronic Information and Electrical Engineering, Shanghai Jiao Tong University, Shanghai, China.
National Engineering Research Center for Nanotechnology, Shanghai, China.
Thorac Cancer. 2023 Jul;14(21):2018-2025. doi: 10.1111/1759-7714.14984. Epub 2023 Jun 7.
There have been reports of increased glutamate pyruvate transaminase 2 (GPT2) expression in certain cancers including breast cancer. Although the role of GPT2 as a metabolic enzyme is well understood in breast cancer progression, little is known about the other roles of GPT2, especially exosomal GPT2.
BT549 and BT474 Cells were cultured and their exosomes were isolated by using ultracentrifugation. Cells migrated through the membrane were stained with crystal violet, and then were observed by microscope. Total RNA was extracted from culture cells and transcribed into cDNA, quantitative real-time RT-PCR was used to detect mRNA expression of ICAM1, VCAM1, and MMP9 using SYBR Green qPCR Mix with a 7500 Fast Real-time PCR system. Western blot was used to detect the gene expression of p-lkBa and TSG101 and GPT2 in breast cancer cells. Immunohistochemistry was used to detect the protein expression of GPT2 and BTRC in cancer cells, animal models loaded with metastasis breast cancer cells were established via tail vein injections. Interaction between GPT2 and BTRC in breast cancer cells was investigated via Co-immunoprecipitation.
GPT2 was up-regulated in TNBC. Exosomes were isolated effectively from TNBC cells, and confirmed that GPT2 was overexpressed inexosomes. QRT-PCR showed that mRNA expression levels of ICAM1, VCAM1, and MMP9 in TNBC were high. Exosomal GPT2 derived from TNBC enhanced migration and invasion of breast cancer via in vitro cell experiment and in vivo animal model experiment. Exosomal GPT2 binds with BTRC to degrade p-lkBa, and improved metastasis of breast cancer cells.
We demonstrated that GPT2 was upregulated in TNBC as well as in exosomes derived from triple-negative breast cancer (TNBC) cells. GPT2 expression was associated with the malignancy of breast cancer and promoted metastasis of breast cancer cells. Moreover, exosomal GPT2 derived from TNBC cells was verified to increase the capacity of breast cancer cells to metastasize through activating beta-transducin repeat containing E3 ubiquitin protein ligase (BTRC). This suggested that exosomal GPT2 may be useful for breast cancer patients as a potential biomarker and treatment target.
已有研究报道，谷氨酸丙酮酸转氨酶 2（GPT2）在某些癌症中表达增加，包括乳腺癌。虽然 GPT2 作为一种代谢酶在乳腺癌进展中的作用已被充分阐明，但人们对 GPT2 的其他作用知之甚少，尤其是外泌体 GPT2。
采用超速离心法分离 BT549 和 BT474 细胞的外泌体。用结晶紫染色细胞穿过膜的部位，然后用显微镜观察。从培养细胞中提取总 RNA，逆转录成 cDNA，用 SYBR Green qPCR Mix 通过 7500 Fast Real-time PCR 系统定量实时 RT-PCR 检测 ICAM1、VCAM1 和 MMP9 的 mRNA 表达。Western blot 检测乳腺癌细胞中 p-lkBa 和 TSG101 和 GPT2 的基因表达。免疫组化检测癌细胞中 GPT2 和 BTRC 的蛋白表达，通过尾静脉注射建立携带转移性乳腺癌细胞的动物模型。通过免疫共沉淀研究乳腺癌细胞中 GPT2 和 BTRC 之间的相互作用。
TNBC 中 GPT2 上调。TNBC 细胞中有效分离出外泌体，并证实外泌体中 GPT2 过表达。QRT-PCR 显示 TNBC 中 ICAM1、VCAM1 和 MMP9 的 mRNA 表达水平较高。体外细胞实验和体内动物模型实验表明，源自 TNBC 的外泌体 GPT2 增强了乳腺癌的迁移和侵袭。外泌体 GPT2 与 BTRC 结合降解 p-lkBa，从而改善乳腺癌细胞的转移。
我们证明 GPT2 在 TNBC 以及源自三阴性乳腺癌（TNBC）细胞的外泌体中上调。GPT2 的表达与乳腺癌的恶性程度有关，并促进了乳腺癌细胞的转移。此外，源自 TNBC 细胞的外泌体 GPT2 被证实通过激活β-转导素重复包含 E3 泛素蛋白连接酶（BTRC）增加乳腺癌细胞转移的能力。这表明外泌体 GPT2 可作为潜在的生物标志物和治疗靶点用于乳腺癌患者。
