Papain-like protease (PLpro), a crucial functional domain of the SARS-CoV-2 non-structural protein 3 (nsp3), plays a dual role in both hydrolyzing viral polyprotein precursors and modulating host immune responses. These critical functions position PLpro as a key target in the ongoing development of antiviral therapies for SARS-CoV-2. This review analyzes more than 100 PLpro-ligand co-crystal structures and summarizes the major binding modes between these ligands and PLpro. Most of these ligands bind to sites analogous to those targeted by the classical non-covalent inhibitor GRL0617, primarily involving the P3 and P4 subsites and the BL2 loop. Based on these structural insights, optimized inhibitors have expanded targeting beyond the canonical binding site to auxiliary regions such as the BL2 groove and the Val70 site, and in some cases toward the catalytic Cys111 buried within a narrow pocket. Certain ligands identified through various screening approaches bind to non-canonical or allosteric regions, such as the S1 and S2 sites or the zinc-finger domain, engaging PLpro through distinct interaction modes and thereby offering additional opportunities for PLpro inhibitor design. The review also discusses potential strategies for future PLpro inhibitor development informed by recent structural advances. Taken together, these structural and functional insights support ongoing efforts in the structure-guided design and optimization of PLpro inhibitors.

Lung adenocarcinoma (LUAD) is the most common histological subtype of lung cancer, characterized by a high incidence in late stages, high mortality rate, and poor prognosis. Src Homology 2 Domain Containing Protein 5 (SH2D5) is a mammalian-specific, uncharacterized scaffolding protein, and its role in LUAD remains unclear. In the present study, we investigated the function and potential mechanisms of SH2D5 in the progression of LUAD. We found aberrant expression of SH2D5 in LUAD tissues and cells, and its high expression is closely associated with poor prognosis in LUAD patients. Through loss-of-function and gain-of-function experiments, we revealed that overexpression of SH2D5 promotes the proliferation and migration abilities of lung adenocarcinoma cells. Gene set enrichment analysis (GSEA) revealed that SH2D5 positively regulates the epithelial-mesenchymal transition (EMT) process in lung adenocarcinoma cells. Additionally, we found that regulating the expression of SH2D5 influenced the phosphorylation levels of AKT, and the rescue experiments with AKT pathway activators/inhibitors partially reversed the tumor progression and EMT processes induced by SH2D5. In summary, our study demonstrated that SH2D5 promotes the migration and EMT process of LUAD cells through the AKT signaling pathway, suggesting that SH2D5 may serve as a crucial potential target for the treatment of metastatic LUAD.

Background Studying the mechanism of action of LncRNAs in lung adenocarcinoma (LUAD) is of great importance for an in-depth understanding of the molecular mechanism of lung adeno carcinogenesis and development. Objective The aim is to identify a long non-coding RNA LINC01117 that is specifically and highly expressed in LUAD cells and to investigate its biological functions and molecular mechanisms in LUAD cells, providing a new potential target for targeting LUAD therapy. Methods This study used publicly available data downloaded from The Cancer Genome Atlas (TCGA) database. Construction of siRNA and overexpression plasmid-packed lentiviral constructs were used to knock down and increase the expression of LINC01117 in LUAD cells. The effect of LINC01117 on LUAD cell migration and invasion was verified by scratch assays and Transwell assays. Western blot assays were performed to verify the effect of knocking down LINC01117 expression on key proteins of the EMT process. The effect of overexpression and knockdown LINC01117 expression on key proteins of the EMT process and the nuclear and cytoplasmic distribution of YAP1, a key effector molecule of the Hippo pathway, was verified by Western blot assays. Results LINC01117 expression was upregulated in LUAD tissues and cell lines. Clinical correlation and prognostic analyses showed that LINC01117 was associated with poorer clinical features (staging and N classification) and poorer prognosis and could be analyzed as an independent prognostic factor. Cell migration and invasion were significantly inhibited in the knockdown group compared to the control group; in contrast, cell migration and invasion were promoted in the overexpression group. Overexpression of LINC01117 resulted in down-regulation of E-cadherin expression and increased expression levels of N-cadherin, vimentin, ZEB1, snail and slug; in contrast, knockdown of LINC01117 appeared to have the opposite effect. Furthermore, knockdown of LINC01117 increased the enrichment of YAP1 protein in the cytoplasm and reduced its level in the nucleus; overexpression of LINC01117 produced the opposite intracellular distribution results. Conclusions LINC01117 was highly expressed in LUAD, and knockdown of LINC01117 significantly inhibited the migration and invasion of LUAD cells, while overexpression of LINC01117 significantly promoted the migration and invasion of LUAD cells, and affected the EMT process, and was able to alter the distribution of YAP1 in the nucleus and cytoplasm. This suggests that LINC01117 may regulate the activity of the Hippo pathway by altering the nuclear and cytoplasmic distribution of YAP1, which in turn induces the EMT process in lung adenocarcinoma cells and thus exerts a pro-cancer effect. It suggests that LINC01117 may play a key role in the occurrence and development of LUAD.

