Detection of Sialylated N-Linked Glycans by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone（THAP）and 2,5-dihydroxybenzoic acid（DHB）as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer（MALDI-TOF MS）.High quality negative-ion spectra of commercial sialylated glycan were obtained with THAP as matrix.Detection limit of the glycan was less than 0.1 pmol.After methyl esterification of sialic acid（SA）residue,sialylated glycans were detected sensitively in the positive-ion mode using DHB as matrix.Neutral and sialylated glycans from the mixture of asialofetuin and fetuin were methylesterified and simultaneously recognized in one manipulation.Methyl esterification of SA residue offers a convenient and sensitive way to identify the structure of N-linked glycans for glycan profiling.

Dietary eubiotics of microbial muramidase and glycan improve intestinal villi,ileum microbiota composition and production trait of broiler

Background Optimal gut health is important to maximize growth performance and feed efficiency in broiler chickens.A total of 1,365 one-day-old male Ross 308 broiler chickens were randomly divided into 5 treatments groups with 21 replicates,13 birds per replicate.The present research investigated effects of microbial muramidase or a precision glycan alone or in combination on growth performance,apparent total tract digestibility,total blood carotenoid content,intestinal villus length,meat quality and gut microbiota in broiler chickens.Treatments included:NC:negative control(basal diet group);PC:positive control(basal diet+0.02%probiotics);MR:basal diet+0.035%microbial muramidase;PG:basal diet+0.1%precision glycan;and MRPG:basal diet+0.025%MR+0.1%PG,respectively.Results MRPG group increased the body weight gain and feed intake(P<0.05)compared with NC group.Moreover,it significantly increased total serum carotenoid(P<0.05)and MRPG altered the microbial diversity in ileum contents.The MRPG treatment group increased the abundance of the phylum Firmicutes,and family Lachnospiraceae,Ruminococcaceae,Oscillospiraceae,Lactobacillaceae,Peptostreptococcaceae and decreased the abundance of the phylum Campilobacterota,Bacteroidota and family Bacteroidaceae.Compared with the NC group,the chickens fed MRPG showed significantly increased in duodenum villus length at end the trial.Conclusion In this study,overall results showed that the synergetic effects of MR and PG showed enhancing growth performance,total serum carotenoid level and altering gut microbiota composition of broilers.The current research indicates that co-supplementation of MR and PG in broiler diets enhances intestinal health,consequently leading to an increased broiler production.

Comparing different domains of analysis for the characterisation of N-glycans on monoclonal antibodies

With the size of the biopharmaceutical market exponentially increasing,there is an aligned growth in the importance of data-rich analyses,not only to assess drug product safety but also to assist drug development driven by the deeper understanding of structure/function relationships.In monoclonal antibodies,many functions are regulated by N-glycans present in the constant region of the heavy chains and their mechanisms of action are not completely known.The importance of their function focuses analytical research efforts on the development of robust,accurate and fast methods to support drug development and quality control.Released N-glycan analysis is considered as the gold standard for glycosylation characterisation;however,it is not the only method for quantitative analysis of glycoform heterogeneity.In this study,ten different analytical workflows for N-glycan analysis were compared using four monoclonal antibodies.While observing good comparability between the quantitative results generated,it was possible to appreciate the advantages and disadvantages of each technique and to summarise all the observations to guide the choice of the most appropriate analytical workflow according to application and the desired depth of data generated.

