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Journal of Peking University (Health Sciences) ›› 2025, Vol. 57 ›› Issue (5): 884-894. doi: 10.19723/j.issn.1671-167X.2025.05.012

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HDAC2-mediated H3K27 acetylation promotes the proliferation and migration of hepatocellular carcinoma cells

Shaohai TANG1, Baoming YANG1, Jiankun LI1, Lili ZHAO2, Yifan WANG3, Shunxiang WANG1,*()   

  1. 1. Department of Hepatobiliary Surgery, the Fourth Hospital of Hebei Medical University, Shijiazhuang 050000, China
    2. College of Pharmacy, Hebei Medical University, Shijiazhuang 050000, China;
    3. Department of Colorectal Surgery, the Fourth Hospital of Hebei Medical University, Shijiazhuang 050000, China
  • Received:2024-06-28 Online:2025-10-18 Published:2025-06-18
  • Contact: Shunxiang WANG
  • Supported by:
    the Natural Science Foundation Project of Hebei Province(H2020206455); the 2023 Annual Medical Science Research Project Plan of Hebei Province(20230875); the Key Medical Science Research Project Plan of Hebei Province in 2018(20180537); the 2015 and 2016 Hebei Provincial Government' s Plan for Funding the Cultivation of Outstanding Clinical Medicine Talents and Basic Research Projects(361006)

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Abstract: Objective: To explore the specific mechanism of histone deacetylase 2 (HDAC2) mediated histone H3 lysine 27 acetylation (H3K27ac) modification in promoting the proliferation and migration of hepatocellular carcinoma cells. Methods: Samples of 40 cases of hepatocellular carcinoma and paracancerous tissues resected from January 2021 to January 2023 were collected. The expressions of HDAC2 and H3K27ac in hepatocellular carcinoma, paracancerous tissues and cell lines were detected by immunohistochemistry and Western blotting. The correlation between the expression levels of HDAC2 and H3K27ac and the relationship between HDAC2 expression and clinicopathological characteristics of patients with hepatocellular carcinoma were analyzed. The proliferation, migration and invasion of Hep3B and HepG2 cells were determined by MTS, clone formation, scratch and Transwell experiments. The acetylation of H3K27 mediated by HDAC2 was verified by Western blotting, real-time fluorescence quantitative PCR (qRT-PCR) and chromatin immunoprecipitation high-throughput sequencing (ChIP-seq). In vivo xenotransplantation experiment, the tumorigenicity of cells in each group was measured, and the expression of proteins related to phosphoinositide 3-kinases/phosphatase and tensin homolog deleted on chromosome ten/protein kinase B/mammalian target of rapamycin (PI3K/PTEN/AKT/mTOR) signal pathway was detected. Results: High expression of HDAC2 and low expression of H3K27ac were found in hepatocellular carcinoma tissues and cell lines (P < 0.05), and there was a negative correlation between them (r=-0.477, P=0.002). The expression of HDAC2 was related to tumor size, hepatitis B virus infection, TNM stage and portal vein tumor thrombus (P < 0.05). Compared with the sh-NC group of Hep3B and HepG2 cells, the proliferation, clone formation, migration and invasion ability of sh-HDAC2 group were decreased (P < 0.05). Compared with the Empty group, the HDAC2 group exhibited increased expression levels and activity of HDAC2, as well as enhanced cell proliferation, clone formation, migration, invasion ability, tumor volume and mass in vivo, and elevated expression levels of p-PI3K, p-AKT, and p-mTOR (P < 0.05). Conversely, the enrichment and expression levels of H3K27ac, along with the expression level of PTEN, were decreased (P < 0.05). In the iHDAC2 group, the expression levels and activity of HDAC2, as well as the proliferation, clone formation, migration, invasion ability, tumor volume and mass in vivo, and expression levels of p-PI3K, p-AKT, and p-mTOR were reduced (P < 0.05). Additionally, the expression levels of H3K27ac and PTEN were increased (P < 0.05). To validate the involvement of the PI3K/PTEN/AKT/mTOR signaling pathway in HDAC2-mediated regulation of malignant behaviors in liver cancer cells through H3K27ac, the PI3K activator 740Y-P was introduced. Compared with the iHDAC2 group, the iHDAC2+740Y-P group exhibited increased proliferation, clone formation, migration, invasion ability, tumor volume and mass in vivo, and elevated expression levels of p-PI3K, p-AKT, and p-mTOR (P < 0.05). Conversely, the expression level of PTEN was decreased (P < 0.05). Conclusion: HDAC2 initiates PI3K/PTEN/AKT/mTOR signal pathway by mediating H3K27 acetylation, which promotes the occurrence and development of hepatocellular carcinoma.

