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International Journal of Ophthalmology(English edition) 2015年第5期8卷 866-871页

作者： Lin Cong Yi-Ping Xia Gui-Qiu Zhao Jing Lin Qiang Xu Li-Ting Hu Jian-Qiu Qu Xu-Dong Peng Department of Ophthalmology the Affiliated Hospital of Qingdao University

AIMTo observe the expression of vitamin D receptor (VDR) in human specimen and immortalized human corneal epithelium cells (HCEC) when challenged with fusarium solani. Moreover, we decided to discover the pathway of VDR expression. Also, we would like to detect the expression of cathelicidin antimicrobial peptide (CAMP) in the downstream pathway of VDR.METHODSImmunohistochemistry was used to examine the VDR expression in HCEC from healthy and fungal keratitis patients. Real time quantitative polymerase chain reaction (qPCR) was performed to observe the messenger ribonucleic acid (mRNA) change of VDR when immortalized HCEC were challenged with fusarium solani for different hours. CAMP was detected at both mRNA and protein levels.RESULTSWe found out that the VDR expression in fusarium solani keratitis patients' specimen was much more than that in healthy people. The mRNA and protein expression of VDR increased when we stimulated HCEC with fusarium solani antigen (P<0.01) and it could be inhibited by toll like receptor 2 (TLR2) monoclonal antibody. The CAMP expression was decreased because of fusarium solani antigen stimulation (P<0.01).CONCLUSIONThe VDR expression can be increased via TLR2/1-VDR pathway while the CAMP expression is decreased by the stimulation of fusarium solani antigen.

关键词： vitamin D receptor fusarium solani corneal epithelium cathelicidin

Bicarbonate Effects on Antibacterial Immunity and Mucus Glycobiology in the Cystic Fibrosis Lung:A Review With Selected Experimental Observations

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Infectious Microbes & Diseases 2022年第3期4卷 103-110页

作者： Ruth Siew Tzung-Lin Ou Samira Dahesh Kathryn Akong Victor Nizet Department of Pediatrics University of California San DiegoLa JollaCaliforniaUSA School of Medicine China Medical UniversityTaichungTaiwan Skaggs School of Pharmacy and Pharmaceutical Sciences University of California San DiegoLa JollaCaliforniaUSA

The primary defect in cystic fibrosis(CF)is abnormal chloride and bicarbonate transport in the CF transmembrane conductance regulator epithelial ion channel.The apical surface of the respiratory tract is lined by an airway surface liquid(ASL)layer composed of mucin comprising mainly MUC5A and MUC5B glycoproteins.ASL homeostasis depends on sodium bicarbonate secretion into the airways and secretion deficits alter mucus properties leading to airway obstruction,inflammation and infections.Downstream effects of abnormal ion transport in the lungs include altered intrinsic immune defenses.We observed that neutrophils killed Pseudomonas aeruginosa more efficiently when it had been exposed to sodium bicarbonate,and formation of neutrophil extracellular traps by neutrophils was augmented in the presence of increasing bicarbonate concentrations.Physiological levels of bicarbonate sensitized P.aeruginosa to the antimicrobial peptide cathelicidin LL-37,which is present in both lung ASL and neutrophil extracellular traps.Sodium bicarbonate has various uses in clinical medicine and in the care of CF patients and could be further explored as a therapeutic adjunct against Pseudomonas infections.

关键词： cystic fibrosis bicarbonate mucus pneumonia bacterial infection neutrophil cathelicidin neutrophil extracellular traps

cathelicidin在烟曲霉菌性角膜炎中的作用及机制的研究

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Cathelicidin在烟曲霉菌性角膜炎中的作用及机制的研究

作者：侯晓晨青岛大学

学位级别：硕士

目的:研究cathelicidin在烟曲霉菌角膜炎中的作用及对中性粒细胞吞噬、自噬等功能的影响。方法:1.建立小鼠烟曲霉菌角膜炎动物模型,采用RT-PCR、Western blot法检测感染后0、1、2、4天小鼠角膜中cathelicidin(CRAMP)m RNA和蛋白的表达;... 详细信息