N-linked glycoproteomic profiling in esophageal squamous cell carcinoma

BACKGROUND Mass spectrometry-based proteomics and glycomics reveal post-translational modifications providing significant biological insights beyond the scope of genomic sequencing.AIM To characterize the N-linked glycoproteomic profile in esophageal squamous cell carcinoma(ESCC)via two complementary approaches.METHODS Using tandem multilectin affinity chromatography for enrichment of N-linked glycoproteins,we performed N-linked glycoproteomic profiling in ESCC tissues by two-dimensional gel electrophoresis(2-DE)-based and isobaric tags for relative and absolute quantification(iTRAQ)labeling-based mass spectrometry quantitation in parallel,followed by validation of candidate glycoprotein biomarkers by Western blot.RESULTS 2-DE-based and iTRAQ labeling-based quantitation identified 24 and 402 differentially expressed N-linked glycoproteins,respectively,with 15 in common,demonstrating the outperformance of iTRAQ labeling-based quantitation over 2-DE and complementarity of these two approaches.Proteomaps showed the distinct compositions of functional categories between proteins and glycoproteins with differential expression associated with ESCC.Western blot analysis validated the up-regulation of total procathepsin D and high-mannose procathepsin D,and the downregulation of total haptoglobin,high-mannose clusterin,and GlcNAc/sialic acid-containing fraction of 14-3-3ζ in ESCC tissues.The serum levels of glycosylated fractions of clusterin,prolinearginine-rich end leucine-rich repeat protein,and haptoglobin in patients with ESCC were remarkably higher than those in healthy controls.CONCLUSION Our study provides insights into the aberrant N-linked glycoproteome associated with ESCC,which will be a valuable resource for future investigations.

Recent progress in targeting the sialylated glycan-SIGLEC axis in cancer immunotherapy

Malignant tumors are complex structures composed of cancer cells and tumor microenvironmental cells.In this complex structure,cells cross-talk and interact,thus jointly promoting cancer development and metastasis.Recently,immunoregulatory molecule-based cancer immunotherapy has greatly improved treatment efficacy for solid cancers,thus enabling some patients to achieve persistent responses or cure.However,owing to the development of drug-resistance and the low response rate,immunotherapy against the available targets PD-1/PD-L1 or CTLA-4 has limited benefits.Although combination therapies have been proposed to enhance the response rate,severe adverse effects are observed.Thus,alternative immune checkpoints must be identified.The SIGLECs are a family of immunoregulatory receptors(known as glyco-immune checkpoints)discovered in recent years.This review systematically describes the molecular characteristics of the SIGLECs,and discusses recent progress in areas including synthetic ligands,monoclonal antibody inhibitors,and Chimeric antigen receptor T(CAR-T)cells,with a focus on available strategies for blocking the sialylated glycan-SIGLEC axis.Targeting glyco-immune checkpoints can expand the scope of immune checkpoints and provide multiple options for new drug development.

Antitumor Active Protein-containing Glycans from the Body of Ganoderma tsugae

To explore the effects of traditional herbal medicine Ganoderrna tsugae（G, tsugae） on immunomodulatory and antitumor activities, the crude polysaccharides of G. tsugae were purified by filtration, diethylaminoethyl（DEAE） sepharose-fast flow chromatography and sephadex G-100 size-exclusion chromatography. Two main fractions, pro- tein-containing glyeans CSSLP-1 and CSSLP-2, were obtained via the gradient elution. The protein content, molecu- lar weight, and monosaccharide composition of the two fractions were analyzed. Furthermore, the influence of the protein-containing glycans from G. tsugae on the activation of human acute monocytic leukemia cell line（THP-l） and their antitumor activities to the human hepatocellular liver carcinoma cell（HepG-2） in vitro were evaluated. The re- sults indicate that CSSLP-1 and CSSLP-2 could increase the pinocytie activity of THP-1 cells and induce THP-1 cells to produce the eytokines of TNFa and IL-2, significantly. CSSLP-1 and CSSLP-2 also played an inhibiting effect on the cancer cell（HepG-2）. Moreover, the anti-proliferation activity of CSSLP-1 and CSSLP-2 increased with the par- ticipation of TNFa and IL-2 or other antitumor factors induced from THP-1 cells by G. tsugae protein-containing glycan fractions.

Partial characterization of the N-linked oligosaccharide structures on P-selectin glycoprotein ligand-1 (PSGL-1)

PSGL-1, a specific ligand for P-, E- and L-selectin, was isolated from in vivo [3H]-glucosamine labeled HL- 60 cells by a combination of wheat germ agglutinin-agarose and P- or E-selectin-agarose chromatography. N- linked oligosaccharides were released from the purified, denatured ligand molecule by peptide: N-glycosidase F treatment and, following separation by Sephacryl S-200 chromatography, partially characterized using lectin, ion-exchange and size-exclusion chromatography in combination with glycosidase digestions. The data obtained suggest that the N-glycans on PSGL-1 are predominantly core-fucosylated, multiatennary complex type structures with extended, poly- N- acetyllactosamine contaniing outer chains. A portion of the outer chains appears to be substituted with fucose indicating that the N-glycans, in addition to the O-glycans on PSGL-1, may be involved in selectin binding.