Lung cancer is one of the most lethal malignant tumors in the world. Non-small cell lung cancer (NSCLC) is the most common pathological subtype. However, the molecular mechanism of NSCLC progress is still unclear. We extracted the expression data of the Bruton's tyrosine kinase (BTK) gene in NSCLC tissues from the TCGA database. The results of paired t-test showed that the BTK gene was significantly underexpressed in NSCLC tissues. To further verify the above results, we detected the expression of the BTK gene in NSCLC cell lines A549, H1299, and H1650 at the RNA and protein levels by real-time fluorescent quantitative polymerase chain reaction and Western Blot analysis, respectively. The results showed that BTK was low expressed in NSCLC tissues and cells. More importantly, the expression of the BTK gene is also significantly related to the patient's age, gender, tumor range (T), lymph node invasion (N), tumor stage, and prognosis, and its expression level gradually decreases with the progress of the disease. It is speculated that BTK may be an independent prognostic factor of NSCLC. Our experimental results are consistent with the above clinical correlation analysis results. Overexpression of BTK can significantly inhibit the proliferation, migration, and invasion of NSCLC cells and can block the G0/G1 tumor cell cycle, indicating that overexpression of BTK can inhibit the growth, migration, and invasion of NSCLC cells.

Background Emerging evidence implicates the correlation of embryonic germline genes with the tumor progress and patient’s outcome. However, the prognostic value of these genes in lung adenocarcinoma (LUAD) has not been fully studied. Here we systematically evaluated this issue, and constructed a novel signature and a nomogram associated with embryonic germline genes for predicting the outcomes of lung adenocarcinoma. Methods The LUAD cohorts retrieved from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) database were used as training set and testing set, respectively. The embryonic germline genes were downloaded from the website https://venn.lodder.dev. Then, the differentially expressed embryonic germline genes (DEGGs) between the tumor and normal samples were identified by limma package. The functional enrichment and pathway analyses were also performed by clusterProfiler package. The prognostic model was constructed by the least absolute shrinkage and selection operator (LASSO)-Cox regression method. Survival and Receiver Operating Characteristic (ROC) analyses were performed to validate the model using training set and four testing GEO datasets. Finally, a prognostic nomogram based on the signature genes was constructed using multivariate regression method. Results Among the identified 269 DEGGs, 249 were up-regulated and 20 were down-regulated. GO and KEGG analyses revealed that these DEGGs were mainly enriched in the process of cell proliferation and DNA damage repair. Then, 103 DEGGs with prognostic value were identified by univariate Cox regression and further filtered by LASSO method. The resulting sixteen DEGGs were included in step multivariate Cox regression and an eleven embryonic germline gene related signature (EGRS) was constructed. The model could robustly stratify the LUAD patients into high-risk and low-risk groups in both training and testing sets, and low-risk patients had much better outcomes. The multi-ROC analysis also showed that the EGRS model had the best predictive efficacy compared with other common clinicopathological factors. The EGRS model also showed robust predictive ability in four independent external datasets, and the area under curve (AUC) was 0.726 (GSE30219), 0.764 (GSE50081), 0.657 (GSE37745) and 0.668 (GSE72094). More importantly, the expression level of some genes in EGRS has a significant correlation with the progression of LUAD clinicopathology, suggesting these genes might play an important role in the progression of LUAD. Finally, based on EGRS genes, we built and calibrated a nomogram for conveniently evaluating patients’ outcomes.

