The immune influence of a parabiosis model on tumour-bearing mice
The immune influence of a parabiosis model on tumour-bearing mice
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DOI：
10.4414/smw.2018.14678
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发表时间：
2018-09-24
影响因子：
2.9
通讯作者：
Guo Ximin
中科院分区：
文献类型：
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作者：
Feng Nan;Luo Jianmin;Guo Ximin
目的：本研究的目的是分析联体共生模型对荷瘤小鼠免疫功能的影响。在表达绿色荧光蛋白（GFP+）的C57 BL/6野生型小鼠和不含绿色荧光蛋白（GFP-）的C57 BL/6野生型小鼠之间建立联体共生，以确保动物之间的血液交叉循环，然后检测CD 4 + T细胞、CD 8 + T细胞和白细胞介素2的表达，流式细胞仪检测联体共生模型小鼠脾细胞中IFN-γ和IFN-4、IFN-10的表达。于联体手术后第8天和第14天取联体小鼠肿瘤组织，免疫组化法检测肿瘤组织中CD 3、CD 4、CD 8、CD 31、IFN-γ和血管内皮生长因子（VEGF）的变化。GFP+ C57小鼠的健康细胞通过连接的毛细血管网进入荷瘤小鼠的血液循环，对荷瘤小鼠的CD 4+和CD 8+细胞起到刺激作用，使荷瘤小鼠的CD 4+细胞比阳性对照组增加更多（p < 0.05）。在荷瘤小鼠中检测到的GFP+细胞数量很少，但GFP+细胞可以刺激小鼠自身产生更多的CD 4 +/白细胞介素（IL）-4，CD 4 +/IL-10（p < 0.05）。GFP+小鼠的CD 4 +/IL-2、CD 4 +/IL-4和CD 4 +/IL-10均高于阴性对照组（p < 0.05）。在联体共生模型中，两种小鼠中IFN-γ的水平均降低（p < 0.05）。联体GFP+小鼠的CD 4 +/CD 8+比值高于阴性对照组（p < 0.05）。免疫组化检测中，CD 3、CD 4、CD 8和IFN-γ阳性细胞率均高于阳性对照组，其光密度值分别为0.32 ± 0.63、0.33 ± 0.00、0.31 ± 0.91和0.28 ± 0.14（p < 0.05）。CD 31（0.19 ± 0.50）和VEGF（0.19 ± 0.21）的表达较阳性对照组低，但差异无统计学意义。与阳性对照组相比，CD 31和VEFG细胞表达较低，分别为0.19 +/- 0.50和0.19 +/- 0.21（p > 0.05）。阳性对照组中的CD 31和VEGF细胞值较高，分别为0.32 +/- 0.35和0.29 +/- 0.35，但与共生荷瘤组相比，无显著差异。在第8天，CD 3、CD 4、CD 8和IFN-γ细胞的表达较低：分别为0.22、0.17、0.15和0.16。结论：所建立的异基因联体小鼠模型能较好地适应联体小鼠的状态，可广泛应用于医学实验。联体共生模型在研究免疫调节方面发挥了重要作用，为未来的肿瘤免疫治疗提供了基础。联体共生模型可刺激荷瘤小鼠产生CD 3、CD 4、CD 8和IFN-γ，在免疫调节和肿瘤杀伤中发挥显著作用。联体荷瘤组CD 31、VEFG细胞阳性表达率较低，但与阳性对照组比较，差异无统计学意义。
AIM: The aim of this study was to analyse the immune influence of a parabiosis model on tumour-bearing mice.METHODS: Parabiosis was established between C57BL/6 wild-type mice expressing green fluorescent protein (GFP+) and C57BL/6 wild-type mice without green fluorescent protein (GFP-) to ensure blood cross-circulation between animals, and then the expression of CD4+ T cells, CD8+ T cells and interleukins 2, 4 and 10, and interferon- gamma (INF-gamma) in spleen cells of parabiosis model mice were examined with flow cytometry. At day 8 and day 14 after conjoined surgery, we were aiming to sample tumour tissue in the parabiosis mice and observe changes of CD3, CD4, CD8, CD31, IFN-gamma and vascular endothelial growth factor (VEGF) through immunohistochemical analysis.RESULTS: The interaction of blood was established on the third day with modelling rate of 85.7% after blood interaction. The healthy cells of GFP+ C57 mice entered the blood circulation of tumour-bearing mice via a connecting capillary network, playing a role in stimulating CD4+ and CD8+ cells in the tumour-bearing mice so that CD4+ cells increased more in tumour-bearing mice than in the positive control group (p < 0.05). The number of GFP+ cells that were detected in a tumour-bearing mouse was small, but GFP+ cells can stimulate the mouse itself to generate more CD4+/interleukin (IL)-4, CD4+/IL-10 (p < 0.05). The numbers of CD4+/IL-2, CD4+/IL-4 and CD4+/IL-10 among the GFP+ mice were higher than those in the negative control group(p < 0.05). The levels of IFN-gamma in both mice in the parabiosis model were decreased (p < 0.05). The rate of CD4+/CD8+ in parabiosis GFP+ mice was higher than in the negative control group (p < 0.05). In immunohistochemical tests, the rates of CD3, CD4, CD8 and IFN-gamma positive cells was higher than in the positive control group, with their optical densities of 0.32 +/- 0.63, 0.33 +/- 0.00, 0.31 +/- 0.91 and 0.28 +/- 0.14 respectively (p < 0.05). The expression of CD31 (0.19 +/- 0.50) and VEGF (0.19 +/- 0.21) were lower when compared with the positive control group, with no significant difference. CD31 and VEFG cell expression was low, at 0.19 +/- 0.50 and 0.19 +/- 0.21, respectively, compared with the positive control group (p > 0.05). Values for CD31 and VEGF cells in the positive control group were higher, at 0.32 +/- 0.35 and 0.29 +/- 0.35, respectively, but when compared with the parabiosis tumour-bearing group, there was no significant difference. The expression of CD3, CD4, CD8 and IFN-gamma cells at day 8 was low: 0.22, 0.17, 0.15 and 0.16, respectively. When compared with the parabiosis tumour-bearing group, there was no significant difference.CONCLUSIONS: The established allogeneic parabiosis mice model can be well adapted to the conjoined state of mice and be applied in wide medical experiments. The parabiosis model has played an important role in studying immune regulation, which provides a basis for the future tumour immunotherapy. Parabiosis models can stimulate tumour-bearing mice to generate CD3, CD4, CD8 and IFN-gamma, and play a notable role in immune regulation and tumour destruction. The positive expression rates of CD31 and VEFG cells in the parabiosis tumour-bearing group were lower; however, when compared with the positive control group, there was no significant difference.