N-glycans released from glycoproteins using a commercial kit and comprehensively analyzed with a hypothetical database

The glycosylation of proteins is responsible for their structural and functional roles in many cellular activities.This work describes a strategy that combines an efficient release, labeling and liquid chromatography–mass spectral analysis with the use of a comprehensive database to analyze N-glycans. The analytical method described relies on a recently commercialized kit in which quick deglycosylation is followed by rapid labeling and cleanup of labeled glycans. This greatly improves the separation, mass spectrometry（MS） analysis and fluorescence detection of N-glycans. A hypothetical database, constructed using Glyc Resoft, provides all compositional possibilities of N-glycans based on the common sugar residues found in N-glycans. In the initial version this database contains ＆gt; 8,700 N-glycans, and is compatible with MS instrument software and expandable. N-glycans from four different well-studied glycoproteins were analyzed by this strategy. The results provided much more accurate and comprehensive data than that had been previously reported. This strategy was then used to analyze the N-glycans present on the membrane glycoproteins of gastric carcinoma cells with different degrees of differentiation. Accurate and comprehensive N-glycan data from those cells was obtained efficiently and their differences were compared corresponding to their differentiation states. Thus, the novel strategy developed greatly improves accuracy, efficiency and comprehensiveness of N-glycan analysis.

HBGH患者血清CGN、SDC-1水平与病情及疾病转归的关系

目的 探讨血清扣带蛋白(CGN)、多配体聚糖-1(SDC-1)水平与高血压性基底节脑出血(HBGH)患者病情及疾病转归的关系。方法 选取2019年2月至2022年2月连云港市第二人民医院(下称该院)收治的123例HBGH患者为研究对象,另选择该院同期行健康体检的体检健康者120例为健康组。检测两组纳入对象血清CGN、SDC-1表达水平。根据疾病转归情况将患者分为好转组92例,恶化组31例。采用受试者工作特征(ROC)曲线及曲线下面积(AUC)分析血清CGN、SDC-1表达水平对HBGH患者疾病转归的预测价值。结果 重度组血清CGN、SDC-1表达水平均高于中度组、轻度组,且中度组血清CGN、SDC-1表达水平均高于轻度组,差异有统计学意义(P<0.05),3组HBGH患者血清CGN、SDC-1表达水平均高于健康组,差异有统计学意义(P<0.05)。恶化组血清CGN、SDC-1表达水平高于好转组,差异有统计学意义(P<0.05)。血清CGN、SDC-1预测HBGH患者疾病转归的AUC分别为0.742(95%CI:0.792~0.697)、0.861(95%CI:0.906~0.910),两者联合预测的AUC为0.917(95%CI:0.962~0.870)。恶化组出血量、破入脑室情况明显高于好转组,入院格拉斯哥昏迷量表(GCS)评分低于好转组,差异有统计学意义(P<0.05)。多因素Logistic回归分析显示,血清CGN≥51.63 pg/mL(OR=3.815)、血清SDC-1≥450.67μg/L(OR=4.230)、入院GCS评分≤8分(OR=5.333)是HBGH患者疾病转归的影响因素(P<0.05)。结论 血清CGN、SDC-1表达水平升高与HBGH患者病情加重、疾病转归恶化密切相关,二者对HBGH患者疾病转归具有一定预测价值。