目的:研究cathelicidin在烟曲霉菌角膜炎中的作用及对中性粒细胞吞噬、自噬等功能的影响。方法:1.建立小鼠烟曲霉菌角膜炎动物模型,采用RT-PCR、Western blot法检测感染后0、1、2、4天小鼠角膜中cathelicidin(CRAMP)m RNA和蛋白的表达;收集临床真菌性角膜炎患者角膜、角膜移植术后剩余的供体周边角膜,RT-PCR、Western blot和免疫荧光技术检测角膜中cathelicidin(LL-37)的表达和定位。2.使用外源性CRAMP或磷酸盐缓冲液(PBS)对小鼠角膜进行干预,利用裂隙灯照相、临床评分评估CRAMP的干预效果;RT-PCR和ELISA实验检测各组角膜的促炎症因子的表达、用苏木精-伊红(HE)染色评估角膜组织损伤程度、流式细胞技术分析中性粒细胞募集水平,以评估CRAMP在体内对炎症反应的调控;通过过碘酸-希夫染色(PAS)、菌落形成单位(CFU)计数实验评估CRAMP在小鼠角膜内的抗真菌疗效。3.在体外,使用CRAMP或PBS预处理小鼠中性粒细胞后与羧基荧光素琥珀酰氨基酯(CFSE)标记的烟曲霉菌分生孢子共培养,流式细胞术实验分析CRAMP在体外对中性粒细胞吞噬分生孢子能力的影响;将分生孢子注入小鼠腹腔,通过流式细胞技术分析CRAMP在体内对中性粒细胞吞噬作用的影响。4.在用LL-37或PBS处理人中性粒细胞前加入或不加入LL-37功能性受体CXCR2的拮抗剂SB225002或磷脂酶Cγ(PLCγ)抑制剂U73122,最后与CFSE标记的分生孢子共培养,通过流式细胞技术评估各组吞噬水平的变化,Western blot检测各组中性粒细胞PLCγ的磷酸化水平。5.CRAMP或PBS预处理的小鼠中性粒细胞与CFSE标记的分生孢子共培养,之后将中性粒细胞洗净并再培养一段时间,流式细胞实验分析CRAMP在体外对中性粒细胞清除胞内分生孢子能力的影响;体内实验中将CFSE标记的分生孢子注入小鼠腹腔并给予CRAMP或PBS腹腔注射,通过流式细胞技术分析CRAMP在体内对中性粒细胞胞内分生孢子清除力的影响。7.LL-37或PBS处理人中性粒细胞前用或不用SB225002或自噬抑制剂巴弗洛霉素A进行预处理,最后与CFSE标记的分生孢子共培养,通过流式细胞技术评估各组胞内真菌杀伤力的变化,并用RT-PCR、Western blot检测各组中性粒细胞自噬相关基因和蛋白的表达。结果:1.感染烟曲霉菌的人或小鼠角膜中LL-37或CRAMP的表达增加。2.CRAMP处理2天后,小鼠烟曲霉菌性角膜炎的病变程度开始减轻;CRAMP处理4天后,角膜的临床评分明显降低。与病变程度变化趋势相同的是,与PBS处理组相比,CRAMP降低了角膜组织中的促炎因子IL-6、TNF-a和IL-1β的表达和中性粒细胞浸润水平。PAS染色和CFU计数试验显示,与PBS处理组相比,CRAMP处理明显抑制了角膜中的真菌生长,并减少了真菌负荷。3.流式细胞技术显示,在体内、外,CRAMP都能上调中性粒细胞吞噬烟曲霉菌分生孢子的能力。4.LL-37通过其在中性粒细胞上的功能性受体CXCR2促进细胞内PLCγ的磷酸化活化增强中性粒细胞的吞噬功能。5.流式细胞技术显示,在体内、外,CRAMP都能增强中性粒细胞的胞内真菌清除力。7.LL-37/CXCR2上调中性粒细胞自噬相关蛋白(Beclin-1、LC3-II)的表达,以加速对胞内分生孢子的清除。结论:cathelicidin能抑制烟曲霉菌角膜炎的炎症反应并减少真菌负荷。cathelicidin能加速中性粒细胞吞噬和降解烟曲霉菌分生孢子。在机制上,LL37/CXCR2激活中性粒细胞PLCγ以增强吞噬作用,并促进中性粒细胞自噬以消除胞内的分生孢子。

关键词： cathelicidin 烟曲霉菌角膜炎抗炎中性粒细胞抗真菌活性

NMR assisted studies on the solution structures and functions of antimicrobial peptides Dedicated to Professor Chaohui Ye on the occasion of his 80th birthday

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Magnetic Resonance Letters 2022年第4期2卷 214-223页

