Abstract：
OBJECTIVE: Exosome exchange between cancer cells or between cancer and stromal cells is involved in cancer metastasis. We have previously developed in vivo color-coded labeling of cancer cells and stromal cells with spectrally-distinct fluorescent genetic reporters to demonstrate the role of exosomes in metastasis. In the present study, we studied exosome transfer between different pancreatic-cancer cell lines in vivo and in vitro and its potential role in metastasis.
METHODS: Human pancreatic-cancer cell lines AsPC-1 and MiaPaCa-2 were used in the present study. AsPC-1 cells contain a genetic exosome reporter gene labeled with green fluorescent protein (pCT-CD63-GFP) and MiaPaCa-2 cells express red fluorescent protein (RFP). Both cell lines were co-injected into the spleen of nude mice (n=5) to further study the role of exosome exchange in metastasis. Three weeks later mice were sacrificed and tumors at the primary and metastatic sites were cultured and observed by confocal fluorescence microscopy for exosome transfer.
RESULTS: The primary tumor formed in the spleen and metastasized to the liver, as observed macroscopically. Cells were cultured from the spleen, liver, lung, bone marrow and ascites. Transfer of exosomes from AsPC-1 to MiaPaCa-2 was demonstrated in the cultured cells by confocal fluorescence microscopy. Moreover, cell fusion was also observed along with exosome transfer. Exosome transfer did not occur during in vitro co-culture between the two pancreatic-cancer cell lines, suggesting a role of the tumor microenvironment (TME) in exosome transfer.
CONCLUSIONS: The transfer of exosomes between different pancreatic-cancer cell lines was observed during primary-tumor and metastatic growth in nude mice. This cell-cell communication might be a trigger of cell fusion and promotion of cancer metastasis. Exosome transfer between the two pancreatic-cancer cell lines appears to be facilitated by the TME, as it did not occur during in vitro co-culture.
METHODS: Human pancreatic-cancer cell lines AsPC-1 and MiaPaCa-2 were used in the present study. AsPC-1 cells contain a genetic exosome reporter gene labeled with green fluorescent protein (pCT-CD63-GFP) and MiaPaCa-2 cells express red fluorescent protein (RFP). Both cell lines were co-injected into the spleen of nude mice (n=5) to further study the role of exosome exchange in metastasis. Three weeks later mice were sacrificed and tumors at the primary and metastatic sites were cultured and observed by confocal fluorescence microscopy for exosome transfer.
RESULTS: The primary tumor formed in the spleen and metastasized to the liver, as observed macroscopically. Cells were cultured from the spleen, liver, lung, bone marrow and ascites. Transfer of exosomes from AsPC-1 to MiaPaCa-2 was demonstrated in the cultured cells by confocal fluorescence microscopy. Moreover, cell fusion was also observed along with exosome transfer. Exosome transfer did not occur during in vitro co-culture between the two pancreatic-cancer cell lines, suggesting a role of the tumor microenvironment (TME) in exosome transfer.
CONCLUSIONS: The transfer of exosomes between different pancreatic-cancer cell lines was observed during primary-tumor and metastatic growth in nude mice. This cell-cell communication might be a trigger of cell fusion and promotion of cancer metastasis. Exosome transfer between the two pancreatic-cancer cell lines appears to be facilitated by the TME, as it did not occur during in vitro co-culture.
摘要：
目的：癌细胞之间或癌细胞与基质细胞之间的外泌体交换参与癌症转移。我们先前已经开发了具有光谱上不同的荧光基因报告基因的癌细胞和基质细胞的体内颜色编码标记，以证明外泌体在转移中的作用。在本研究中,我们在体内和体外研究了不同胰腺癌细胞系之间的外泌体转移及其在转移中的潜在作用。
方法：本研究使用人胰腺癌细胞系AsPC-1和MiaPaCa-2。AsPC-1细胞含有用绿色荧光蛋白（pCT-CD63-GFP）标记的遗传外泌体报告基因，MiaPaCa-2细胞表达红色荧光蛋白（RFP）。将两种细胞系共同注射到裸鼠(n=5)的脾脏中以进一步研究外泌体交换在转移中的作用。三周后,处死小鼠,培养原发和转移部位的肿瘤,并通过共聚焦荧光显微镜观察外泌体转移。
结果：原发性肿瘤在脾脏中形成并转移到肝脏，如宏观观察。从脾脏培养细胞，肝脏,肺，骨髓和腹水.通过共聚焦荧光显微镜在培养的细胞中证明了外泌体从AsPC-1转移到MiaPaCa-2。此外，还观察到细胞融合以及外泌体转移。在两种胰腺癌细胞系之间的体外共培养过程中，未发生外泌体转移，提示肿瘤微环境（TME）在外泌体转移中的作用。
结论：在裸鼠的原发肿瘤和转移生长过程中观察到外泌体在不同胰腺癌细胞系之间的转移。这种细胞-细胞通讯可能是细胞融合和促进癌症转移的触发因素。两种胰腺癌细胞系之间的外泌体转移似乎是由TME促进的，因为它在体外共培养期间没有发生。
方法：本研究使用人胰腺癌细胞系AsPC-1和MiaPaCa-2。AsPC-1细胞含有用绿色荧光蛋白（pCT-CD63-GFP）标记的遗传外泌体报告基因，MiaPaCa-2细胞表达红色荧光蛋白（RFP）。将两种细胞系共同注射到裸鼠(n=5)的脾脏中以进一步研究外泌体交换在转移中的作用。三周后,处死小鼠,培养原发和转移部位的肿瘤,并通过共聚焦荧光显微镜观察外泌体转移。
结果：原发性肿瘤在脾脏中形成并转移到肝脏，如宏观观察。从脾脏培养细胞，肝脏,肺，骨髓和腹水.通过共聚焦荧光显微镜在培养的细胞中证明了外泌体从AsPC-1转移到MiaPaCa-2。此外，还观察到细胞融合以及外泌体转移。在两种胰腺癌细胞系之间的体外共培养过程中，未发生外泌体转移，提示肿瘤微环境（TME）在外泌体转移中的作用。
结论：在裸鼠的原发肿瘤和转移生长过程中观察到外泌体在不同胰腺癌细胞系之间的转移。这种细胞-细胞通讯可能是细胞融合和促进癌症转移的触发因素。两种胰腺癌细胞系之间的外泌体转移似乎是由TME促进的，因为它在体外共培养期间没有发生。