Immunotherapeutic strategies based on Epstein-Barr virus (EBV) latent membrane protein 2 (LMP2) antigen-specific cytotoxic T lymphocytes (CTLs) have been proven to boost LMP2-specific CTL responses in patients with nasopharyngeal carcinoma (NPC). Such strategies can produce clinical benefits in some patients with NPC. Currently, the major challenge limiting the use of immunotherapy for NPC is its low clinical response rate. The efficacy of immunotherapy based on EBV-LMP2 specific CTLs depends mainly on their cytotoxic activity, but no studies have been conducted to elucidate this activity. In this study, laser confocal scanning microscopy (LCSM) and real-time cell analysis (RTCA) were used to evaluate the killing function and its underlying mechanism of LMP2-specific CTLs. LCSM showed that LMP2-specific CTLs recognize and kill target cells expressing viral escape protein LMP2, and that the killing rate is related to the number of CTLs adhering to the target cells. LMP2-specific CTL-mediated cytotoxicity is rate limited by the time required for effective contact and recognition between CTLs and target cells. RTCA showed that the protective effect of LMP2-specific CTLs required an appropriate effector-to-target ratio, and that LMP2-specific CTLs could not eradicate residual target cells at a low effector-to-target ratio. Moreover, our results revealed that LMP2-specific CTL responses involve two independent but complementary mechanisms: the perforin/granzyme and Fas/FasL pathways. Therefore, we have elucidated, for the first time, the selective cytotoxicity and mechanism by which LMP2-specific CTLs induced by the rAd-LMP2 vaccine kill target cells and have explored the killing mode and several key parameters of killing mediated by LMP2-specific CTLs. Our study will contribute to the knowledge of vaccines targeting EBV-LMP2 and to the improvement of immunotherapeutic strategies.

目的:构建并鉴定表达HIV-1 CRF01_AE亚型结构基因的小鼠模型.方法:使用哺乳动物密码子优化的HIV-1 CRF01_AE gp160基因,通过慢病毒包装系统构建重组慢病毒LV-GFP-AE gp160,将上述重组慢病毒感染小鼠肺上皮细胞TC-1,经嘌呤霉素抗性筛选获得稳定表达gp160基因的TC-1细胞.采用RT-PCR、流式细胞术检测gp160基因在细胞内的表达稳定性,将稳定表达Gp160蛋白的TC-1-HIV AE gp160细胞接种小鼠,用免疫组化方法检测小鼠体内细胞团块中HIV Gp160蛋白的表达.结果:菌落PCR、酶切鉴定和测序表明重组质粒pLVX-AE gp160构建正确,RT-PCR、GFP荧光及流式细胞术结果均显示gp160基因能在细胞TC-1中稳定表达,免疫组化结果也表明小鼠体内接种的细胞可以稳定表达HIV Gp160蛋白.结论:建立了稳定表达HIV-1 CRF01_AE亚型Gp160蛋白的TC-1细胞及小鼠模型,为HIV-1 CRF01_AE亚型HIV疫苗的临床前研究提供了可靠的体外、体内免疫原性评价工具,为该疫苗的进一步开发奠定了坚实的实验基础.

