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

Abstract

Objective. To investigate the killing effect of rukangyin (RKY) activated γδT 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 γδT cells. There are 8 groups including the Circled digit one PBMC control group, Circled digit two RKY group, Circled digit three ZOL group, Enclosed digit four PHA group, Circled Digit FiveRKY+ZOL group, Circled Digit SixRKY+PHA group, Circled Digit SevenZOL+PHA group, and Circled Digit Eight RKY+ZOL+PHA group. At 0 and 14 days of culture, cell viability and γδT cell expansion were detected by flow cytometry. The 8 groups of amplified γδT 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 γδT cells on breast cancer MDA-MB-231 cells. Results. The optimal concentrations of RKY, ZOL, and PHA to activate γδT cell proliferation were 4.5 mg/l, 3 μM, and 60 μg/ml, respectively. On day 0 of culture, the values (x¯±s, %) of γδT cells in groups Circled digit one to Circled Digit Eight 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 γδT cells in groups Circled digit one to Circled Digit Eight 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 γδT cells were compared before and after culture for 0 and 14 days. The t values of group Circled digit one to group Circled Digit Eight 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 γδT 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 Circled digit one to group Circled Digit Eight, the γδT 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 Circled digit one to group Circled Digit Eight 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 γδT cells against MDA-MB-231 cells by activating the proliferation of γδT cells, which provides a basis for Chinese medicine combined with immunotherapy for breast cancer.