超声血流参数联合NLR、CA125、PLR对子宫内膜癌鉴别诊断价值

目的:探讨超声联合中性粒细胞/淋巴细胞比值(NLR)、血小板/淋巴细胞比值(PLR)NLR、糖类抗原125(CA125)对子宫内膜癌鉴别诊断价值。方法:收集2020年1月-2022年10月本院收治的子宫内膜癌患者92例为内膜癌组,子宫内膜良性病变患者82例为良性病变组,超声检测两组患者血流搏动指数(PI)、阻力指数(RI)、收缩期峰值流速(PSV),检测患者血清NLR、PLR、CA125水平,采用受试者工作特征(AUC)曲线分析各指标对子宫内膜癌鉴别诊断价值。结果:内膜癌组超声血流参数PSV高于对照组,PI、RI低于良性病变组,且内膜癌组Ⅲ-Ⅳ期患者PSV高于Ⅰ-Ⅱ期患者,PI、RI低于Ⅰ-Ⅱ期患者;血清CA125、NLR、PLR水平,内膜癌组均高于良性病变组,且内膜癌组Ⅲ-Ⅳ期患者高于Ⅰ-Ⅱ期患者(均P<0.05);ROC曲线分析显示,超声血流参数、CA125、NLR、PLR及联合应用鉴别诊断子宫内膜癌的曲线下面积分别为0.813、0.892,0.717、0.632、0.952,联合诊断效能最佳,敏感度98.2%。结论:超声血流参数,血清CA125、NLR、PLR在子宫内膜癌患者中发生异常,各指标联合应用对子宫内膜癌的鉴别诊断效力较高,有较好的临床指导价值。

色谱在糖组学分析中的应用

糖组学是继基因组学和蛋白组学后发展起来的组学技术,是研究细胞、组织或生物体内糖组的组成、结构及功能的一门学科。糖组学研究对深入了解生命活动规律、疾病的预防和治疗以及药物的质控和研发具有重要意义。糖组在生物样品中的丰度较低,且受单糖组成、糖苷键连接位置、连接方式、分支结构等因素的影响,使得糖链的组成与结构复杂多样,给糖组学研究带来了巨大挑战。液相色谱分离技术以及液相色谱-质谱联用技术被广泛应用于糖组的结构解析,在糖组学研究中发挥了不可或缺的作用。本文首先从色谱分离原理的角度讨论了糖链的富集方法及其在糖组学研究中的应用案例,并比较分析了各种方法的优势与不足;在糖链的分离与分析方面,从色谱分离模式的角度,分别列举了反相色谱、高效阴离子色谱、亲水相互作用色谱和多孔石墨化碳色谱分离糖链、糖缀合物或糖衍生物的分离原理,以及各类色谱模式联合质谱解析在糖组学分析中的研究进展。本文对各类色谱方法的分离特色进行了总结与讨论,为研究人员针对特定样品或特定目标物的分离与分析选择合适的方法提供了参考。近年来,在色谱技术的推动下,糖组学研究取得了相当大的进展。随着色谱新材料和新方法的不断开发,色谱技术将在糖组学研究中发挥更加重要的作用。

鹅颈藤壶多糖提取、理化性质及N-糖链表达谱分析研究

本文以南海海域鹅颈藤壶(Lepas anatifera)为原料,提取鹅颈藤壶背甲与肌肉质柄中的多糖,基于其基本理化性质的分析,比较不同生长时期鹅颈藤壶N-糖链表达谱的差异性。经过脱脂、碱水解/蛋白酶解、乙醇沉淀方法提取得到粗多糖;利用高效液相色谱法、高效凝胶渗透色谱法、苯酚-硫酸法、考马斯亮蓝法对其单糖组成、分子量分布、总糖含量及蛋白质含量进行分析;采用液质分析(PGC-ESI-MS/MS)方法对鹅颈藤壶成体(A-GB)与幼体(L-GB)肌肉质柄中N-糖链进行分析并比较。表明肌肉质柄中多糖分子量为18.40 kDa;总糖含量为11.99%,蛋白质含量60.14%,富含甘露糖、葡萄糖、氨基半乳糖,其中甘露糖的含量高达80%以上。在鹅颈藤壶成体与幼体中均鉴定得到26条N-糖链,成体中高甘露糖型N-糖链占总糖链的20.81%,且相对丰度较高的糖型组成为F_(1)H_(3)N_(2)(39.06%)、F_(1)H_(3)N_(3)(24.44%)、H_(3)N_(3)(9.35%);幼体中高甘露糖型N-糖链占总糖链的52.17%,相对丰度较高的糖型组成为F_(1)H_(3)N_(2)(21.65%)、H_(6)N_(2)(15.93%)、H_(5)N_(2)(14.75%)。鹅颈藤壶多糖是一类N-糖基化的蛋白复合物,推测N-糖链在鹅颈藤壶长发育过程中发挥着至关重要的作用,可能高甘露糖型N-糖链与其附着黏附能力密切相关。

