Benzidine promotes the stemness of bladder cancer stem cells via activation of the Sonic hedgehog pathway

Wang,Dengdian; Xie,Dongdong; Bi,Liangkuan; Wang,Yi; Zou,Ci; Chen,Lei; Geng,Hao; Qian,Weiwei; Li,Yuan; Sun,Hongliang; Wang,Xin; Lu,Youlu; Yu,Dexin; Zhong,Caiyun

doi:10.3892/ol.2020.12407

Benzidine promotes the stemness of bladder cancer stem cells via activation of the Sonic hedgehog pathway

Authors:
- Dengdian Wang
- Dongdong Xie
- Liangkuan Bi
- Yi Wang
- Ci Zou
- Lei Chen
- Hao Geng
- Weiwei Qian
- Yuan Li
- Hongliang Sun
- Xin Wang
- Youlu Lu
- Dexin Yu
- Caiyun Zhong
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Affiliations: Department of Urology, The Second Hospital of Anhui Medical University, Hefei, Anhui 230601, P.R. China, Department of Clinical Nutrition, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Nanjing, Jiangsu 210008, P.R. China, Department of Nutrition and Food Safety, Center for Global Health, School of Public Health, Nanjing Medical University, Nanjing, Jiangsu 211166, P.R. China
Published online on: December 23, 2020 https://doi.org/10.3892/ol.2020.12407
Article Number: 146
Copyright: © Wang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Substantial evidence suggests that cancer stem cells (CSCs) are the main cause of the initiation, progression and recurrence of tumors. Benzidine has been identified as a risk factor for bladder cancer. The aim of the present study was to investigate the effects of benzidine on bladder CSCs (BCSCs) and the possible mechanism underlying its action. The bladder cancer cell lines UM‑UC‑3 and EJ were maintained in serum‑free medium and cells forming three‑dimensional spheres were characterized as BCSCs. The sphere‑forming cells were exposed to different concentrations of benzidine and vismodegib, and western blotting was performed to evaluate the expression of markers associated with CSCs and the Sonic hedgehog (SHH) signaling pathway. Flow cytometry was used to detect the distribution of cells in different phases of the cell cycle, and immunofluorescence staining was used to detect the protein expression of CD44. The results revealed that the levels of BCSC markers, namely CD133, CD44, aldehyde dehydrogenase 1‑A1, Nanog and octamer‑binding transcription factor‑4, in the cell spheres were markedly elevated compared with those in cells cultured in serum‑supplemented medium. Furthermore, benzidine increased the expression of BCSC markers and promoted the sphere‑forming ability of the cells. In addition, it was observed that benzidine activated the SHH pathway, while inhibition of the Shh pathway using vismodegib diminished the promoting effects of benzidine on BCSCs. The findings of the present study indicate that benzidine promoted the stemness of BCSCs via activation of the SHH pathway, which may support further exploration of the molecular basis of the association between benzidine exposure and bladder oncogenesis.

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