Key words: Histone deacetylase 2, Histone H3 lysine 27 acetylation, Hepatocellular carcinoma, Proliferation, Migration

CLC Number: 

  • R735.7

Table 1

Primer sequence"

Gene Primer sequences (5′-3′)
HDAC2 Forward：ATGGCGTACAGTCAAGGAGGC
Reverse：AAATCAGAACAGCTCAGCAAC
GAPDH Forward：ATGGGGAAGGTGAAGGTCGG
Reverse：TTACTCCTTGGAGGCCATGT

Figure 1

Expression of HDAC2 and H3K27ac in hepatocellular carcinoma A, immunohistochemical detection of HDAC2 and H3K27ac expression in hepatocellular carcinoma; B, correlation analysis of HDAC2 and H3K27ac expression in hepatocellular carcinoma; C, detection of HDAC2 and H3K27ac expression in hepatocellular carcinoma cell line by Western blotting. HDAC, histone deacetylase; H3K27ac, histone H3 lysine 27 acetylation; GAPDH, glyceraldehyde-3-phosphatedehydrogenase. *P < 0.05, vs. L02 cell."

Table 2

Relationship between HDAC2 expression level and clinicopathological features of patients with hepatocellular carcinoma"

Items Total HDAC2 expression χ2 P
High (n=20) Low (n=20)
Age/years
  ≥59 25 (62.50) 14 (35.00) 11 (27.50) 0.960 0.327
  ＜59 15 (37.50) 6 (15.00) 9 (22.50)
Gender
  Male 21 (52.50) 13 (32.50) 8 (20.00) 2.506 0.113
  Female 19 (47.50) 7 (17.50) 12 (30.00)
Tumor size/cm
  ≥3 28 (70.00) 17 (42.50) 11 (27.50) 4.286 0.038
  ＜3 12 (30.00) 3 (7.50) 9 (22.50)
HBV infection
  Positive 30 (75.00) 19 (47.50) 11 (27.50) 8.533 0.003
  Negative 10 (25.00) 1 (2.50) 9 (22.50)
Alpha-fetoprotein/(μg/L)
  ≥400 27 (67.50) 12 (30.00) 15 (37.50) 1.026 0.311
  ＜400 13 (32.50) 8 (20.00) 5 (12.50)
Histologic grade
  Well and moderate 25 (62.50) 10 (25.00) 15 (37.50) 2.667 0.102
  Low 15 (37.50) 10 (25.00) 5 (12.50)
TNM stage
  Ⅰ to Ⅱ 19 (47.50) 6 (15.00) 13 (32.50) 4.912 0.027
  Ⅲ to Ⅳ 21 (52.50) 14 (35.00) 7 (17.50)
Portal vein tumor thrombus
  Yes 11 (27.50) 9 (22.50) 2 (5.00) 6.144 0.013
  No 29 (72.50) 11 (27.50) 18 (45.00)
Cirrhosis
  Yes 15 (37.50) 6 (15.00) 9 (22.50) 0.960 0.327
  No 25 (62.50) 14 (35.00) 11 (27.50)

Figure 2

Effects of inhibition of HDAC2 on proliferation, migration and invasion of hepatocellular carcinoma cells A, qRT-PCR assay results; B, MTS assay result; C, clone formation experiment results; D, wound scratch assay results; E, transwell assay results. HDAC, histone deacetylase; GAPDH, glyceraldehyde-3-phosphatedehydrogenase. *P < 0.05, vs. sh-NC group."

Figure 3

Acetylation of H3K27 mediated by HDAC2 A, Western blotting assay result; B, qRT-PCR assay results; C, result of HDAC2 activity detection; D, ChIP-seq assay results. HDAC, histone deacetylase; H3K27ac, histone H3 lysine 27 acetylation; GAPDH, glyceraldehyde-3-phosphatedehydrogenase. *P < 0.05, vs. sh-NC group. # P < 0.05, vs. empty group."

Figure 4

In vitro cell experiments show that HDAC2-mediated H3K27 acetylation promotes the proliferation, migration and invasion of hepatocellular carcinoma cells A, MTS assay result; B, clone formation experiment results; C, wound scratch assay results; D, transwell assay results. HDAC, histone deacetylase; H3K27ac, histone H3 lysine 27 acetylation; GAPDH, glyceraldehyde-3-phosphatedehydrogenase. *P < 0.05, vs. empty group. #P < 0.05, vs. iHDAC2 group."

Figure 5

In vivo cell experiments show that HDAC2-mediated H3K27 acetylation initiates PI3K/PTEN/AKT/mTOR signal pathway to enhance the tumorigenicity of hepatocellular carcinoma cells A, experimental results of xenotransplantation in vivo; B, Western blotting assay results of transplanted tumor. HDAC, histone deacetylase; GAPDH, glyceraldehyde-3-phosphatedehydrogenase. H3K27ac, histone H3 lysine 27 acetylation; PI3K/PTEN/AKT/mTOR, phosphoinositide 3-kinases/phosphatase and tensin homolog deleted on chromosome ten/protein kinase B/mammalian target of rapamycin. *P < 0.05, vs. empty group. #P < 0.05, vs. iHDAC2 group."

Figure 6

Specific mechanism of HDAC2-mediated H3K27 acetylation modification in promoting the progression of hepatocellular carcinoma HDAC, histone deacetylase; H3K27ac, histone H3 lysine 27 acetylation; PI3K/PTEN/AKT/mTOR, phosphoinositide 3-kinases/phosphatase and tensin homolog deleted on chromosome ten/protein kinase B/mammalian target of rapamycin."

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