Background::Liver cancer is largely resistant to chemotherapy. This study aimed to identify the effective chemotherapeutics for β-catenin-activated liver cancer which is caused by gain-of-function mutation of catenin beta 1 ( CTNNB1), the most frequently altered proto-oncogene in hepatic neoplasms. Methods::Constitutive β-catenin-activated mouse embryonic fibroblasts (MEFs) were established by deleting exon 3 ( β-cateninΔ(ex3)/+ ), the most common mutation site in CTNNB1 gene. A screening of 12 widely used chemotherapy drugs was conducted for the ones that selectively inhibited β-cateninΔ(ex3)/+ but not for wild-type MEFs. Untargeted metabolomics was carried out to examine the alterations of metabolites in nucleotide synthesis. The efficacy and selectivity of methotrexate (MTX) on β-catenin-activated human liver cancer cells were determined in vitro. Immuno-deficient nude mice subcutaneously inoculated with β-catenin wild-type or mutant liver cancer cells and hepatitis B virus ( HBV); β-cateninlox(ex3)/+ mice were used, respectively, to evaluate the efficacy of MTX in the treatment of β-catenin mutant liver cancer. Results::MTX was identified and validated as a preferential agent against the proliferation and tumor formation of β-catenin-activated cells. Boosted nucleotide synthesis was the major metabolic aberration in β-catenin-active cells, and this alteration was also the target of MTX. Moreover, MTX abrogated hepatocarcinogenesis of HBV; β-cateninlox(ex3)/+ mice, which stimulated concurrent Ctnnb1-activated mutation and HBV infection in liver cancer. Conclusion::MTX is a promising chemotherapeutic agent for β-catenin hyperactive liver cancer. Since repurposing MTX has the advantages of lower risk, shorter timelines, and less investment in drug discovery and development, a clinical trial is warranted to test its efficacy in the treatment of β-catenin mutant liver cancer.

Fragile X syndrome(FXS)is caused by abnormal duplication and amplification of the FMR1 gene CGG.This article reports a pair of brothers diagnosed with FXS by genetic testing.Two patients,aged 15 and 14 years old respectively,both had clinical manifestations such as language disorders,intellectual disabilities,attention deficit disorder,autism spectrum disorder,and FXS's characteristic facial features.The proband had a rare late-onset epileptic seizure,which was well treated with levetiracetam,while his younger brother had no electroencephalogram abnormalities after repeated follow-up.This pair of cases suggests that the clinical phenotype of FXS has diversity and heterogeneity.

Infectious bronchitis in chickens is a serious threat to the global poultry industry.Despite the availability of commercial vaccines,the epidemic has not been effectively controlled.Therefore,the development of novel vaccines may provide new ways to prevent and control this disease.In this study,BLP-S1,a bacterium-like particle displaying the S1 subunit of infectious bronchitis virus on its surface,was constructed using the GEM-PA system.The immunoprotection results showed that BLP-S1 effectively induced the production of specific IgG and sIgA in commercial chickens and provided effective protection against a heterologous strain with a protection rate of up to 90％.This study demonstrated that BLP-S1 has good immunogenicity and immunoprotection,with the poten-tial to develop a novel vaccine against infectious bronchitis.

Objective To investigate a convenient and quantitative solution to activation levels and functional characterization of CAR-T cells by inserting T cell activity-responsive promoter (TARP) nanoluciferase reporter gene system into a lentiviral plasmid containing the gene encoding the chimeric antigen receptor (CAR). Methods The recombinant plasmid was constructed by using whole gene synthesis and molecular cloning techniques. The lentivirus was packaged and was infected with human primary T lymphocytes. Flow cytometry was used to detected the positive rate of lentivirus-infected T cells. The functional characterization of CAR-T cells was identified by luciferase reporter gene system, Western blot, flow cytometry, and small animal live imaging techniques. Results The results of enzyme digestion identification and the plasmid sequencing showed that the recombinant plasmids were constructed, and flow cytometry displayed the normal preparation of CAR-T cells. This system could dynamically respond to the activation of CAR-T cells by luciferase reporter gene system. The functional assay in vitro confirmed that the system could reflect the exhaustion of CAR-T cells, and the small animal live imaging results demonstrated that the system can be used as a tracer of CAR-T cells in mice. Conclusion TARP nanoluciferase reporter gene system provides a more convenient, sensitive and quantitative method for evaluating CAR-T cells activation level, exhaustion phenotype and tracing.
Humans ; Animals ; Mice ; T-Lymphocytes ; Cell Line, Tumor ; Receptors, Chimeric Antigen/genetics* ; Promoter Regions, Genetic ; Immunotherapy, Adoptive/methods*

