Anti-tumor Proliferation And Metastasis Effects Of Licochalcone B On Bladder Cancer And Its Mechanisms

Objective:To examine the mechanisms by which licochalcone B (LCB) inhibits the proliferation and metastasis of bladder cancer.Methods:Cell viability was evaluated using a3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. The apoptotic rate was determined via flow cytometry using an annexin V-FITC apoptosis detection kit. The expressions of Bcl-2, Bax, caspase-3, survivin, poly (ADP-ribose) polymerase (PARP), cell division cycle25A (Cdc25A) and cell division cycle25B (Cdc25B) in protein level were analyzed via Western blot. The changes in mRNA level of cyclinA, cyclin-dependent kinase-1(CDK1) and cyclin-dependent kinase-2(CDK2) were detected by real-time quantitative PCR (qRT-PCR) assay. Cell-matrix adhesion of T24cells was measured through MTT assay. The migration of T24cells was measured by wound healing assay. The matrix metalloproteinase (MMP-2and MMP-9) activity in T24cells was measured by qRT-PCR and gelatin zymography methods. In addition, tumorigenicity, HE staining, and TUNEL staining were employed to investigate the influence of LCB on the growth of murine bladder cancer in vivo. The expressions of activator protein-1(AP-1) and nuclear factor-KB (NF-κB) were analyzed via enzyme-linked immunosorbent assay (ELISA).Results:(1) The Findings indicated that LCB (40,60,80μM) significantly inhibited proliferation of bladder cancer T24and EJ cells in a concentration-and time-dependent manner.(2) LCB significantly induced cell cycle arrest in the S phase after72h of treatment and caused a decrease in cyclin A, CDK1, CDK2mRNA expressions accompanied with a decrease in Cdc25A and Cdc25B protein levels.(3) Hoechst dye staining of condensed nuclei and annexin V-FITC/PI staining revealed the manifestation of apoptosis after LCB-treated for72h. LCB treatment downregulated Bcl-2and survivin expressions, enhanced Bax expression, activated caspase-3and cleaved PARP protein. Consistently, the tumorigenicity of LCB-treated bladder cancer MB49cells was limited significantly in vitro and the MB49tumor model performed in C57BL/6mice in vivo.(4) LCB signifieantly inhibited cell-matrix adhesion and decreased the MMP-9mRNA and protein expressions after LCB-treated T24cells for24h, as well as NF-κB level.Conclusions:These findings provide support for the use of LCB in chemoprevention and bladder cancer therapy.