ABSTRACT
Objective:To investigate the regulatory effects of ring finger protein 43 (RNF43) on CD8 + T cell-mediated anti-tumor immune reaction in melanoma. Methods:RNF43 gene was over-expressed and knockdown in mouse melanoma cells line B16-OVA by lentivirus infection; In vivo proliferation of mouse melanoma cells line B16-OVA in the Lv-Ctrl-OE, Lv-RNF43-OE, Lv-Ctrl-KD and Lv-RNF43-KD groups was detected by subcutaneous tumorigenesis assay in mice, and the expression levels of CD8 + T cells perforin and interferon γ (IFN-γ) in tumor immune microenvironment of melanoma were detected by flow cytometry; The expression levels of β-catenin and programmed death-ligand 1 (PD-L1) mRNA in cells were detected by quantitative real-time PCR assay; The effect of RNF43 on the transcriptional regulation of PD-L1 was detected by dual-luciferase reporter gene assay. Results:Stable RNF43 over-expressing and RNF43 knockdown mouse melanoma cells lines Lv-RNF43-OE and Lv-RNF43-KD were successfully constructed. The results of subcutaneous tumorigenesis experiment in mice showed that the tumor mass of the Lv-RNF43-OE group was (0.08±0.06) g, which was significantly smaller than that of the Lv-Ctrl-OE group [ (1.04±0.52) g], with a statistically significant difference ( t=3.71, P=0.032) ; The tumor mass of Lv-RNF43-KD group was (1.94±0.29) g, with no statistically significant difference ( t=-1.70, P=0.164) compared with that of the Lv-Ctrl-KD group (1.15±0.74) g. The flow cytometry results showed that the fluorescence intensity of CD8 + T cell perforin in the Lv-RNF43-OE group was 9 034 ± 2 628, which was significantly higher than that in the Lv-Ctrl-OE group (3 847 ±1 637), with a statistically significant difference ( t=-3.35, P=0.015) ; The fluorescence intensity of CD8 + T cell perforin in the Lv-RNF43-KD group was 966±247, which was significantly lower than that in the Lv-Ctrl-KD group (2 226±646), with a statistically significant difference ( t=3.16, P=0.034) ; The fluorescence intensity of IFN-γ of CD8 + T cell in the Lv-RNF43-OE group was 2 422±429, which was significantly higher than that of 1 688±324 in the Lv-Ctrl-OE group, with a statistically significant difference ( t=-2.73, P=0.034) ; The fluorescence intensity of IFN-γ of CD8 + T cell in the Lv-RNF43-KD group was 614 (454, 863), with a statistically significant difference ( Z=-1.96, P=0.050) compared with 1 159 (1 152, 2 068) in the Lv-Ctrl-KD group. The results of quantitative real-time PCR showed that the relative expression level of β-catenin mRNA in the Lv-RNF43-OE group was 0.67±0.16, which was significantly lower than that of 1.00±0.11 in the Lv-Ctrl-OE group, with a statistically significant difference ( t=2.98, P=0.041) ; The relative expression level of PD-L1 mRNA in the Lv-RNF43-OE group was 0.32±0.09, which was significantly lower than that of 1.00±0.09 in the Lv-Ctrl-OE group, with a statistically significant difference ( t=9.13, P=0.001). The results of the dual-luciferase reporter gene assay showed that the PD-L1 promoter luciferase activity in the pCMV6-NC, RNF43, RNF43+β-catenin and β-catenin groups were 1.00±0.00, 0.84±0.00, 1.49±0.00 and 1.57±0.03 ( F=2 218.33, P<0.001). Further pairwise comparison showed that compared with the pCMV6-NC group, PD-L1 promoter luciferase activity was significantly lower in the RNF43 group ( P<0.001) and significantly higher in the RNF43+β-catenin and β-catenin groups ( P<0.001; P=0.003) ; compared with the RNF43 group, PD-L1 promoter luciferase activity was significantly higher in the RNF43+β-catenin group ( P<0.001) . Conclusion:RNF43 may reduce the expression of PD-L1 mRNA in melanoma by inhibiting the expression of β-catenin and promote CD8 + T cell-mediated anti-tumor immune reaction.
ABSTRACT
BackgroundCurrently, the prevention and control of the novel coronavirus disease (COVID-19) outside Hubei province in China, and other countries have become more and more critically serious. We developed and validated a diagnosis aid model without computed tomography (CT) images for early identification of suspected COVID-19 pneumonia (S-COVID-19-P) on admission in adult fever patients and made the validated model available via an online triage calculator. MethodsPatients admitted from Jan 14 to February 26, 2020 with the epidemiological history of exposure to COVID-19 were included [Model development (n = 132) and validation (n = 32)]. Candidate features included clinical symptoms, routine laboratory tests, and other clinical information on admission. Features selection and model development were based on the least absolute shrinkage and selection operator (LASSO) regression. The primary outcome was the development and validation of a diagnostic aid model for S-COVID-19-P early identification on admission. ResultsThe development cohort contained 26 S-COVID-19-P and 7 confirmed COVID-19 pneumonia cases. The final selected features included 1 variable of demographic information, 4 variables of vital signs, 5 variables of blood routine values, 7 variables of clinical signs and symptoms, and 1 infection-related biomarker. The model performance in the testing set and the validation cohort resulted in the area under the receiver operating characteristic (ROC) curves (AUCs) of 0.841 and 0.938, the F-1 score of 0.571 and 0.667, the recall of 1.000 and 1.000, the specificity of 0.727 and 0.778, and the precision of 0.400 and 0.500. The top 5 most important features were Age, IL-6, SYS_BP, MONO%, and Fever classification. Based on this model, an optimized strategy for S-COVID-19-P early identification in fever clinics has also been designed. ConclusionsS-COVID-19-P could be identified early by a machine-learning model only used collected clinical information without CT images on admission in fever clinics with a 100% recall score. The well-performed and validated model has been deployed as an online triage tool, which is available at https://intensivecare.shinyapps.io/COVID19/.
ABSTRACT
Objective To explore the therapeutic effect, mechanism and safety of Qingrehuashi recipe in treating chronic superficial gastritis with erosion pertaining to spleen-stomach damp-heat syndrome. Methods 84 cases of chronic superficial gastritis with erosion were recruited into a control group and a treatment group randomly, with each groups containing 42 cases. The treatment group was treated with Qingrehuashi recipe, while the control group was treated with Qingweizhitong granule. Both groups were treated for 8 weeks. The clinical manifestations and signs, gastroscopic results and pathological effects were observed before and after the treatment. Results The total effective rate and curative rate in the treatment group were superior to those in the control group (P<0.05) . Conclusion Qingre Huashi recipe is effective and safe in in treating chronic superficial gastritis with erosion pertaining to spleen-stomach damp-heat syndrome. The mechanism might be related to its effects of anti-inflammation, anti-oxidation, eradicating Hp, promoting gastric motility and enhancing gastric mucosal barrier.
