Int J Stomatol

Int J Stomatol ›› 2024, Vol. 51 ›› Issue (5): 538-549.doi: 10.7518/gjkq.2024067

• Oral mucosal disease • Previous Articles     Next Articles

Effect of areca nut extract on the biological function of oral epithelial cells

Ming Li1,2(),Zhangui Tang1,2(),Zhenying Yuan3   

  1. 1.Xiangya School/Hospital of Stomatology, Central South University, Changsha 410000, China
    2.Hunan Key Labora-tory of Oral Health Research, Changsha 410000, China
    3.Dept. of Cariology and Endodontics, Wuyi Road District, Changsha Stomatological Hospital, Changsha 410000, China
  • Received:2023-10-16 Revised:2024-04-28 Online:2024-09-01 Published:2024-09-14
  • Contact: Zhangui Tang E-mail:limingq248@163.com;zhgtang@csu.edu.cn
  • Supported by:
    National Natural Science Foundation of China(82170972)

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Abstract:

Objective　This study aimed to investigate the effects of areca nut extract (ANE) on the biological function of human oral epithelial cells. Methods　First, we analyzed the effects of ANE with gradient concentrations on the proli-feration of human oral keratinocytes (HOKs). Next-generation RNA sequencing was used to examine the differential genes between control HOKs and those treated with ANE. Flow cytometry was employed to evaluate the apoptosis and cycle arrest of HOKs. β-galactosidase staining kit was used to detect cell senescence. Immunofluorescence was applied to detect the expression of Ki67 and γH2AX protein in HOKs. Real-time PCR (RT-PCR) was utilized to detect the expression of senescence-related phenotypic genes interleukin (IL)-6, matrix metalloproteinase-2 (MMP2), and TGFB1. Wes-tern blot was adopted to detect the expression of autophagy-related proteins (LC3-Ⅰ/Ⅱ and p62) and epithelial-mesenchymal transition (EMT)-related proteins (SMAD2, p-SMAD2, E-cadherin, and Vimentin). Enzyme-linked immunosorbent assay was employed to detect the expression of exocrine TGFB1 protein in HOKs. Results　Sequencing results showed that ANE caused abnormality in cell cycle, p53, autophagy, FoxO, cellular senescence, and other signal pathways. Flow cytometry showed that ANE induced HOK apoptosis and G2/M phase arrest. The number of senescent cells and the expression of senescent phenotypic secretion genes (IL-6, MMP2, and TGFB1) increased in the ANE group. Compared with those in the control group, the expression levels of γH2AX, LC3-Ⅱ, TGFB1, p-SMAD2, and Vimentin protein increased and those of Ki67, LC3-Ⅰ, and E-cadherin protein decreased in the ANE group. Conclusion　ANE could induce HOK apoptosis, G2/M phase arrest, senescence, DNA double-strand damage, autophagy, and EMT, possibly forming a cell microenvironment that promotes tumor occurrence.

Key words: areca nut, areca nut extract, oral keratinocyte, cell senescence, cycle arrest, autophagy, epithelial-mesenchymal transformation

CLC Number: 

  • Q257

TrendMD: 

Fig 1

Determination of 4 kinds of areca alkaloids in areca nut extract by high performance liquid chromatography"

Fig 2

The effect of ANE on the proliferation of HOK cells was detec-ted by CCK-8 assay"

Fig 3

Volcano map of differential genes between control group and 10 mg/mL ANE group"

Fig 4

KEGG and GO analyzed the signaling pathway and gene annotation involved in the differential genes between the control group and the 10 mg/mL ANE group HOK cells, respectively"

Fig 5

Flow cytometry results showed that ANE could promote HOK cell apoptosis and increase the proportion of G2/M phase cells"

Fig 6

Compared with the control group, the number of Ki67 protein stained HOK cells in 10 mg/mL ANE group was reduced immunofluorescence ×100"

Fig 7

10 mg/mL ANE promoted the senescence of HOK cells, decreased the expression of Ki67 protein and increased the expression of senescence-related phenotype genes"

Fig 8

Areca nut extract induced DNA double-strand damage and autophagy in HOK cells"

Fig 9

Areca nut extract induced the secretion of TGFB1 protein, promoting EMT of HOK cells"

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