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Volume 37 Issue 9
Sep.  2021
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Article Navigation >  CHINESE JOURNAL OF BURNS AND WOUNDS  > 2021  >  37(9): 853-859
Zhang C,Zhang Q,Zhang JH,et al.Effects and molecular mechanism of histone deacetylase 6 inhibitor Tubastatin A on the proliferation and movement of human skin fibroblasts[J].Chin J Burns,2021,37(9):853-859.DOI: 10.3760/cma.j.cn501120-20200519-00274.
Citation: Zhang C,Zhang Q,Zhang JH,et al.Effects and molecular mechanism of histone deacetylase 6 inhibitor Tubastatin A on the proliferation and movement of human skin fibroblasts[J].Chin J Burns,2021,37(9):853-859.DOI: 10.3760/cma.j.cn501120-20200519-00274.
shu

Effects and molecular mechanism of histone deacetylase 6 inhibitor Tubastatin A on the prolifera- tion and movement of human skin fibroblasts

doi: 10.3760/cma.j.cn501120-20200519-00274
  • 1.

    Department of Plastic Surgery, the First Affiliated Hospital of Army Medical University (the Third Military Medical University), Chongqing 400038, China

  • 2.

    State Key Laboratory of Trauma, Burns, and Combined Injury, Institute of Burn Research, the First Affiliated Hospital of Army Medical University (the Third Military Medical University), Chongqing 400038, China

  • 3.

    Department of Endocrinology, the First Affiliated Hospital of Army Medical University (the Third Military Medical University), Chongqing 400038, China

Funds:

General Program of National Natural Science Foundation of China 81873936

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    Abstract
  •   Objective  To explore the effects and possible molecular mechanism of histone deacetylase 6 (HDAC6) inhibitor Tubastatin A on the proliferation and movement of human skin fibroblasts (HSFs).  Methods  The experimental research method was used. HSFs in logarithmic growth phase were taken and divided into negative control group, 1 μmol/L Tubastatin A group, 5 μmol/L Tubastatin A group, and 10 μmol/L Tubastatin A group according to the random number table. The HSFs in negative control group were added with Dulbecco′s modified eagle medium with the final volume fraction of 0.1% dimethyl sulfoxide (hereinafter referred to as the complete medium), and the other three groups were added with the complete medium with the corresponding final molarity of Tubastatin A. After 24 h of conventional culture, the cell proliferation activity was detected using cell counting kit 8 (CCK-8) method and 5-ethynyl-2'-deoxyuridine (EdU) staining; the range of motion of cells within 3 h was observed under the living cell workstation, and the curve movement velocity of the cells was calculated. The protein expressions of extracellular signal-regulated kinase 1/2 (ERK1/2) and phosphorylated ERK1/2 (p-ERK1/2) were detected by Western blotting, and the ratio of p-ERK1/2 to ERK1/2 was calculated to represent the activity of ERK1/2. The sample number in cell proliferation activity detection with CCK-8 method was 6, while the sample numbers in other experiments were 3. Data were statistically analyzed with one-way analysis of variance and least significant difference test.  Results  After 24 h of culture, CCK-8 method and EdU staining showed that compared with negative control group, the cell proliferation activities in 1 μmol/L Tubastatin A group, 5 μmol/L Tubastatin A group, and 10 μmol/L Tubastatin A group were significantly decreased (P<0.01). After 24 h of culture, CCK-8 method showed that compared with 1 μmol/L Tubastatin A group, the cell proliferation activity in 10 μmol/L Tubastatin A group was significantly decreased (P<0.05); EdU staining showed that compared with 1 μmol/L Tubastatin A group, the cell proliferation activities in 5 μmol/L Tubastatin A group and 10 μmol/L Tubastatin A group were significantly decreased (P<0.05 or P<0.01). Within 3 h of observation, the ranges of cell motion in 1 μmol/L Tubastatin A group, 5 μmol/L Tubastatin A group, and 10 μmol/L Tubastatin A group were obviously reduced compared with that in negative control group. Within 3 h of observation, the curve movement velocity of cells in negative control group was (0.780±0.028) μm/min, which was obviously faster than (0.594±0.023), (0.469±0.028), and (0.391±0.021) μm/min of 1 μmol/L Tubastatin A group, 5 μmol/L Tubastatin A group, and 10 μmol/L Tubastatin A group (P<0.01); the curve movement velocity of cells in 1 μmol/L Tubastatin A group was obviously faster than those in 5 μmol/L Tubastatin A group and 10 μmol/L Tubastatin A group (P<0.01); the curve movement velocity of cells in 5 μmol/L Tubastatin A group was obviously faster than that in 10 μmol/L Tubastatin A group (P<0.05). After 24 h of culture, compared with negative control group, the activities of ERK1/2 of cells in 1 μmol/L Tubastatin A group, 5 μmol/L Tubastatin A group, and 10 μmol/L Tubastatin A group were decreased significantly (P<0.01); compared with 1 μmol/L Tubastatin A group, the activities of ERK1/2 of cells in 5 μmol/L Tubastatin A group and 10 μmol/L Tubastatin A group were decreased significantly (P<0.01); compared with 5 μmol/L Tubastatin A group, the activity of ERK1/2 of cells in 10 μmol/L Tubastatin A group was decreased significantly (P<0.05).  Conclusions  HDAC6 inhibitor Tubastatin A may mediate the inhibitory effect on proliferation and movement of HSFs by inhibiting the activity of ERK1/2.

     

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