作者： Yaying Zhang Chunyang Cao College of Chemistry and Materials Science Shanghai Normal University100 Guilin RoadShanghai200234China State Key Laboratory of Bioorganic and Natural Product Chemistry Center for Excellence in Molecular SynthesisShanghai Institute of Organic ChemistryChinese Academy of Sciences345 Lingling RoadShanghai200032China

Microbial resistance has now become a global public health concern,and the spread of multidrug-resistant bacteria also threatens human health.Antimicrobial peptides(AMPs)are a class of small peptides with antibacterial,anti-inflammatory,anti-infective,antioxidation,anti-tumor,antiviral functions and immune regulation activities.Due to the small sizes,their structures are easily studied by nuclear magnetic resonance(NMR)techniques.Compared to traditional antibiotics,AMPs have specific antibacterial mechanisms,and do not easily result in the production of drug-resistant strains.Thus,the development of new antimicrobial peptides and their wide use instead of chemical antibiotics are of great significance to human health.In this review,we first summarized the relationship between the structures and functions of antimicrobial peptides.Then,we focused on examples,cathelicidins,a group of cationic antimicrobial peptides with multiple biological activities.Especially,cathelicidin BF30 or BF34,composed of 30 or 34 amino acids,were from the venom glands of the Bungarus fasciatus snake and were considered to be the most active antibacterial peptides among different cathelicidin members.Their solution structures determined by NMR are a-helixes,which are useful in designing new and stable peptides with similar framework,including stapple peptides by inducing chemical modifications in the sidechains of some residues,as well as cyclic peptides by inducing disulfide bond between cysteines in the sequences.

关键词： Nuclear magnetic resonance Antimicrobial peptides cathelicidin Structure Function

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中华肿瘤杂志 2018年第3期40卷 172-177页

作者：权文强潘星昊吴军录姚懿雯李冬同济大学附属同济医院检验科上海200065 华侨大学生物医学学院厦门361021

目的研究肿瘤相关巨噬细胞分泌的抗菌肽cathelicidin促进小鼠结直肠癌生长的作用及机制。方法通过氧化偶氮甲烷/葡聚糖硫酸钠（AOM/DSS）方法建立小鼠肠炎相关的结直肠癌模型。对诱导成瘤的小鼠，同时每隔3 d注射1次小鼠cathelicidin中... 详细信息

目的研究肿瘤相关巨噬细胞分泌的抗菌肽cathelicidin促进小鼠结直肠癌生长的作用及机制。方法通过氧化偶氮甲烷/葡聚糖硫酸钠（AOM/DSS）方法建立小鼠肠炎相关的结直肠癌模型。对诱导成瘤的小鼠，同时每隔3 d注射1次小鼠cathelicidin中和抗体、IgG抗体（阳性对照）和PBS（阴性对照），持续1个月，取小鼠结肠，拍照、计数肿瘤数目并进行评价。利用实时荧光定量PCR和Western blot法检测小鼠结肠肿瘤组织、非肿瘤组织以及组织中巨噬细胞cathelicidin的表达；采用免疫组化法检测小鼠结肠肿瘤组织与非肿瘤组织中的cathelicidin、肿瘤增殖抗原Ki-67和CD68的表达；采用原位末端转移酶标记技术（TUNEL）检测小鼠结肠肿瘤组织中细胞的凋亡情况。结果cathelicidin在小鼠结肠肿瘤组织的炎症免疫细胞（巨噬细胞）中呈高表达状态，而在肿瘤细胞中表达较弱。注射cathelicidin中和抗体组小鼠的肿瘤数目和长径分别为（4.50±1.18）个和（1.74±0.18）mm，明显低于注射PBS组小鼠的肿瘤数目[（13.88±1.98）个，P〈0. 01]和肿瘤长径[（3.74±0.38）mm，P〈0. 01]；也明显低于注射IgG抗体小鼠的肿瘤数目[（15.25±1.82）个，P〈0. 01]和肿瘤长径[（3.40±0.36）mm，P〈0.01]。注射cathelicidin中和抗体组肿瘤组织中Ki-67阳性肿瘤细胞百分比为（28.20±3.44）%，低于注射PBS组[（68.20±3.51）%，P〈0.01]和注射IgG抗体组[（69.20±3.41）%，P〈0.01]。免疫组织化学染色检测结果显示，注射cathelicidin中和抗体组小鼠组织中CD68的表达明显低于注射IgG抗体组和注射PBS组。TUNEL染色显示，注射cathelicidin中和抗体对肿瘤细胞的凋亡影响不大。结论肿瘤相关巨噬细胞分泌的cathelicidin具有促进小鼠结直肠癌增殖的作用，中和cathelicidin的表达可以抑制结直肠癌的生长和增殖。cathelicidin通过募集炎症细胞（巨噬细胞）进入肿瘤微环境可能是其促进结直肠癌增殖的主要作用机制。

