Objective:To screen the differentially expressed exosomal miRNAs derived from liver cancer stem cells (LCSCs) and its effect on the malignant biological characteristics of liver cancer cells.Methods:miRNA expression profile chip was used to analyze the differentially expressed exosomal miRNA derived from LCSCs. The effects of miRNA on malignant phenotypes of LCSCs were identified. The cells were further treated with doxorubicin at different concentrations (0, 150, 300 μmol/L), and the expression level of miR-196a was detected by quantitative real-time PCR (qRT-PCR). The apoptosis of liver cancer cells cultured by exosomes derived from LCSCs (Exo-NC group) and exosomes derived from miR-196a inhibited LCSCs (Exo-Inhibitor group) and the activity of caspase3/7 under the action of exosomes from LCSCs were detected. Nude mice were randomly divided into Do-PBS group, Do-Exo-Inhibitor group and Do-Exo-NC group using random number table method, with 5 mice in each group, and the effect of miR-196a on nude mice xenograft tumor model with liver cancer cells was analyzed.Results:In this study, exosomes were isolated and purified from CD133 + Huh7 stem cell culture supernatant. miR-7162-3p, miR-1910-5, miR-3613-3p, miR-196a and miR-155-5p were up-regulated, while miR-1246 and miR-3613-5p were down-regulated. miR-7162-3p, miR-196a and miR-155-5p in exosomes had important effects on the self-renewal ability of LCSCs. miR-1910-5p, miR-196a and miR-155-5p had important effects on the invasion ability of liver cancer stem cells, among which miR-196a had the most significant inhibitory effect. Treatment for 24 h, the miR-196a expression level of the 0, 150 and 300 μmol/L doxorubicin was 0.96±0.05, 1.23±0.05 and 2.33±0.03 respectively, with a statistically significant difference ( F=996.90, P<0.001). Treatment for 48 h, the miR-196a expression level of the 0, 150 and 300 μmol/L doxorubicin were 1.02±0.07, 2.35±0.05 and 2.89±0.55 respectively, with a statistically significant difference ( F=303.00, P<0.001). When the concentration of doxorubicin was 0 and 300 μmol/L, the apoptosis rates of the Exo-NC group were 9.37%±0.19% and 11.64%±0.27%, and those of the Exo-Inhibitor group were were 18.80%±1.91% and 22.79%±1.57%, with statistically significant differences ( t=4.41, P=0.048; t=4.96, P=0.038). When doxorubicin was not used, the ratios of caspase3/7 in the Exo-NC group at 24 h and 48 h were 0.94±0.08 and 0.97±0.09, and those in the Exo-Inhibitor group were 1.56±0.01 and 1.58±0.01, with statistically significant differences ( t=11.41, P=0.008; t=6.07, P=0.026). Under 300 μmol/L doxorubicin, the ratios of caspase3/7 in the Exo-NC group at 24 h and 48 h were 0.95±0.07 and 1.36±0.08, and those in the Exo-Inhibitor group were 2.84±0.08 and 3.20±0.14, with statistically significant differences ( t=24.20, P=0.002; t=15.78, P=0.004). The results of xenograft tumor in nude mice showed that the tumor volumes of Do-PBS, Do-Exo-Inhibitor and Do-Exo-NC groups increased successively, which were (1 051.86±89.90) mm 3, (1 310.91±86.66) mm 3 and (2 185.14± 352.34) mm 3 respectively, with a statistically significant difference ( F=30.28, P<0.001). The weights of the transplanted tumors in the 3 groups increased successively, which were (0.36±0.10) g, (0.39±0.12) g and (0.76±0.16) g respectively, with a statistically significant difference ( F=11.81, P=0.002). The expression of miR-196a in tumors was significantly decreased after miR-196a inhibitor transfection. The expression levels of the 3 groups were 1.05±0.16, 0.38±0.08 and 2.17±0.26, with a statistically significant difference ( F=48.93, P<0.001). Conclusion:The exosomal secreted by LCSCs can enhance the resistance of liver cancer cells to doxorubicin by miR-196a.

