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. 2021 Jun 10;12:645835. doi: 10.3389/fimmu.2021.645835

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Copyright © 2021 Zhang, Yang, Liu, Zhang and Jin

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Liver-infiltrating non-specific and HBV core-specific Th9/Tc9 cells in hepatitis B-related hepatocellular carcinoma (HCC). Intrahepatic lymphocytes (IHLs) were isolated from fresh HCC specimens and non-tumor site liver specimens in thirteen HCC patients (seven in stage A and six in stage B) who underwent hepatic carcinectomy. IHLs were stimulated with either phorbol myristate acetate (50 ng/ml)+ionomycin (1 μg/ml) (for non-specific stimulation) or HBsAg (for HBV-specific stimulation) in the presence of brefeldin A (10 μg/ml) for 6 hours, and were then stained with anti-CD3, anti-CD4, anti-CD8, and anti-IL-9. CD3⁺CD4⁺IL-9⁺ Th9 cells and CD3⁺CD8⁺IL-9⁺ Tc9 cells were analyzed by flow cytometry. (A) The percentage of liver-infiltrating non-specific Th9 cells was compared between non-tumor site and tumor site. Significance was assessed using Student t test. (B) The percentage of liver-infiltrating HBV-specific Th9 cells was compared between non-tumor site and tumor site. Significance was assessed using Student t test. (C) The percentage of liver-infiltrating non-specific Tc9 cells was compared between non-tumor site and tumor site. Significance was assessed using Student t test. (D) The percentage of liver-infiltrating HBV-specific Tc9 cells was compared between non-tumor site and tumor site. Significance was assessed using Student t test.