源于细菌Dyella caseinilytica的新型N-糖酰胺酶的特性及其功能分析

N-糖酰胺酶(N-glycosidase,PNGase)是一种糖苷水解酶,主要功能为释放糖肽或糖蛋白上的N-糖链,在食品、生物医药等领域中具有重要的应用价值。目前,商业化的PNGase在使用过程中具有一定的局限性,因此,开发性能优良的新型PNGase具有研究意义。该研究在细菌Dyella caseinilytica中利用编码PNGase的基因,通过在大肠杆菌中进行重组表达,成功获得一种新型PNGase,命名为PNGase Dc,该酶分子质量为63.01 kDa。之后对其酶学特性、功能和应用潜力进行分析。酶学特性结果表明,PNGase Dc的最适pH和温度分别为2.0和37℃,酶促反应无金属离子依赖性;此外,该酶稳定性优良,在4℃下保存45 d仍能维持最高活性;分子对接结果表明,PNGase Dc与底物N′N-二乙酰壳二糖的相互作用力以氢键和范德华力为主;定点突变证明,Asp102和Glu236是影响重组PNGase Dc活性的关键氨基酸;PNGase Dc对标准糖蛋白辣根过氧化物酶和牛乳铁蛋白均具有良好的去糖基化效果;最后,以拟南芥和小鼠血浆作为底物评估PNGase Dc应用潜力,证实PNGase Dc在动植物源糖蛋白的研究中具有一定的应用潜力。综上所述,该研究为动植物源糖蛋白的研究提供了一个新的工具酶PNGase Dc,并为该酶的开发利用奠定理论基础。

半乳糖上调内皮细胞Siglec-9聚糖配体水平抑制巨噬细胞活性

目的探究半乳糖对人原代肺动脉内皮细胞上唾液酸特异性Ig样凝集素-9(sialic-acid-binding immunoglobulin-like lectin 9,Siglec-9)聚糖配体表达的影响及作用机制。方法采用流式细胞术确定内皮细胞表面Siglec-9聚糖配体的表达并鉴定该聚糖配体末端糖苷键类型;分别用左旋葡萄糖、葡萄糖、N-乙酰基葡萄糖、甘露糖、N-乙酰基甘露糖、半乳糖、唾液酸、蔗糖处理内皮细胞48 h,采用流式细胞术检测内皮细胞Siglec-9聚糖配体的表达;将内皮细胞分为对照组和半乳糖组(n=3),Western blot、流式细胞术、免疫荧光分析半乳糖对内皮细胞Siglec-9聚糖配体表达的影响;α2-3,6,8唾液酸酶处理内皮细胞后,Western blot检测半乳糖对内皮细胞Siglec-9聚糖配体恢复时间的影响;半乳糖处理内皮细胞后,Western blot检测分析细胞内唾液酸合成关键酶(GNE、NANS、NANP、CMAS、NPL以及ST3Gals)表达水平,并用RT-qPCR验证相关蛋白的mRNA表达;siRNA敲低NANP、CMAS及ST3Gal-Ⅲ,Western blot检测内皮细胞Siglec-9聚糖配体水平变化;巨噬细胞与内皮细胞共培养实验分析Siglec-9聚糖配体的增加对巨噬细胞的凋亡与吞噬能力的影响。结果内皮细胞上表达有Siglec-9聚糖配体且该配体为末端结构为α2-3连接的唾液酸糖蛋白;与对照组比较,半乳糖组内皮细胞上该聚糖配体表达水平显著增加(P<0.01),但半乳糖的添加对该配体的自我恢复时间无影响;与对照组比较,半乳糖处理的内皮细胞中NANP、CMAS、ST3Gal-Ⅲ表达水平升高(P<0.05,P<0.01),且RT-qPCR验证结果与其一致;抑制内皮细胞中NANP、CMAS、ST3Gal-Ⅲ的表达后,内皮细胞Siglec-9聚糖配体水平下降(P<0.01)。共培养实验中,与未处理组比较,半乳糖处理的内皮细胞促进巨噬细胞凋亡增加(P<0.01),并导致其吞噬能力减弱(P<0.05)。结论半乳糖通过NANP-CMAS-ST3Gal-Ⅲ通路上调内皮细胞上Siglec-9聚糖配体水平,从而抑制巨噬细胞活性。

