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Journal of Peking University (Health Sciences) ›› 2026, Vol. 58 ›› Issue (1): 60-67. doi: 10.19723/j.issn.1671-167X.2026.01.008

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Effects of cold atmosphere plasma treatment on the biological behavior of human gingival fibroblasts

Miao ZHENG1, Xinrong MA1, Hao CHEN2, Hengxin ZHAO3, Yu ZHANG4, Jianguo TAN5, Heping LI3,*(), Xiao WANG1,*()   

  1. 1. Department of Stomatology, Peking University Third Hospital, Beijing 100191, China
    2. School of Clinical Medicine, Tsinghua University (Beijing Tsinghua Changgung Hospital), Beijing 100084, China
    3. Department of Engineering Physics, Tsinghua University, Beijing 100084, China
    4. School of Basic Medical Sciences, Tsinghua University, Beijing 100084, China
    5. Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Center for Stomatology & National Clinical Research Center for Oral Diseases & National Engineering Research Center of Oral Biomaterials and Digital Medical Devices, Beijing 100081, China
  • Received:2025-09-24 Online:2026-02-18 Published:2026-01-05
  • Contact: Heping LI, Xiao WANG
  • Supported by:
    Beijing Natural Science Foundation(L232144)

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Abstract:

Objective: To preliminarily investigate the effects of direct treatment with cold atmosphere plasma (CAP) on the migration and proliferation capabilities of human gingival fibroblasts (HGFs), as well as the correlation between the doses and the effects. Methods: The CAP Bio-Med Platform was used to generate the CAP in this study. The characteristics of the CAP source were kept constant by fixing the discharge voltage, frequency, and gas flow rate. Different CAP doses were generated by adjusting the discharge time (20 s, 60 s, 120 s, 180 s) and used to treat HGFs. The temperature, pH, and reactive oxygen species (ROS) levels in the HGFs culture medium were measured following treatment with different CAP doses. The morphology of the HGFs after treatment was observed via immunofluorescence staining, and the cell perimeter and area were calculated. The migration ability of the HGFs after treatment was assessed using a scratch assay, and their proliferation ability was evaluated using a cell counting kit. Results: As the treatment duration increased, the CAP dose generated by the platform ranged from 0 J to 210.6 J. Different CAP doses did not affect the temperature of the HGFs culture medium. As the CAP dose increased, the pH of the HGFs culture medium first decreased from an initial 8.18±0.06 to 8.13±0.20, then gradually increased to 8.63±0.15 (P < 0.05). The concentration of H2O2 in the culture medium peaked at (55.96±1.51) μmol/L in the 60 s CAP treatment group. With an extension in treatment time, the concentration decreased gradually to (22.92±0.57) μmol/L (P < 0.05). Following low-dose CAP treatment (20 s), HGFs exhibited a larger surface area and more pseudopodia extensions. In contrast, following excessively high-dose CAP treatment(180 s), some HGFs displayed a narrow, elongated spindle shape with a smaller surface area than the low-dose group. Compared with the untreated group, low-dose CAP treatment significantly enhanced the migration and proliferation abilities of HGFs (P < 0.05), whereas excessively high-dose CAP treatment inhibited HGFs migration and proliferation (P < 0.05). Conclusion: Treatment with different doses of CAP alters the pH and ROS levels of the HGFs culture medium. CAP treatment has a dose-dependent effect on the biological behavior of HGFs: Low-dose treatment enhances migration and proliferation, while excessively high-dose treatment inhibits these abilities.

Key words: Cold atmospheric plasma, Fibroblasts, Gingiva, Reactive oxygen species

CLC Number: 

  • R783.6

Figure 1

Cold atmosphere plasma Bio-Med Platform"

Figure 2

Temperature, pH, and concentration of H2O2 in the cell culture medium of the control group and the CAP-treated groups CAP-0 s, CAP-20 s, CAP-60 s, CAP-120 s, and CAP-180 s represent no CAP treatment, CAP treatment for 20 s, 60 s, 120 s, and 180 s respectively. Differences in H2O2 concentration among groups are indicated by uppercase letters. Differences in pH among groups are indicated by lowercase letters. Different letters indicate that the difference between groups is statistically significant (P < 0.05). CAP, cold atmosphere plasma."

Figure 3

Observation of HGFs' morphology in control and CAP-treated groups by high content analysis system (immunofluorescence staining ×20) A, no CAP treatment group; B, CAP treatment for 20 s group; C, CAP treatment for 60 s group; D, CAP treatment for 120 s group; E, CAP treatment for 180 s group. CAP, cold atmosphere plasma; HGFs, human gingival fibroblasts."

Figure 4

Spreading area (A) and perimeter (B) of HGFs in the control and CAP-treated groups CAP-0 s, CAP-20 s, CAP-60 s, CAP-120 s and CAP-180 s represent no CAP treatment, CAP treatment for 20 s, 60 s, 120 s and 180 s respectively. Different lowercase letters indicate that the difference between groups is statistically significant (P < 0.05). CAP, cold atmosphere plasma; HGFs, human gingival fibroblasts."

Figure 5

Representative images of HGFs migration in the control and CAP-treated groups after scratch wounding CAP-0 s, CAP-20 s, CAP-60 s, CAP-120 s and CAP-180 s represent no CAP treatment, CAP treatment for 20 s, 60 s, 120 s and 180 s respectively. The initial wound boundaries (0 h) are outlined in blue. The migrated edges after 12 h and 24 h are outlined in yellow. CAP, cold atmosphere plasma; HGFs, human gingival fibroblasts."

Figure 6

Migratory ability of HGFs at 12 h (A) and 24 h (B) after treatment in control and CAP-treated groups CAP-0 s, CAP-20 s, CAP-60 s, CAP-120 s and CAP-180 s represent no CAP treatment, CAP treatment for 20 s, 60 s, 120 s and 180 s respectively. Different lowercase letters indicate that the difference between groups is statistically significant (P < 0.05). CAP, cold atmosphere plasma; HGFs, human gingival fibroblasts."

Figure 7

The proliferation ability of HGFs in the control and CAP-treated groups measured by CCK-8 assay after 24 h (A), 48 h (B) and 72 h (C) of culture CAP-0 s, CAP-20 s, CAP-60 s, CAP-120 s and CAP-180 s represent no CAP treatment, CAP treatment for 20 s, 60 s, 120 s and 180 s respectively. Different lowercase letters indicate that the difference between groups is statistically significant (P < 0.05). CAP, cold atmosphere plasma; HGFs, human gingival fibroblasts; CCK-8, cell counting kit 8."

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