Study on the Killing Effect of γδT Cells Activated by Rukangyin on Breast Cancer MDA-MB-231 Cells

Objective To investigate the killing effect of rukangyin (RKY) activated γδ T cells on breast cancer cells MDA-MB-231 and to provide a basis for Chinese medicine combined with immunotherapy for breast cancer.Methods Thus, study isolates peripheral blood mononuclear cells (PBMC) and uses CCK8 to select the optimal concentration of Rukang drink, ZOL (zoledronic acid), and PHA (phytoagglutinin) to activate γδ T cells. There are 8 groups including the ① PBMC control group, ② RKY group, ③ ZOL group, ④ PHA group, ⑤RKY+ZOL group, ⑥RKY+PHA group, ⑦ZOL+PHA group, and ⑧ RKY+ZOL+PHA group. At 0 and 14 days of culture, cell viability and γδ T cell expansion were detected by flow cytometry. The 8 groups of amplified γδ T were cocultured with breast cancer MDA-MB-231 cells labeled with fluorescent dye CFSE at a ratio of 10 : 1 to determine the lethality of γδ T cells on breast cancer MDA-MB-231 cells.Results The optimal concentrations of RKY, ZOL, and PHA to activate γδ T cell proliferation were 4.5 mg/l, 3  μ M, and 60  μ g/ml, respectively. On day 0 of culture, the values ( x ¯ ± s , %) of γδ T cells in groups ① to ⑧ were 3.50 ± 0.72, 3.97 ± 0.45, 3.99 ± 0.15, 4.37 ± 0.24, 4.47 ± 0.97, 4.59 ± 1.35, 3.45 ± 0.40, and 3.89 ± 0.48, while when a comparison between groups was made, F = 1.093 and p = 0.412; there is no significant difference between groups. Besides, when being cultured for 14 days, the values ( x ¯ ± s , %) of γδ T cells in groups ① to ⑧ were 4.77 ± 0.78, 23.22 ± 2.73, 26.4 ± 0.92, 28.66 ± 1.43, 27.99 ± 1.10, 30.21 ± 1.91, 32.51 ± 0.74, and 33.21 ± 0.42. Then, based on the comparison between groups, F = 119.917 and p < 0.001, there are obvious statistical differences between groups. Furthermore, the expansion values of γδ T cells were compared before and after culture for 0 and 14 days. The t values of group ① to group ⑧ were 2.072, 12.051, 41.641, 29.015, 27.777, 18.972, 59.836, and 79.622. Except for the PBMC control group ( p = 0.107), there are significant statistical differences ( p < 0.001). The number of γδ T cell expansion at 14 days was the RKY+ZOL+PHA group>ZOL+PHA group>RKY+PHA group>PHA group>RKY+ZOL group>ZOL group>RKY group>PBMC control group. From group ① to group ⑧, the γδ T cell expansion multiples were 1.14 ± 0.44, 5.25 ± 0.77, 5.70 ± 0.89, 6.05 ± 1.03, 6.21 ± 0.09, 6.76 ± 1.46, 7.52 ± 1.05, and 7.97 ± 1.55, respectively, while the comparison between groups was F = 17.772 and p < 0.001. As for the amplification factor, there was RKY+ZOL+PHA group>ZOL+PHA group>RKY+PHA group>RKY+ZOL group>PHA group>ZOL group>RKY group>PBMC control group. In the killing experiment, the killing rate ( x ¯ ± s , %) of group ① to group ⑧ was 1.08 ± 0.03, 1.89 ± 0.14, 1.22 ± 0.11, 1.31 ± 0.09, 1.48 ± 0.10, 2.02 ± 0.21, 2.18 ± 0.27, and 2.37 ± 0.35, whereas the comparison between groups was F = 20.498 and p < 0.001. In terms of killing rate, there was RKY+ZOL+PHA group>ZOL+PHA group>RKY+PHA group>RKY group>RKY+ZOL group>PHA group>ZOL group>PBMC control group.Conclusion Rukangyin can increase the lethality of γδ T cells against MDA-MB-231 cells by activating the proliferation of γδ T cells, which provides a basis for Chinese medicine combined with immunotherapy for breast cancer.