Objective:To explore the clinical and CT imaging features of immune checkpoint inhibitor-associated pneumonia (CIP) and to improve the early diagnostic ability of CIP.Methods:From June 1, 2020 to October 31, 2021, the clinical data and chest CT images of 2 067 patients with advanced malignant tumor treated with immune checkpoint inhibitor (ICI) in the First Medical Center, Chinese PLA General Hospital were retrospectively analyzed. Patients with CIP were enrolled according to the guidelines for CIP diagnosis, and the incidence, time from the start of medication to the onset of CIP, medication cycle, imaging features, imaging patterns, CT grade and outcomes were analyzed. χ 2 test was used to compare the incidence of CIP in patients with or without basic lung disease. Results:Among 2 067 patients with malignant tumors treated with ICI, 67 patients developed CIP, the incidence of CIP was 3.2%. The incidence of CIP was significantly different between 386 patients with basic lung disease (7.00%, 27/386) and 1 681 patients without basic lung disease (2.4%, 40/1 681) (χ 2=21.32, P<0.001). The time from the start of medication to the onset of CIP was 7-367 d (median 52 days), and the duration of medication was 1-12 cycles (median 2 cycles). The imaging features of CIP presented as ground glass opacities in 54 cases (80.6%), solid nodules in 26 cases (38.8%), consolidations in 25 cases (37.3%) and irregular reticular opacities in 24 cases (35.8%). The main radiologic pattern was organizing pneumonia (OP, 34 cases, 50.7%), and followed by diffuse alveolar damage (DAD) pattern (14 cases, 20.9%). According to CT grading, there were 26 cases in low risk grade, 17 cases in moderate risk grade and 24 cases in high risk grade. Of 43 low-and medium-risk grade cases, 25 were OP pattern, accounting for 58.1%, and among 24 high-risk grade patients, 13 were DAD pattern, accounting for 54.2%. Forty-three of the 52 patients were initially untreated, of which 23 patients progressed, 17 had lesion shrinkage, and 3 had resolution, and relapsed in 8 cases after resolution or drug withdrawal. Conclusions:The imaging manifestations of CIP are mainly ground glass opacities, nodules, consolidations, and irregular reticular opacities. The radiologic patterns are mainly OP and DAD. OP is the most common pattern in low-moderate risk grade CIP and DAD is the most common pattern in high risk grade CIP. Patients with basic lung disease are more likely to get CIP.

