Guilu Erxian Glue (龟鹿二仙胶) Inhibits Chemotherapy-Induced Bone Marrow Hematopoietic Stem Cell Senescence in Mice May via p16 INK4a -Rb Signaling Pathway

Objective To evaluate the effect of Guilu Erxian Glue (龟鹿二仙胶, GEG) on cyclophosphamide (CTX)-induced bone marrow hematopoietic stem cells (HSCs) senescence in mice and explore the underlying mechanism. Methods The H 22 liver cancer ascites lump model was established in male Kunming mice by injecting intraperitoneally (i.p.) with 5 × 10 6 /mL H 22 cells per mouse. Fifty tumor-bearing mice were divided into the control, model, pifithrin- α , GEG, and GEG+pifithrin- α groups using a random number table, 10 mice in each group. CTX (100 mg/kg i.p.) was administrated to mice from day 1 to day 3 (d1–d3) continuously except for the control group. The mice in the pifithrin- α , GEG and GEG+pifithrin- α groups were treated with pifithrin- α (2.2 mg/(kg·d) i.p.) for 6 consecutive days (d4–d9), GEG (9.5 g/(kg·d) i.p.) for 9 consecutive days (d1–d9), and GEG plus pifithrin- α , respectively. HSCs were collected after 9-d drug treatment. The anti-aging effect of GEG was studied by cell viability, cell cycle, and β -galactosidase (β -gal) assays. The mRNA and protein expressions of cyclin-dependent kinase 2 (CDK2), CDK4, inhibitor of cyclin-dependent kinase 4a encoding the tumor suppressor protein p16 (p16 INK4a ), p21 Cip1/Waf1 , p53, and phosphorylated retinoblastoma (pRb) were evaluated by quantitative real-time reverse transcription-polymerase chain reaction and semi-quantitative Western blot, respectively. Results Compared with the model group, GEG increased cell viability as well as proliferation ( P <0.05 or P <0.01) and reduced β -gal expression. Furthermore, GEG significantly decreased the expressions of p16 INK4a , p53 and p21 Cip1/Waf1 proteins, and increased the expressions of CDK2, CDK4 and pRb proteins compared with the model group ( P <0.05 or P <0.01). Conclusion GEG can alleviate CTX-induced HSCs senescence in mice, and the p16 INK4a -Rb signaling pathway might be the underlying mechanism.