Perioperative dynamic alterations in peripheral regulatory T and B cells in patients with hepatocellular carcinoma

Abstract

Background: Intratumoral and circulating regulatory T cells (Tregs) have been shown to be critical in the pathogenesis of hepatocellular carcinoma (HCC). However there is limited knowledge on the alterations of regulatory B cells (Bregs). We here investigated perioperative dynamic alterations of peripheral circulating Tregs and Bregs in HCC patients to reveal the relationship between regulatory lymphocytes and its clinical implications.

Methods: 36 patients with HCC, 6 with chronic hepatitis B infection and 10 healthy donors were enrolled for this study. Frequencies of peripheral Tregs and Bregs were measured by flow cytometry with antibodies against CD4, CD25, CD127, CD19 and IL-10 before, and after radical surgery. Then, clinical informatics of HCC patients was achieved through Digital Evaluation Score System (DESS) for the assessment of disease severity. Finally, we analysed correlations between digitalized clinical features and kinetics of circulating regulatory lymphocytes.

Results: Level of circulating CD4+CD25+CD127- Tregs in HCC patients was significantly lower than that in healthy donors and patients with chronic hepatitis B infection before surgery, but was increased after surgery. Preoperative level of CD19+ IL-10+ Bregs in HCC patients was also significantly lower than the other groups. However it dramatically was elevated right after surgery and remained elevated compared to controls (about 7 days after surgery, P = 0.04). Frequency of circulating Tregs was correlated with circulating leukocytes, ferritin, and clinical features suggesting tumor aggressiveness including portal vein thrombosis, hepatic vein involvement and advanced clinical stages. Frequency of circulating Bregs was associated with Hepatitis B e Antigen (HBeAg) and Hepatitis B virus (HBV) DNA copy number. In addition, DESS was significantly and positively correlated with other staging systems.

Conclusion: Frequencies of peripheral Tregs and Bregs in HCC patients increased after surgery. These results suggest that a postoperative combination of therapies against Tregs and Bregs may be beneficial for better outcome of HCC patients after resection.

Figure 1

Frequencies of peripheral regulatory T cells of different study groups. Frequencies of peripheral regulatory T cells were analysed in the healthy donor (Healthy), patients with chronic hepatitis B infection (CHB), and patients with hepatocellular carcinoma (HCC) before surgery (HCCb), 1-2 days (HCCa1) or about 7 days (HCCa7) after surgery. Representative flow cytometry analysis of peripheral CD4⁺CD25⁺CD127^- populations were measured and presented (A). Frequency of CD4⁺CD25⁺CD127^- cells of total CD4⁺ T cells in the peripheral blood were calculated (B). The mean ± SEM is shown,* and ** stand for P < 0.05 and 0.01, respectively.

Figure 2

Frequencies of peripheral regulatory B cells of different study groups. Frequencies of peripheral regulatory B cells were analysed in the healthy donor (Healthy), patients with chronic hepatitis B infection (CHB), and patients with hepatocellular carcinoma (HCC) before surgery (HCCb), 1-2 days (HCCa1) or about 7 days (HCCa7) after surgery. Representative flow cytometry analysis of peripheral IL10⁺CD19⁺ cells were measured and presented (A). Frequency of IL10⁺CD19⁺ cells of total CD19⁺ B cells in the peripheral blood (B). The mean ± SEM is shown, *, **and *** stand for P < 0.05, 0.01 and 0.001 respectively.

Figure 3

Perioperative kinetics of peripheral lymphocytes of patients with hepatocellular carcinoma. Frequency (A) and numbers (B) of lymphocytes in patients with hepatocellular carcinoma (HCC) before surgery (HCCb), 1-2 days (HCCa1) or about 7 days (HCCa7) after surgery. The mean ± SEM is shown, ** and *** stand for P < 0.01 and 0.001, respectively.