Objective:To investigate the complement C3a receptor 1 (C3AR1) expression in glioma and its mechanism in progressing malignancy.Methods:(1) The C3AR1 mRNA expression data and clinical information were obtained in 607 glioma patients from The Cancer Genome Atlas (TCGA) database and 656 glioma patients from Chinese Glioma Genome Atlas (CGGA) database; the differences in C3AR1 mRNA expression were analyzed among gliomas with different World Health Organization (WHO) grading. The overall survival and disease-free survival were compared between high and low C3AR1 mRNA expression patients obtained from TCGA database by Gene expression profiling interactive analysis (GEPIA). Gene body (GO) function analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of C3AR1 related differentially expressed genes were performed by DAVID database. Correlation of C3AR1 mRNA expression with immune cell infiltration was analyzed using TIMER online website. (2) The brain tissues from 3 non-tumor patients and 9 glioma patients surgically resected in Department of Neurosurgery, First Affiliated Hospital of Xinxiang Medical University from January 2019 to September 2021 were collected; the C3AR1 protein expression was detected by Western blotting. (3) The in vitro cultured U87 and U251 cells were divided into negative control group and C3AR1 knockdown group ( C3AR1 being knocked down by lentivirus transfection); and CCK-8 assay, plate cloning assay and Transwell assay were used to detect the proliferation rate, number of colony formation and number of membrane penetrating cells. Western blotting was used to detect the nuclear factor-κB (NF-κB) signaling pathway protein expressions. Results:(1) In TCGA database, the C3AR1 mRNA expression in gliomas of WHO grading II, grading III and grading IV increased sequentially, with significant differences ( P<0.05). In CGGA database, the C3AR1 mRNA expression in glioma of WHO grading IV was statistically higher than that in gliomas of WHO grading II and grading III ( P<0.05). GEPIA showed that the overall survival and disease-free survival in the low C3AR1 mRNA expression group were statistically higher than those in the high C3AR1 mRNA expression group ( P<0.05). GO function analysis and KEGG pathway enrichment analysis revealed that C3AR1 related differentially expressed genes were more enriched in such biological processes and signaling pathways as calcium homeostasis, membrane structural valves, proton transmembrane transporter protein activity, chemokine signaling pathway and NF-κB signaling pathway. TIMER showed that C3AR1 mRNA expression in glioblastoma and low-grade glioma was positively correlated with infiltration degrees of B cells, CD4 + T cells, neutrophils, macrophages and dendritic cells, and C3AR1 mRNA expression in glioblastoma was negatively correlated with infiltration degree of CD8 + T cells ( P<0.05). (2) C3AR1 protein expression in glioma tissues was significantly higher than that in non-tumor tissues. (3) Compared with the negative control group, the C3AR1 knockdown group group had significantly lower proliferation rate, smaller numbers of colony formation and membrane penetrating cells, and lower expressions of NF-κB, phosphorylated (p)-NF-κB, p-NF-κB inhibitory protein (IκB)α, p-I-κB kinase (IKK)α and N-cadherin, and significantly higher E-cadherin expression. Conclusion:C3AR1 is highly expressed in glioma and progresses malignancy through NF-κB signaling pathway.

目的：探讨环状 RNA circ_0091579 作为分子海绵吸附 miR-330-3p 介导环指蛋白 126（ring finger protein 126， RNF126）对结直肠癌（colorectal cancer，CRC）细胞增殖、凋亡、侵袭的影响。方法：选取 2019 年 2 月至 2020 年 5 月在大理大学 第一附属医院行手术治疗的 60 例 CRC 患者的癌组织和癌旁组织。构建 circ_0091579、miR-330-3p 的过表达或敲减的 CRC LoVo 细胞，qPCR 检测 CRC 组织和细胞中 circ_0091579、miR-330-3p 和 RNF126 的表达；MTT、Transwell、流式细胞术分别检测 细胞的增殖、侵袭、凋亡情况；生物信息学方法预测 circ_0091579 和 miR-330-3p、miR-330-3p 和 RNF126 的靶向关系并用双荧光 素酶报告实验和 RNA 免疫沉淀实验验证。结果：CRC 组织和多种细胞（HCT116、SW620、CW-2、LoVo 细胞）中，circ_0091579 和 RNF126 均高表达、miR-330-3p 均低表达（均 P<0.05）。敲减 circ_0091579 可以抑制 LoVo 细胞的增殖、侵袭而促进其凋亡 （均 P<0.05），但该影响在加入 miR-330-3p 后被逆转；过表达 miR-330-3p 使 LoVo 细胞增殖和侵袭能力减弱但凋亡程度加强 （均 P<0.05），该影响在加入 RNF126 后被抵消。circ_0091579、miR-330-3p 和 RNF126 之间存在靶向作用关系。结论：circ_0091579 通过 miR-330-3p/RNF126 轴促进 LoVo 细胞增殖、迁移和侵袭而抑制其凋亡。

