Objective To investigate the changes of regulatory T cells (Treg) in patients with hepatocellular carcinoma (HCC) after ultrasound- guided percutaneous cool- tip radiofrequency ablation (RFA), and to discuss its influence on the prognosis. Methods A total of 30 patients with HCC were enrolled in this study. The percentage of Treg in peripheral blood was estimated with flow cytometry before RFA and one, 4, 7 and 12 months after RFA. During the follow-up period, the therapeutic effects were evaluated by contrast enhanced sonography or contrast enhanced CT scanning. By using the methods of receiver operating characteristic (ROC) curve and Kaplan-Meier survival function, the correlation of Treg dynamic changes with the progression-free survival time was analyzed. Results One month after RTA, the tumor response (TR) rate in the 30 patients was 93.3% (28/30), the tumor progression (TP) rate was 6.67%(2/30). The percentage of Treg before RFA was (9.42 ± 1.16)%, which decreased to (6.55 ± 0.97)% one month after RFA, the difference was statistically significant (t = 15.325, P <0.001). Twelve months after RFA, TR rate became 33.3%(10/30), and TP rate became 66.7%(20/30). The preoperative percentage of Treg of TR group was (8.75 ± 0.72)%, which was significantly lower than that of TP group (9.76 ± 1.20)%, the difference was statistically significant (t=-2.448, P=0.021). ROC curves indicated that the optimal cut-off value of Treg nadir was 4.82%, the sensitivity was 90.0% and the specificity was 60.0%. The optimal cut-off time to reach Treg nadir was 5.5 months, the sensitivity was 70.0% and the specificity was 85.0%. Kaplan-Meier curve analysis showed that after RFA the progression-free survival rate (PFS) of patients with Treg nadir ≤ 4.82% was significantly higher than that of patients with Treg nadir>4.82%. PFS of patients with reaching Treg nadir≥5.5 months was significantly higher than that of patients with reaching Treg nadir<5.5 months. Log-rank test results were字2=5.207, P=0.023; 字2=22.079, P < 0.001, respectively. Conclusion Percutaneous cool-tip radiofrequency ablation can decrease the percentage of Treg cells. Besides, Treg nadir and the time reaching Treg nadir can reflect the prognosis of HCC patients after RFA to a certain extent.

Background There are a variety of microorganisms in ambient air, and susceptible people can be infected once contact with pathogenic microorganisms in the environment. In order to avoid the spread of pathogenic bacteria, disinfection is the simplest and most effective way of killing pathogenic bacteria in the environment to block the contact between pathogenic bacteria and humans. Sodium hypochlorite (NaClO) is the most widely used disinfectant, but its safety in ambient air disinfection is not clear yet. Objective To establish a model of bronchial epithelial cell (BEAS-2B) injury induced by NaClO, and to explore the mechanism of the toxic effect of NaClO disinfectants on BEAS-2B. Methods Cells were treated with concentration gradients of 0, 25, 50,100, 200, and 400 μmol·L⁻¹ of the diluted NaClO (100 mmol·L⁻¹) standard solution, respectively, and cell activity was measured by cell counting kit-8 (CCK-8) assay after 15 and 30 min. Cells treated with 0, 25, and 50 μmol·L⁻¹ NaClO were selected to observe the cell morphology under an inverted microscope, apoptosis was determined by flow cytometry Annexin V FITC / PI double staining to determine the final experimental concentration. The morphology of organelles such as mitochondria was observed under a transmission electron microscope. Mitochondrial membrane potential of the cells was detected by JC-1 staining. Intracellular Ca²⁺ concentration was measured with a Fluo-4 AM fluorescent probe. Total cellular reactive oxygen species (ROS) was detected with a 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescent probe, cell mitochondrial ROS with a dihydroethidium (DHE) fluorescent probe, and lipid peroxidation intermediate malondialdehyde (MDA) with a commercial kit. Results Compared with 0 μmol·L⁻¹, NaClO treatment group, cell morphology did not change a lot after 25 μmol·L⁻¹ NaClO treatment for 30 min, and the cells began to wrinkle and become round after 30 min treatment with 50 μmol·L⁻¹ NaClO, showing about 70% of normal cell viability (P<0.01). So 30 min 50 μmol·L⁻¹ NaClO treatment was selected for the subsequent experiment. The experimental results found that compared with the 0 μmol·L⁻¹ NaClO treatment group, the number of apoptotic cells increased (P<0.05), the mitochondrial membrane potential decreased (P<0.01), the intracellular Ca²⁺ concentration increased (P<0.05), the cellular ROS level increased (P<0.05), the mitochondrial ROS level increased (P<0.01), and the MDA content increased (P<0.01) in the NaClO treatment group.. Conclusion The study has successfully established a model of BEAS-2B injury induced by NaClO, and found that NaClO can lead to cell damage by inducing apoptosis and oxidative stress in BEAS-2B cells. According to the results, there are two possible reasons. First, NaClO solves in water to form hypochlorous acid (HClO) which is oxidative and increases the intracellular ROS level after entering cells, leading to cellular oxidative stress. Second, HClO enters cells to directly attack the mitochondrial membrane, resulting in the imbalance of potential inside and outside the mitochondrial membrane, and apoptosis caused by Ca²⁺ efflux.