ObjectiveTo investigate the value of combined determination of hepatitis C virus (HCV) genotype, the alpha-fetoprotein variant AFP-L3, and P53 antibody in HCV-related hepatocellular carcinoma (HCV-HCC). MethodsA total of 84 patients with HCV-HCC who were diagnosed in our hospital from January 2016 to December 2019 were enrolled as HCV-HCC group, and 84 patients with benign liver diseases (hepatitis C and HCV liver cirrhosis) were enrolled as control group. The PCR-reverse dot blot hybridization technique was used to determine HCV genotype, ELISA was used to measure P53 antibody, and electrochemical luminescence was used to measure AFP-L3. The t-test and the Kruskal-Wallis H test were used for comparison between two groups; the chi-square test was used for comparison of categorical data between two groups. The logistic regression analysis and the receiver operating characteristic (ROC) curve were used to compare the value of each index in the diagnosis of HCV-HCC. ResultsCompared with the control group, the HCV-HCC group had a significantly higher proportion of patients with HCV 1b genotype or AFP-L3 and a significantly higher level of P53 antibody (χ2=5714, Z=-9.27, Z=-9.92, all P＜0.05). The logistic regression analysis showed that HCV genotype, AFP-L3, and P53 antibody had significant effects on HCV-HCC (all P＜0.05). The above indices were fitted to establish a model of Logit(Y)=-3.881+0031XAFP-L3(%)+0.043XP53+1218XHCV genotype, in which Y was the positive probability value of combined determination. In the screening of HCV-HCC, Y had a significantly larger area under the ROC curve than HCV genotype (0.945 vs 0.758, Z=6.17, P＜0001), AFP-L3 (0.945 vs 0.863, Z=3.97, P＜0.001), and P53 antibody (0.945 vs 0.887, Z=3.07, P=0.002). Y had higher AUC (0.945), sensitivity (90.90%), specificity (94.00%), positive predictive value (93.80%), negative predictive value (9116%), and diagnostic accuracy (92.44%) than each index alone. ConclusionHCV 1b genotype, AFP-L3, and P53 antibody level are associated with the risk of HCV-HCC, and the combined determination of the three indices has important clinical significance in the early diagnosis of HCV-HCC.

A purified polysaccharide with a galactose backbone(SPR-1,Mw 3,622 Da)was isolated from processed Polygonati Rhizoma with black beans(PRWB)and characterized its chemical properties.The backbone of SPR-1 consisted of[(4)-β-D-Galp-(1]9→ 4,6)-β-D-Galp-(1 → 4)-α-D-GalpA-(1 → 4)-α-D-GalpA-(1 →4)-α-D-Glcp-(1 → 4,6)-α-D-Glcp-(1 → 4)-α/β-D-Glcp,with a branch chain of R1:β-D-Galp-(1 → 3)-β-D-Galp-(1 → connected to the →4,6)-β-D-Galp-(1 → via O-6,and a branch chain of R2:α-D-Glcp-(1 →6)-α-D-Glcp-(1 → connected to the →4,6)-α-D-Glcp-(1 → via O-6.Immunomodulatory assays showed that the SPR-1 significantly activated macrophages,and increased secretion of NO and cytokines(i.e.,IL-1β and TNF-α),as well as promoted the phagocytic activities of cells.Furthermore,isothermal titration calorimetry(ITC)analysis and molecular docking results indicated high-affinity binding between SPR-1 and MD2 with the equilibrium dissociation constant(KD)of 18.8 μM.It was suggested that SPR-1 activated the immune response through Toll-like receptor 4(TLR4)signaling and downstream responses.Our research demon-strated that the SPR-1 has a promising candidate from PRWB for the TLR4 agonist to induce immune response,and also provided an easily accessible way that can be used for PR deep processing.

[摘 要] 目的：探讨胶原三螺旋重复蛋白1（CTHRC1）在膀胱癌组织和细胞中的表达及其对膀胱癌5637细胞迁移和侵袭的影响及其机制。方法：利用TCGA和Arrayexpress数据库中膀胱癌基因表达数据，分析CTHRC1转录和翻译水平。收集2014年9月至2020年12月重庆医科大学附属第一医院手术切除的144例膀胱癌组织和25例全膀胱切除的癌旁组织标本，以及人膀胱癌细胞RT4、5637、T24、UMUC-3、TCCSUP和输尿管上皮永生化细胞SV-HUC-1。采用免疫组织化学染色法、qPCR法和WB法检测膀胱癌组织和细胞中CTHRC1的表达水平，通过Kaplan-Meier曲线分析CTHRC1表达对总生存期（OS）的影响。运用RNAi技术，敲降5637细胞CTHRC1表达后，通过细胞划痕实验和Transwell实验检测CTHRC1表达下调对5637细胞迁移和侵袭的影响。利用基因集富集分析（GSEA）预测CTHRC1相关的潜在信号通路，WB法检测敲降CTHRC1表达对FAK-ERK1/2通路相关蛋白表达的影响。结果：CTHRC1的转录和翻译水平在肌层浸润性膀胱癌（MIBC）组织和细胞中表达显著上调（均P<0.05），CTHRC1高表达组患者5年OS较低表达患者缩短（P<0.05）。干扰CTHRC1表达后，膀胱癌5637细胞迁移及侵袭能力均显著降低（均P<0.01）。GSEA预测显示，CTHRC1高表达组主要富集在黏着斑激酶（FAK）、肌动蛋白细胞骨架调节、FAK和ERK1/2信号通路。WB法实验结果表明，重组CTHRC1蛋白促进膀胱癌5637细胞FAK-ERK1/2信号通路活化（P<0.05或P<0.01）。结论：CTHRC1在MIBC中表达上调，且与膀胱癌患者不良预后密切相关；CTHRC1促进膀胱癌细胞迁移和侵袭，该过程可能与FAK-ERK1/2信号通路的激活有关。