关键词：结直肠肿瘤 cathelicidin 肿瘤微环境炎症肿瘤相关巨噬细胞

cathelicidin a potential therapeutic peptide for gastrointestinal inflammation and cancer

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World Journal of Gastroenterology 2013年第18期19卷 2731-2735页

作者： Jimmy Yip Chuen Chow Zhi Jie Li William Ka Kei Chi Hin Cho Department of Medicine University of California San Diego La Jolla CA 92037 United States Western Australia Institute for Medicine Research the University of Western Australia Institute of Digestive Diseases the Chinese University of Hong Kong Hong Kong China School of Biomedical Sciences Faculty of Medicine the Chinese University of Hong Kong Hong Kong China

cathelicidins, are host defense peptides synthesized and stored in circulating leukocytes and numerous types of epithelial tissues in particular the gastrointestinal (GI) tract and skin. They have been known for their antimicrobial activities against a variety of microbes. Recently it was discovered that they have other significant biological functions and produce appealing pharmacological actions against inflammation and cancer in the GI tract through defined mechanisms. Experimental evidence shows that these actions could be tissue and disease specific and concentration dependent. This article reviews some of the physiological functions of cathelicidins and also their therapeutic potential in the treatment of inflammation and cancer and also the delivery system for this peptide as targeted therapy for various disorders in the GI tract both in animals and humans.

关键词： cathelicidin Gastrointestinal tract Cancer Inflammation Ulcer repair

中华大蟾蜍皮肤cathelicidin家族新型抗菌肽的鉴定及其抗菌活性

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中国中药杂志 2016年第4期41卷 630-635页

作者：展波高媛媛林维平贾广韬孙同毅李文辉潍坊医学院生物科学与技术学院山东潍坊261053 潍坊医学院临床医学院山东潍坊261053 中国科学院昆明动物研究所云南省动物模型与人类疾病机理重点实验室云南昆明650223

对中华大蟾蜍皮肤转录组进行了测定,通过对得到转录组数据的组装、注释和序列比对得到1个新型cathelicidin家族前体c DNA。按照cathelicidin家族抗菌肽的加工方式,推导出该前体可能编码2个成熟的中华大蟾蜍cathelicidin家族抗菌肽,命名... 详细信息

对中华大蟾蜍皮肤转录组进行了测定,通过对得到转录组数据的组装、注释和序列比对得到1个新型cathelicidin家族前体c DNA。按照cathelicidin家族抗菌肽的加工方式,推导出该前体可能编码2个成熟的中华大蟾蜍cathelicidin家族抗菌肽,命名为BG-CATH25,BG-CATH29。圆二色散测定了合成BG-CATH25,BG-CATH29的二级结构特征,结果揭示BG-CATH25,BG-CATH29在不同缓冲体系中以无规则卷曲的构象为主。对实验所用4种细菌的抗菌活性研究表明BG-CATH25,BGCATH29对嗜水气单胞菌有较强的抑菌活性,MIC分别为1.25,10 mg·L-1。

关键词：中华大蟾蜍 cathelicidin 抗菌肽转录组嗜水气单胞菌

一种新型改良硫氧还蛋白融合表达体系的建立及cathelicidin家族Lf-CATH2的表达

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生物工程学报 2015年第3期31卷 403-410页

作者：鲁一灵高久香乔雪王义鹏于海宁大连理工大学生命科学与技术学院辽宁大连116024 苏州大学药学院江苏苏州215123

通过改良硫氧还蛋白融合表达体系,原核表达cathelicidin家族抗菌肽Lf-CATH2。首先在Lf-CATH2基因上游加入凝血酶位点,并去除p ET32α载体的凝血酶序列和S标签序列,构建优化的Lf-CATH2-p ET32α-TS载体,于大肠杆菌中表达。产物融合蛋白... 详细信息

