Dual control of Shuanghuang Shengbai granule on upstream and downstream signal modulators of CyclinD-CDK4/6 signaling pathway of cell cycle in Lewis-bearing mice with cyclophosphamide-induced myelosuppression

Xian Gu,¹ Zhen-ye Xu,¹ Ling-yu Zhu,¹ Li-fang Wang,¹ Kai Li,¹ Qiang Pei<sup>2

1</sup>Longhua Hospital Shanghai University of TCM, Shanghai, People’s Republic of China; ²The First Affiliated Hospital of Xinxiang Medical University, Weihui, People's Republic of China

Background: This study investigated the dual control mechanism of the Shuanghuang Shengbai granule in modulating the cell cycle in Lewis-bearing mice with cyclophosphamide induced myelosuppression.
Methods: Thirty Lewis-bearing mice were randomly grouped into an untreated group, control group, and treated group. Both treated and untreated groups were intraperitoneally injected with cyclophosphamide to produce a myelosuppression model. Mice in the treated group were fed with the Shuanghuang Shengbai granule (40 g/day) for 6 consecutive days. Standard blood tests and the count of bone marrow nuclear cells were performed, and the cell reproductive cycles of bone marrow and tumors were measured in these mice. In addition, the western blot approach was used to measure the upstream activating signals of CyclinD-CDK4/6 such as c-Myc and CDC25A, the upstream suppression signals such as p16INK4a, and the expression of downstream activated signals such as Rb, pRB, and E2F. All of the tested results were validated by reverse transcription quantitative real-time polymerase chain reaction.
Results: The results showed that the Shuanghuang Shengbai granule could elevate the count of leukocyte and bone marrow nuclear cells of Lewis-bearing mice with cyclophosphamide induced myelosuppression. It could also stimulate bone marrow cells to move from G0/G1 phases to S phase, accelerating the progress of the cell reproductive cycle and increasing the cell proliferation index. Simultaneously, the Shuanghuang Shengbai granule could also suppress cancer cells moving from G0/G1 phase to S phase, reducing the proliferation index. The tumor weight of Lewis-bearing mice in the treated group was much less than those of the control group. Expression levels of c-Myc, CDC25A, Rb, pRb, and E2F of bone marrow in Shuanghuang Shengbai granule-treated mice was higher compared to the control group, whereas they were lower in the cancer cells.
Conclusion: The experimental results demonstrate that the Shuanghuang Shengbai granule has dual control on the cell reproductive cycles in cancer cells and bone marrow nuclear cells in Lewis-bearing mice.

Keywords: Shuanghuang Shengbai granule, CyclinD-CDK4/6, c-Myc, Rb, pRb, E2F