生物制药是利用生命科学基础理论研发生产用于预防、诊断和治疗相关疾病生物制品的应用型学科,是当前国家重点发展的高新领域.为适应国家发展及市场的需求,更好地培养实用性生物制药人才,本文尝试从课程设置目的、课程内容优化以及教学方式等不同方面对生物制药创新课程设计的教学进行探讨,以期获得更好的教学效果.

目的:以非小细胞肺癌A549细胞为模型,探讨miR-490-3p在肺癌发生发展过程中的作用及其调控机制.方法:通过miRBase数据库获得miR-490-3p序列,设计miR-490-3p mimics并转染A549细胞,CCK8、细胞划痕及Transwell实验分别检测miR-490-3p过表达对A549细胞增殖、迁移和侵袭能力的影响;使用miRwalk在线工具预测miR-490-3p可能的调控基因,通过实时荧光定量PCR及Western印迹对候选调控基因进行筛选,最后通过双萤光素酶报告基因实验验证miR-490-3p与调控基因之间的靶向关系.结果:过表达miR-490-3p可显著抑制A549细胞的增殖、侵袭和迁移能力;在预测的miR-490-3p候选靶基因中,选择与细胞增殖、迁移等表型相关的RASAL2、TGFBR1、PAPPA、HMGA2、TGFA靶基因进行实时荧光定量PCR及Western印迹筛选,结果仅TGFBR1基因在mRNA和蛋白水平的表达与miR-490-3p水平呈负相关,且双萤光素酶报告实验证实miR-490-3p可直接与TGFBR1的3'-UTR结合并抑制其表达.结论:miR-490-3p通过靶向调控TGFBR1的表达抑制非小细胞肺癌A549细胞的增殖和侵袭.

Background Lung cancer has the highest morbidity and mortality worldwide, and lung adenocarcinoma (LADC) is the most common pathological subtype. Accumulating evidence suggests the tumor microenvironment (TME) is correlated with the tumor progress and the patient’s outcome. As the major components of TME, the tumor-infiltrated immune cells and stromal cells have attracted more and more attention. In this study, differentially expressed immune and stromal signature genes were used to construct a TME-related prognostic model for predicting the outcomes of LADC patients. Methods The expression profiles of LADC samples with clinical information were obtained from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO). The differentially expressed genes (DEGs) related to the TME of LADC were identified using TCGA dataset by Wilcoxon rank sum test. The prognostic effects of TME-related DEGs were analyzed using univariate Cox regression. Then, the least absolute shrinkage and selection operator (LASSO) regression was performed to reduce the overfit and the number of genes for further analysis. Next, the prognostic model was constructed by step multivariate Cox regression and risk score of each sample was calculated. Then, survival and Receiver Operating Characteristic (ROC) analyses were conducted to validate the model using TCGA and GEO datasets, respectively. The Kyoto Encyclopedia of Genes and Genomes analysis of gene signature was performed using Gene Set Enrichment Analysis (GSEA). Finally, the overall immune status, tumor purity and the expression profiles of HLA genes of high- and low-risk samples was further analyzed to reveal the potential mechanisms of prognostic effects of the model. Results A total of 93 TME-related DEGs were identified, of which 23 DEGs were up-regulated and 70 DEGs were down-regulated. The univariate cox analysis indicated that 23 DEGs has the prognostic effects, the hazard ratio ranged from 0.65 to 1.25 (p < 0.05). Then, seven genes were screened out from the 23 DEGs by LASSO regression method and were further analyzed by step multivariate Cox regression. Finally, a three-gene (ADAM12, Bruton Tyrosine Kinase (BTK), ERG) signature was constructed, and ADAM12, BTK can be used as independent prognostic factors. The three-gene signature well stratified the LADC patients in both training (TCGA) and testing (GEO) datasets as high-risk and low-risk groups, the 3-year area under curve (AUC) of ROC curves of three GEO sets were 0.718 (GSE3141), 0.646 (GSE30219) and 0.643 (GSE50081). The GSEA analysis indicated that highly expressed ADAM12, BTK, ERG mainly correlated with the activation of pathways involving in focal adhesion, immune regulation. The immune analysis indicated that the low-risk group has more immune activities and higher expression of HLA genes than that of the high-risk group. In sum, we identified and constructed a three TME-related DEGs signature, which could be used to predict the prognosis of LADC patients.