Objective:To establish a cell line stably expressing the transient receptor potential melastatin 2(TRPM2)channel for screening TRPM2 inhibitors based on PiggyBac transposition system.Methods:A plasmid PiggyBac-human TRPM2(pPB-hTRPM2)eukaryotic expression vector was constructed using PiggyBac transposition system.The plasmid and a helper plasmid were co-transfected into HEK293T cells to express TRPM2,which was identified by fluorescence and patch-clamp assays.The high throughput screening performance was assessed with the Z'factor.Calcium imaging and patch clamp techniques were employed to assess the initial activity of eleven compound molecules,confirming the inhibitory effects of the primary molecules on TRPM2.The protective effect of the screened compounds on damaged cells was validated using the oxygen-glucose deprivation/reperfusion(OGD/R)injury model and CCK-8 kit.The level of cellular reactive oxygen species(ROS)was detected by flow cytometry.The neuroprotective effects of the compounds were evaluated using a transient middle cerebral artery occlusion(tMCAO)mouse model.Results:The HEK293T cells transfected with pPB-hTRPM2-EGFP showed high TRPM2 expression.Puromycin-resistant cells,selected through screening,exhibited robust fluorescence.Whole-cell patch results revealed that induced cells displayed classical TRPM2 current characteristics comparable to the control group,showing no significant differences(P＞0.05).With a Z'factor of 0.5416 in calcium imaging,the model demonstrated suitability for high-throughput screening of TRPM2 inhibitors.Calcium imaging and electrophysiological experiments indicated that compound 6 significantly inhibited the TRPM2 channel.Further experiments showed that 1.0 μmol/L of compound 6 enhanced cell viability(P＜0.05)and reduced the level of ROS(P＜0.05)of SH-SY5Y under OGD/R injury.0.3 and 1.0 mg/kg of compound 6 reduced the cerebral infarction volume in tMCAO mice(both P＜0.05).Conclusions:A stable TRPM2 gene expressing cell line has been successfully established using PiggyBac gene editing in this study.TRPM2 channel inhibitors were screened through calcium imaging and patch clamp techniques,and an inhibitor compound 6 was identified.This compound can alleviate cell damage after OGD/R by reducing cellular ROS levels and has a protective effect against cerebral ischemia-reperfusion injury in mice.

Objective To compare the efficacy of endoscopic bougie/balloon dilation(EBD),endoscopic radial incision(ERI),and ERI combined with esophageal stent placement(ESP)for the treatment of benign esophageal stenosis,and evaluate the feasibility and safety of ERI combined with ESP for the treatment of benign esophageal stenosis.Methods 48 Patients with benign esophageal stenosis from January 2019 to January 2023 were recruited,and divided into EBD group(n=24),ERI group(n=17)and ERI+ESP group(n=7).The differences in operating success,restenosis and complications among the three groups were compared.Results The number of previous endoscopic treatment in ERI+ESP group was more than that in EBD group and ERI group,and the differences were statistically significant(P＜0.05).Technical success was achieved in 23 cases and clinical remission in 23 cases in EBD group,technical success in 16 cases and clinical remission in 15 cases in ERI group,technical success in 7 cases and clinical remission in 7 cases in ERI+ESP group.There was no significant difference in technical success rate and clinical remission rate among the three groups(P＞0.05).After 3 months of follow-up,there were 15,9 and 1 cases of esophageal restenosis in the EBD group,ERI group and ERI+ESP group,respectively.There was no significant difference in the rate of esophageal restenosis among the 3 groups(P＞0.05).After 6 months of follow-up,there were 20 cases of esophageal restenosis in the EBD group,13 cases in the ERI group and 1 case in the ERI+ESP group.The rate of esophageal restenosis in the ERI+ESP group was significantly lower than that in the EBD group and the ERI group(P＜0.05).However,there was no statistically significant difference in the esophageal restenosis rate between the EBD group and the ERI group(P＞0.05).The time to the first postoperative restenosis was 74.00(48.75,159.00)days in the EBD group,84.00(54.50,195.00)days in the ERI group,and 250.00(206.00,289.00)days in the ERI+ESP group.The time to the first postoperative restenosis was longer in the ERI+ESP group than that in the EBD and ERI groups.The differences were statistically significant(P＜0.05),but there was no significant difference in restenosis time between EBD group and ERI group(P＞0.05).There were 5,5 and 3 cases of complications in the EBD group,ERI group and ERI+ESP group,respectively,and there was no significant difference in the incidence of complications among the three groups(P＞0.05).Conclusion ERI+ESP is comparable to EBD and ERI in terms of technical success and short-term clinical remission rate for the treatment of benign esophageal stenosis,and is superior to EBD and ERI in terms of long-term restenosis rate and restenosis time,with no influence on the occurrence of complications.

Oral erythema is a rare and potentially malignant disease of oral mucosa with a high tendency to cancer.Traditional surgery is difficult to remove the lesion while preserving the beauty and function of the affected area.Erbium laser is widely used in the treatment of oral diseases and has certain advantages in the treatment of oral erythema.This paper reports a case of excision of large area of oral erythema with erbium laser fractional treatment.

ObjectiveTo visually analyze the current status, hot spots and frontiers of the researches on transcranial magnetic stimulation for stroke in recent five years. MethodsLiterature related to transcranial magnetic stimulation for stroke were retrieved from the Web of Science Core Collection from January 1, 2018 to December 31, 2022, and a visualized analysis was performed using CiteSpace 6.1.R6 software. ResultsThe number of published papers increased year by year, the most prolific author was Abo Masahiro, the country with the largest number of articles was China, and Sun Yat-sen University published the most papers. The hot keywords were cerebral cortex, functional magnetic resonance imaging, cortical excitability, plasticity, etc. The top five keywords of bursting strength were unilateral spatial neglect, poststroke, neuropathic pain, cortical plasticity, and trancallosal inhibition. ConclusionThe popularity of studies on transcranial magnetic stimulation in the treatment of stroke is increasing year by year. In the future, attention can be paid to the therapeutic effect of transcranial magnetic stimulation on unilateral spatial neglect, neuropathic pain, etc. Additionally, the influence of transcranial magnetic stimulation on cortical plasticity can be further studied.