合成生物学在多糖结合疫苗研发中的应用

由于合成生物学的快速发展,目前已经实现了人工设计DNA和蛋白质的合成,对具有重要生物学功能、结构也更为复杂的糖类物质进行精准设计合成,也将是合成生物学未来发展的重要方向。近年来,一种基于细菌寡糖转移酶体系的蛋白多糖偶联技术发展迅速,已被广泛应用于病原细菌多糖结合疫苗的生物合成制备。本文综述了该技术体系中的寡糖转移酶元件、载体蛋白元件、异源多糖抗原合成线路以及工程菌株改造等核心关键模块的最新研究进展。使用生物活体系统,发酵生产多糖结合疫苗,具有产物均一性好、步骤简便、绿色环保等优势,是一种亟待发展的新兴技术,同时也存在一些技术细节需要完善。未来,寡糖转移酶的定向进化、纳米颗粒型蛋白载体的应用、多糖合成基因的组合重排、工程菌株的代谢途径优化,将有望进一步促进多糖结合疫苗的生物合成研究。

N-乙酰氨基葡萄糖苷内切酶的功能与机制

N-乙酰氨基葡萄糖苷内切酶(endo-beta-N-acetylglucosaminidase,ENGase)广泛分布于各种生物中,主要通过降解错误折叠的糖蛋白,参与细胞和生命的调控。ENGase也是糖链编辑的有效工具酶,可专一性水解游离寡糖链及糖肽或糖蛋白上核心五糖的N-乙酰氨基葡萄糖(GlcNAc)之间的β-14糖苷键。其水解产物是寡糖链和一个GlcNAc,或带有一个GlcNAc的糖肽或糖蛋白。本文对ENGase的发现、分布、蛋白质结构、酶学反应及生物学功能进行阐述,为ENGase的生物学研究提供思路,为糖生物学与糖组学的应用研究奠定基础。

N-糖基化修饰的神经细胞生物学及其在神经系统疾病中的作用

神经发育和神经系统功能受到包括环境、遗传和表观遗传在内的多种因素的调控。N-糖基化修饰是由糖基化转移酶催化形成的蛋白质翻译后修饰,参与多种生物学过程。在神经系统中,N-糖基化修饰高丰度存在,调控神经突触的发育、成熟和胶质细胞的炎症反应。异常N-糖基化修饰与阿尔茨海默病、先天性糖基化障碍、精神分裂症和癫痫等多种神经系统疾病密切相关。本文中我们综述了N-糖基化修饰的神经细胞生物学功能及其在神经系统疾病中的作用和机制。

卵巢上皮性癌中inhibin及其受体β-glycan的表达

目的探讨α-inhibin及其受体β-glycan在恶性卵巢上皮性肿瘤中的表达,评估两种多肽信号对卵巢癌生长的调控作用,探讨卵巢癌的发病机制。方法应用EliVision方法检测30例正常卵巢组织、30例卵巢囊腺瘤和80例卵巢囊腺癌中α-in-hibin及其受体β-glycan的表达。结果α-inhibin在正常卵巢、卵巢囊腺瘤和卵巢囊腺癌中的总阳性表达率分别为93.3%、60.0%、36.3%,差异均有显著性(P<0.01);β-glycan在正常卵巢、卵巢囊腺瘤、卵巢囊腺癌的阳性表达率呈逐渐下降的趋势分别为100%、80.0%、75.0%,卵巢囊腺瘤、囊腺癌与正常卵巢组比较,差异有显著性(P<0.05),但卵巢囊腺瘤与囊腺癌比较,差异无显著性(P>0.05)。结论α-inhibin蛋白在卵巢囊腺癌中表达明显下降,而β-glycan在卵巢肿瘤中的表达仍然很高,提示β-glycan对增强α-inhibin抑制卵巢囊腺癌的形成可能起着一定的作用。

Serum N-glycan markers for diagnosing liver fibrosis induced by hepatitis B virus