通过改良硫氧还蛋白融合表达体系,原核表达cathelicidin家族抗菌肽Lf-CATH2。首先在Lf-CATH2基因上游加入凝血酶位点,并去除p ET32α载体的凝血酶序列和S标签序列,构建优化的Lf-CATH2-p ET32α-TS载体,于大肠杆菌中表达。产物融合蛋白经凝血酶切割释放Lf-CATH2,纯化后进行抗菌活性检测。结果表明改良的硫氧还蛋白融合表达体系显著提高酶切效率达37%,Lf-CATH2在新体系中获得了可溶性高表达,且保留了抗菌活性。因此该新型硫氧还蛋白融合表达体系,有望为cathelicidin家族及其他阳离子活性肽提供更好的原核表达载体工具。

关键词： cathelicidin Lf-CATH2 硫氧还蛋白融合表达凝血酶切割

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cathelicidin的特性及其应用

免疫学杂志 2006年第3期22卷 345-347页

作者：李明胡福泉第三军医大学基础部微生物学教研室重庆市微生物工程重点实验室重庆400038

cathelicidin是在哺乳动物体内发现的一类结构多变的抗微生物肽,因在表达的信号肽与成熟肽之间含有一高度保守的cathelin肽段而自成一家族。cathelicidin和防御素一样,也是宿主免疫防御系统的重要组成部分。除了主要的抗菌活性以外,Cath... 详细信息

cathelicidin是在哺乳动物体内发现的一类结构多变的抗微生物肽,因在表达的信号肽与成熟肽之间含有一高度保守的cathelin肽段而自成一家族。cathelicidin和防御素一样,也是宿主免疫防御系统的重要组成部分。除了主要的抗菌活性以外,cathelicidin还具有抑制组织损伤,促进创伤修复,结合内毒素,诱导血管生成等多种生物学功能。在临床耐药菌问题日益严重的今天,进一步了解cathelicidin的生物学活性和临床作用,无疑将推动整个抗感染治疗事业的发展。

关键词： cathelicidin 抗生素生物学功能

Chitosan hydrogel encapsulated with LL-37 peptide promotes deep tissue injury healing in a mouse model

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Military Medical Research 2020年第4期7卷 409-417页

作者： Xu Yang Jing-Lin Guo Jing Han Rui-Juan Si Pan-Pan Liu Zi-Rui Zhang Ai-Min Wang Ju Zhang School of Nursing Qingdao UniversityQingdao 266021China

Background:LL-37 peptide is a member of the human cathelicidin family,and has been shown to promote the healing of pressure ulcers.However,the low stability of this peptide within the wound environment limits its clinical use.Chitosan(CS)hydrogel is commonly used as a base material for wound dressing material.Methods:CS hydrogel(2.5%w/v)was encapsulated with LL-37.Cytotoxicity of the product was examined in cultured NIH3 T3 fibroblasts.Effects on immune response was examined by measuring tumor necrosis factor-α(TNF-α)release from RAW 264.7 macrophages upon exposure to lipopolysaccharides.Antibacterial activity was assessed using Staphylococcus aureus.Potential effect on pressure ulcers was examined using a mouse model.Briefly,adult male C57 BL/6 mice were subjected to skin pressure using magnets under a 12/12 h schedule for 21 days.Mice were randomized to receive naked LL-37(20μg),chitosan gel containing 20μg LL-37(LL-37/CS hydrogel)or hydrogel alone under the ulcer bed(n=6).A group of mice receiving no intervention was also included as a control.Results:LL-37/CS hydrogel did not affect NIH3 T3 cell viability.At a concentration of 1–5μg/ml,LL-37/CS inhibited TNF-αrelease from macrophage.At 5μg/ml,LL-37/CS inhibited the growth of Staphylococcus aureus.The area of the pressure ulcers was significantly lower in mice receiving LL-37/CS hydrogel in comparison to all other 3 groups on days 11(84.24%±0.25%),13(56.22%±3.91%)and 15(48.12%±0.28%).Histological examination on days 15 and 21 showed increased epithelial thickness and density of newly-formed capillary with naked LL-37 and more so with LL-37/CS.The expression of key macromolecules in the process of angiogenesis(i.e.,hypoxia inducible factor-1α(HIF-1α)and vascular endothelial growth factor-A(VEGF-A))in wound tissue was increased at both the mRNA and protein levels.Conclusion:Chitosan hydrogel encapsulated with LL-37 is biocompatible and could promote the healing of pressure ulcers.

关键词： LL-37 Chitosan hydrogel Deep tissue injury Pressure ulcer cathelicidin