Background Lung adenocarcinoma (LUAD) is the major subtype of lung cancer and the most lethal malignant disease worldwide. However, the molecular mechanisms underlying LUAD are not fully understood. Methods Four datasets (GSE118370, GSE85841, GSE43458 and GSE32863) were obtained from the gene expression omnibus (GEO). Identification of differentially expressed genes (DEGs) and functional enrichment analysis were performed using the limma and clusterProfiler packages, respectively. A protein–protein interaction (PPI) network was constructed via Search Tool for the Retrieval of Interacting Genes (STRING) database, and the module analysis was performed by Cytoscape. Then, overall survival analysis was performed using the Kaplan–Meier curve, and prognostic candidate biomarkers were further analyzed using the Oncomine database. Results Totally, 349 DEGs were identified, including 275 downregulated and 74 upregulated genes which were significantly enriched in the biological process of extracellular structure organization, leukocyte migration and response to peptide. The mainly enriched pathways were complement and coagulation cascades, malaria and prion diseases. By extracting key modules from the PPI network, 11 hub genes were screened out. Survival analysis showed that except VSIG4, other hub genes may be involved in the development of LUAD, in which MYH10, METTL7A, FCER1G and TMOD1 have not been reported previously to correlated with LUAD. Briefly, novel hub genes identified in this study will help to deepen our understanding of the molecular mechanisms of LUAD carcinogenesis and progression, and to discover candidate targets for early detection and treatment of LUAD.

目的:探讨桦褐孔菌乙酸乙酯提取物对食管癌细胞EC-109、EC-9706增殖、凋亡和细胞周期等表型的影响.方法:用桦褐孔菌乙酸乙酯提取物处理食管癌细胞株EC-109、EC-9706,采用MTT法、流式细胞术、平板克隆实验分析桦褐孔菌乙酸乙酯提取物对食管癌细胞增殖、凋亡和细胞周期的影响.结果:桦褐孔菌乙酸乙酯提取物可显著抑制食管癌细胞的增殖,并降低肿瘤细胞的克隆形成能力.流式细胞术显示,桦褐孔菌提取物可将食管癌细胞周期阻滞在G1期,并促进肿瘤细胞的凋亡.结论:桦褐孔菌乙酸乙酯提取物可抑制人食管癌细胞株的增殖和克隆形成,阻滞细胞周期并诱导肿瘤细胞凋亡,具有一定的抗食管癌活性.

目的:探讨桦褐孔菌水提物对人类免疫缺陷病毒Ⅰ型(HIV-1)活性的抑制作用及抗病毒作用的潜在靶点.方法:通过细胞毒性、假病毒单轮感染实验、HIV-1 VSV-G包膜实验,评价桦褐孔菌水提物的抗HIV-1药效学;用体外逆转录酶、整合酶活性检测和时间递加分析实验,对桦褐孔菌水提物体外抗HIV-1活性的潜在作用靶点进行初步研究.结果:桦褐孔菌水提物对3株不同包膜的HIV-1均有抑制作用,IC50分别为12.17±2.085、5.661±1.058、8.846±1.334μg/mL.水提物浓度为0.65、12.5、25μg/mL时均能抑制HIV-1的逆转录酶活性,抑制率分别为13.31％±3.39％、26.24％±5.6％、28.36％±4.3％;25μg/mL的桦褐孔菌水提物对整合酶有显著抑制作用(P<0.05).时间递加分析显示,桦褐孔菌水提物在HIV-1感染的融合/进入阶段具有最高的病毒抑制率.结论:桦褐孔菌水提物具有抗HIV-1活性的作用,其对HIV-1的融合/进入、逆转录酶及整合酶均有显著抑制作用,提示桦褐孔菌水提物可能在HIV-1的融合、逆转录、整合阶段存在作用靶点.