BACKGROUND Hepatitis B virus (HBV) infection is the primary cause of hepatitis with chronic HBV infection,which may develop into liver fibrosis,cirrhosis and hepatocellular carcinoma.Detection of early-stage fibrosis related to HBV infection is of great clinical significance to block the progression of liver lesion.Direct liver biopsy is regarded as the gold standard to detect and assess fibrosis;however,this method is invasive and prone to clinical sampling error.In order to address these issues,we attempted to find more convenient and effective serum markers for detecting HBV-induced early-stage liver fibrosis.AIM To investigate serum N-glycan profiling related to HBV-induced liver fibrosis and verify multiparameter diagnostic models related to serum N-glycan changes.METHODS N-glycan profiles from the sera of 432 HBV-infected patients with liver fibrosis were analyzed.Significant changed N-glycan levels (peaks)(P <0.05) in differentfibrosis stages were selected in the modeling group,and multiparameter diagnostic models were established based on changed N-glycan levels by logistic regression analysis.The receiver operating characteristic (ROC) curve analysis was performed to evaluate diagnostic efficacy of N-glycans models.These models were then compared with the aspartate aminotransferase to platelet ratio index (APRI),fibrosis index based on the four factors (FIB-4),glutamyltranspeptidase platelet albumin index (S index),GlycoCirrho-test,and GlycoFibro-test.Furthermore,we combined multiparameter diagnostic models with alanine aminotransferase (ALT) and platelet (PLT) tests and compared their diagnostic power.In addition,the diagnostic accuracy of N-glycan models was also verified in the validation group of patients.RESULTS Multiparameter diagnostic models constructed based on N-glycan peak 1,3,4and 8 could distinguish between different stages of liver fibrosis.The area under ROC curves (AUROCs) of Model A and Model B were 0.890 and 0.752,respectively differentiating fibrosis F0-F1 from F2-F4,and F0-F2 from F3-F4,and surpassing other serum panels.However,AUROC (0.747) in Model C used for the diagnosis of F4 from F0-F3 was lower than AUROC (0.795) in FIB-4.In combination with ALT and PLT,the multiparameter models showed better diagnostic power (AUROC=0.912,0.829,0.885,respectively) when compared with other models.In the validation group,the AUROCs of the three combined models (0.929,0.858,and 0.867,respectively) were still satisfactory.We also applied the combined models to distinguish adjacent fibrosis stages of 432patients (F0-F1/F2/F3/F4),and the AUROCs were 0.917,0.720 and 0.785.CONCLUSION Multiparameter models based on serum N-glycans are effective supplementary markers to distinguish between adjacent fibrosis stages of patients caused by HBV,especially in combination with ALT and PLT.

CEA、CA19-9和基质金属蛋白酶9对于早期结直肠癌诊断价值的评价

目的 通过检测血清中CEA、CA19-9,以及尿液中MMP9(matrix metalloproteinases 9,MMP9)的水平,评价其对于早期结直肠癌(colorectal cancer, CRC)的诊断价值,旨在建立一种早期CRC的辅助诊断方法。方法 采集141例健康对照者、118例结肠息肉患者以及202例CRC患者的血清及尿液样本。二元Logistic回归建立3项指标的联合诊断模型,受试者工作特征曲线(receiver operating characteristic curve, ROC)和曲线下面积(area under curve, AUC)评价3项指标的单独和联合诊断价值。结果 与健康对照组和结肠息肉组相比,CRC组的CEA、CA19-9和MMP9水平均显著升高(P<0.05)。当CEA、CA19-9和MMP9联合建立诊断模型用于区分141例健康对照组与202例CRC患者时,其AUC为0.891(95%置信区间为0.858和0.924),灵敏度和特异性分别为88.12%和70.21%,对于早期CRC和晚期CRC的灵敏度和特异性分别为79.41%和70.21%,92.54%和70.21%。联合诊断模型用于区分118例结肠息肉患者与202例CRC患者时,其AUC为0.815(95%置信区间为0.768和0.861),其灵敏度和特异性分别为78.22%和67.80%,对于早期CRC的AUC为0.819(95%置信区间为0.773和0.863),灵敏度和特异性分别为82.35%和67.80%,晚期CRC的AUC为0.811(95%置信区间为0.760和0.862),其灵敏度和特异性分别为76.27%和67.80%。结论 与常用指标CEA和CA19-9相比,本研究所建立的联合诊断模型诊断性能有显著提升,可作为潜在的CRC辅助诊断方法。