Explore the action of MiRNA-21 on shikonin and epidermal growth factor in regulating the proliferation and Apoptosis of HaCaT Cell
Xiaohong Yang1, Fengling Xing2, Maocan Tao1, Lili Ma1, Wei Ding3, Hongbin Luo1, Yi Cao4
1 Department of Dermatology and STD, Zhejiang Hospital of Traditional Chinese Medicine, Hangzhou, Zhejiang, China
2 Department of Chinese Medicine Surgery, Zhejiang Chinese Medical University, Hangzhou, China
3 Department of Dermatology, Zhejiang Chinese Medical University, Third Affiliated Hospital of Zhejiang University of Traditional Chinese Medicine, Hangzhou, China
4 Department of Dermatology and STD, Zhejiang Hospital of Traditional Chinese Medicine, Hangzhou, Zhejiang; Department of Chinese Medicine Surgery, Zhejiang Chinese Medical University, Hangzhou, China
Dr. Yi Cao
Department of Dermatology and STD, Zhejiang Hospital of Traditional Chinese Medicine, Zhejiang Chinese Medical University, Youdian Rd., 54th, Hangzhou, 310006
Source of Support: None, Conflict of Interest: None
Purpose: The aim of the study is to investigate the effect of MicroRNA-21 (miR-21) and its interaction with epidermal growth factor (EGF) and shikonin on the proliferation, and apoptosis of HaCaT cell line.
Materials and Methods: HaCaT cells were cultured under different concentrations of EGF and shikonin, and to calculate their optimal effect dosages. The transfection was performed using Lipofectamine2000, and then gene expression of miR-21 was detected by quantitative real-time polymerase chain reaction (RT-PCR). MTT assay and flow cytometry were applied to test cell proliferation and apoptosis. Western blot and RT-PCR were used to detect the proliferation (proliferating cell nuclear antigen [PCNA], NF-κB/IKKβ) and apoptosis (caspase-3/caspase-9, bcl-2) signals of HaCaT cell.
Results: MTT assay showed that miR-21 mimic and EGF promoted, whereas, shikonin and miR-21 inhibitor inhibited cell viability of HaCaT cell. MiR-21 was upregulated by miR-21 mimic and EGF, while downregulated by shikonin and miR-21 inhibitor. Besides, EGF and miR-21 mimic promoted proliferation-associated signals (PCNA, NF-κB/IKKβ) expression, which were suppressed by shikonin and miR-21 inhibitor. Yet, shikonin and miR-21 inhibitor induced apoptosis-related signals (caspase-3/caspase-9, bcl-2) expression while reversed by EGF and miR-21 mimic which were confirmed by the result of flow cytometry.
Conclusions: MiR-21 promotes the process of EGF-induced cell growth of HaCaT. The antagonized effect of shikonin in EGF-induced proliferation and apoptosis might be mediated by suppressing the expression of miR-21.