质量风险管理是一种科学的管理方法,常用于药品生产和质量管理过程.药品共线生产是指药品在某些工艺步骤,与其他药品共用生产设施设备进行生产的方式.由于药品共线生产的特殊性,可能会对药品质量带来潜在风险,在本文中笔者对质量风险管理在药品共线生产时的应用进行了深入研究,对当前研究现状进行深入分析,以得出更加权威的结论.

系统生物学是以系统观点研究生命现象的交叉学科,代表着生物学未来研究的方向.然而,系统生物学涉及学科多,内容跨度大,学习曲线陡峭.针对系统生物学本科教学中存在的问题,本文尝试从教材选择、课程内容优化以及教学方式等不同方面探讨进行教学改革的策略,以期获得更好的教学效果.

桦褐孔菌是一种名贵的食药真菌,具有降糖、降血脂的功效,在我国北方及俄罗斯西伯利亚地区作为一种保健食品有着悠久的使用历史.近来的研究显示,桦褐孔菌中的某些活性成分还具有显著的抗肿瘤、抗病毒和寄生虫感染等药理作用,提示其具有较好的药用前景.本文拟对桦褐孔菌的活性成分及相关药理作用的最新进展进行综述.

病毒学实验技术是研究病毒及抗病毒药物的重要手段和工具.笔者结合多年教学经验,并在借鉴兄弟院校相关课程的经验基础上,在教学内容,教学形式及考核方式等方面对病毒学实验技术课程进行了改革,以期获得更好的教学效果.

近年来,食品安全问题受到全世界的共同关注,它关系到人民健康、社会稳定、国家声誉等方方面面.《食品安全学》是高等院校食品质量与安全专业的主干课程,由于其学科交叉性强、内容多、知识点分散等特点,因此存在授课难度大、教学质量难以达到预期效果等问题.作为一门新兴课程,其建设和发展必然要经历循序渐进的过程.辩论式教学是以学生为主体,由小组或全班成员围绕特定的论题辩驳问难,各抒己见,在辩论中主动获取知识、提高素养的一种教学方式.本文结合《食品安全学》课程的特色内容,探讨如何将辩论式教学引入《食品安全学》课程中,以期达到更好的教学效果,从而促进综合型专业人才的培养.

Objective To construct a recombinant double gene co-expressing plasmid of HIV-1 broadly neutralizing antibody and to detect its expression in 293T cell line.Methods HIV-1 neutralizing antibody 2G12 variable region of light chain (VL) and heavy chain (VH) was synthesized,and ligated with vectors containing human IgG constant regions of light and heavy chain to construct a complete 2G12 light and heavy Chain.The VL and VH of 2G12 and IRES were cloned into eukaryotic expression vector pVR by PCR amplification and then the recombinant plasmid was transfected into 293T cells.Expression of the antibody in cell supernatant was detected by ELISA.Binding and neutralizing activity of the cell supernatant were tested by ELISA and micro-neutralization assays.Results The recombinant double gene eukaryotic expression vector which can express human IgG was constructed successfully.The expression level of the supernatant was 6.43 μg/ml,and the antibody retained the binding and neutralizing activity.Conclusions The constructed vector can express the antibody with binding and neutralizing activity,this study provides a good platform for the expression of human HIV neutralizing antibody in the eukaryotic expression vector.

《食品安全学》是食品质量与安全专业主干必修课,涉及知识面广,涵盖学科领域多,其主要内容贯穿于食品生产、加工、储存等相关食品产业的全过程.针对工科院校学生的知识结构特点,在借鉴兄弟院校教学经验基础上对教学内容,教学方式方法及考核形式进行了改革,取得了良好的教学效果.