Objective To analyze the infiltration abundance of macrophage M2 in breast cancer tissues and explore the correlation between VSIG4 and macrophage M2 and the potential mechanism of regulating the invasion and migration of breast cancer patients. Methods We downloaded the RNA-seq data of TCGA-BRCA and assessed the infiltration abundance of immune cells in the samples by CIBERSORT, and established a prognostic risk prediction model. Then, we analyzed the effect of macrophage M2 and VSIG4 on the prognosis of breast cancer patients. In addition, we analyzed the signaling pathway associated with VSIG4 by gene set enrichment analysis and predicted its upstream regulation of miRNA. Results The infiltration abundance of macrophage M2, age, PR status and pathological stage were involved in the establishment of risk prediction model, and the model had a good prediction performance (AUC=0.816). High infiltration of macrophage M2 (HR=1.35, P < 0.05) and high expression of VSIG4 (HR=1.4, P=0.039) suggested poor prognosis of breast cancer patients. VSIG4 could be regulated by upstream miR-29a-3p and significantly correlated with Toll-like receptor, cell adhesion, production and release of cytokine. Conclusion VSIG4 is significantly associated with breast cancer patients' prognosis and infiltration of macrophage M2, regulated by the upstream miR-29a-3p and promotes the invasion and migration of breast cancer cells. It can be used as a potential prognostic marker for breast cancer.

Objective To study the clinical features of hepatic pseudolymphoma.Methods A retrospective study was conducted on 19 patients with hepatic pseudolymphoma who were diagnosed and treated at Zhongshan Hospital in Shanghai from June 2013 to December 2017.Eighteen patients were females and one was a male.The mean age was (55±9) years,range 36 to 68 years.This study mainly analyzed the imaging features,treatment and postoperative results.Results All patients were diagnosed accidentally,and 78.9％ patients did not exhibit any evidence of hepatic B viral infection.A monofocal lesion was found in 14 patients and multifocal lesions in 5 patients.Surgical treatment was performed in all the patients.The lesion size was (1.1±0.4) cm (range 0.5～2.4 cm).Ultrasound revealed hypo-or slightly hypo-echogenicity.On MRI,diffusion weighted imaging showed slight hyperintensity or hyperintensity,all lesions manifested as homogeneous and slightly hyperintensity on T2WI and hypointensity on T1WI.Dynamic enhancement pattern,wash in and wash out,degressive and persistent enhancement were observed in 16(55.2％),12(41.4％),and 1 (3.4％) patients,respectively.More than 70％ of lesions were diagnosed as malignant tumors on preoperative imaging.During a follow-up of 6 ～ 44 months (median:19 months),no patient developed metastasis or recurrence.Conclusions Hepatic pseudolymphoma commonly occurred in women with a small sized lesion.Due to the lack of specific clinical manifestations and imaging findings,preoperative diagnosis was difficult.Surgical resection is still the most optimal treatment.The patients usually have favorable prognosis.

Objective To evaluate the role of protein kinase C in the maintenance of chronic inflammatory pain in rats and the relationship with the expression of Nav1.8 in the dorsal root ganglion (DRG).Methods Thirty pathogen-free healthy female Sprague-Dawley rats,weighing 180-220 g,were divided into 3 groups using a random number table:control group (group C),chronic inflammatory pain group (group CIP) and PKC inhibitor group (group P).Normal saline 20 μl was injected into the plantar surface of the right hindpaw every day for 14 consecutive days in group C.Prostaglandin E2 100 ng was injected into the plantar surface of the right hindpaw every day for 13 consecutive days to establish the model of chronic inflammatory pain,and dimethyl sulfoxide 20μl was injected into the plantar surface of the right hindpaw on 14th day in group CIP.Prostaglandin E2 100 ng was injected into the plantar surface of the right hindpaw every day for 13 consecutive days,and PKC inhibitor GF1O9203X 100 nmol/20 μl was injected into the plantar surface of the right hindpaw on 14th day in group P.The mechanical paw withdrawal threshold (MWT) was measured at 1 day before injection (T0) and 1,3,7 and 14 days after the last injection (T1-4).The DRGs of the lumbar segment (L4.5) were removed for determination of Nav1.8 expression using immunofluorescence and Western blot.Results Compared with group C,the MWT was significantly decreased at T1-4 in CIP and P groups,and the expression of Nav1.8 in DRGs was significantly up-regulated in group CIP (P＜0.05).Compared with group CIP,the MWT was significantly increased at T4,and the expression of Nav1.8 in DRGs was down-regulated in group P (P＜0.05).Conclusion Up-regulated expression of Nav1.8 after PKC activation in DRGs is involved in the maintenance of chronic inflammatory pain in rats.