Objective: To investigate the changes in CD8+CD28–Foxp3+ regulatory T cells (CD8+CD28–Foxp3+Tregs) in peripheral blood and tumor tissue of patients with hepatocellular carcinoma (HCC) and the significance. Methods: The ratio of CD8+CD28–Foxp3+Tregs to CD8+T cells (CD8+CD28–Foxp3+Tregs/CD8+T) in peripheral blood in 72 HCC patients and 22 healthy controls was determined by flow cytometry, and the relations of CD8+CD28–Foxp3+Tregs/CD8+T with clinicopathologic factors of the HCC patients were analyzed. The Foxp3 positive cells and Foxp3 protein expression in tumor tissues and their adjacent tissues from HCC patients were determined by immunohistochemical staining and Western blot analysis, respectively. Results: The peripheral blood CD8+CD28–Fop3+Tregs/CD8+T was significantly increased in HCC patients compared with that in healthy controls (P<0.05), and the peripheral blood CD8+CD28–Fop3+Tregs/CD8+T was significantly associated with the TNM stage, lymph node metastasis and degree of tumor differentiation of the HCC patients (all P<0.05). The average number of Foxp3 positive cells and Foxp3 protein expression level were significantly higher in tumor tissue than those in adjacent tissue (both P<0.05). The peripheral blood CD8+CD28–Fop3+Tregs/CD8+T, and number of Foxp3 positive cells and Foxp3 protein expression level in tumor tissue all presented an increasing trend in the order of well, moderately and poorly differentiated HCC, but all differences did not reach a statistical significance (all P>0.05). Conclusion: CD8+CD28–Fop3+Tregs are increased in the peripheral blood and tumor tissue of HCC patients, which may probably be related to tumor immunosuppression, and their detection may have certain value in assessing the disease states of the HCC patients.