Two intestinal microbiota-derived metabolites, deoxycholic acid and butyrate, synergize to enhance host defense peptide synthesis and alleviate necrotic enteritis

Background Necrotic enteritis(NE)is a major enteric disease in poultry,yet effective mitigation strategies remain elusive.Deoxycholic acid(DCA)and butyrate,two major metabolites derived from the intestinal microbiota,have independently been shown to induce host defense peptide(HDP)synthesis.However,the potential synergy between these two compounds remains unexplored.Methods To investigate the possible synergistic effect between DCA and butyrate in regulating HDP synthesis and barrier function,we treated chicken HD11 macrophage cells and jejunal explants with DCA and sodium butyrate(NaB),either individually or in combination,for 24 h.Subsequently,we performed RNA isolation and reverse transcrip-tion-quantitative PCR to analyze HDP genes as well as the major genes associated with barrier function.To further determine the synergy between DCA and NaB in enhancing NE resistance,we conducted two independent trials with Cobb broiler chicks.In each trial,the diet was supplemented with DCA or NaB on the day-of-hatch,followed by NE induction through sequential challenges with Eimeria maxima and Clostridium perfringens on d 10 and 14,respectively.We recorded animal mortality after infection and assessed intestinal lesions on d 17.The impact of DCA and NaB on the microbiota in the ileum and cecum was evaluated through bacterial 16S rRNA gene sequencing.Results We found that the combination of DCA and NaB synergistically induced multiple HDP genes in both chicken HD11 cells and jejunal explants.Additionally,the gene for claudin-1,a major tight junction protein,also exhibited synergistic induction in response to DCA and NaB.Furthermore,dietary supplementation with a combination of 0.75 g/kg DCA and 1 g/kg NaB led to a significant improvement in animal survival and a reduction in intestinal lesions compared to either compound alone in a chicken model of NE.Notably,the cecal microbiota of NE-infected chickens showed a marked decrease in SCFA-producing bacteria such as Bacteroides,Faecalibacterium,and Cuneatibacter,with lactobacilli becoming the most dominant species.However,supplementation with DCA and NaB largely restored the intestinal microbiota to healthy levels.Conclusions DCA synergizes with NaB to induce HDP and claudin-1 expression and enhance NE resistance,with potential for further development as cost-effective antibiotic alternatives.

The dual antimicrobial and immunomodulatory roles of host defense peptides and their applications in animal production

Host defense peptides(HDPs)are small molecules with broad-spectrum antimicrobial activities against infectious bacteria,viruses,and fungi.Increasing evidence suggests that HDPs can also indirectly protect hosts by modulating their immune responses.Due to these dual roles,HDPs have been considered one of the most promising antibiotic substitutes to improve growth performance,intestinal health,and immunity in farm animals.This review describes the antimicrobial and immunomodulatory roles of host defense peptides and their recent applications in animal production.

Discovery of natural products capable of inducing porcine host defense peptide gene expression using cell-based high throughput screening

Background:In-feed antibiotics are being phased out in livestock production worldwide.Alternatives to antibiotics are urgently needed to maintain animal health and production performance.Host defense peptides(HDPs)are known for their broad-spectrum antimicrobial and immunomodulatory capabilities.Enhancing the synthesis of endogenous HDPs represents a promising antibiotic alternative strategy to disease control and prevention.Methods:To identify natural products with an ability to stimulate the synthesis of endogenous HDPs,we performed a high-throughput screening of 1261 natural products using a newly-established stable luciferase reporter cell line known as IPEC-J2/pBD3-luc.The ability of the hit compounds to induce HDP genes in porcine IPEC-J2 intestinal epithelial cells,3D4/31 macrophages,and jejunal explants were verified using RT-qPCR.Augmentation of the antibacterial activity of porcine 3D4/31 macrophages against a Gram-negative bacterium(enterotoxigenic E.coli)and a Gram-positive bacterium(Staphylococcus aureus)were further confirmed with four selected HDP-inducing compounds.Results:A total of 48 natural products with a minimum Z-score of 2.0 were identified after high-throughput screening,with 21 compounds giving at least 2-fold increase in luciferase activity in a follow-up dose-response experiment.Xanthohumol and deoxyshikonin were further found to be the most potent in inducing pBD3 mRNA expression,showing a minimum 10-fold increase in IPEC-J2,3D4/31 cells,and jejunal explants.Other compounds such as isorhapontigenin and calycosin also enhanced pBD3 mRNA expression by at least 10-fold in both IPEC-J2 cells and jejunal explants,but not 3D4/31 cells.In addition to pBD3,other porcine HDP genes such as pBD2,PG1-5,and pEP2C were induced to different magnitudes by xanthohumol,deoxyshikonin,isorhapontigenin,and calycosin,although clear gene-and cell type-specific patterns of regulation were observed.Desirably,these four compounds had a minimum effect on the expression of several representative inflammatory cytokine genes.Furthermore,when used at HDP-inducing concentrations,these compounds showed no obvious direct antibacterial activity,but significantly augmented the antibacterial activity of 3D4/31 macrophages(P<0.05)against both Gram-negative and Gram-positive bacteria.Conclusions:Our results indicate that these newly-identified natural HDP-inducing compounds have the potential to be developed as novel alternatives to antibiotics for prophylactic and therapeutic treatment of infectious diseases in livestock production.

Geographically Distinct Expression Profile of Host Defense Peptides in the Skin of the Chinese Odorous Frog, Odorrana margaretae

Odorrana margaretae （Anura： Ranidae） is widely distributed in the southern provinces of China. Previously, 72 antimicrobial peptides （AMPs） belonging to 21 families were identified from the skin of O. margaretae, which were captured in the Hunan province. In the present study, five O. margaretae frogs were captured from the Guizhou province and a total of 28 cDNAs encoding 17 host defense peptides （HDPs） belonging to 14 families were cloned from the skin cDNA library of O. margaretae. Among the 17 HDPs, only one （brevinin-1-Omar5） had been characterized. The distinct HDP expression profiles for O. margaretae in the previous and present study may be attributed to the environmental differences between the sampling locations and the genetic divergence among O. margaretae populations. Besides, 11 of the 17 HDPs identified in the present study were novel for ranids. In order to understand their roles in host defense reactions, three HDPs （odorranain-H-OM1, odorranain-M-OM and ranatuerin-2-OM）, which possess low sequence similarity with the known amphibian HDPs, were selected for further chemical synthesis and functional analysis. Odorranain-H-OM1 showed direct antimicrobial activity against bacteria and fungi. Odorranain-M-OM exhibited concentration-dependent anti-oxidant activity. Ranatuerin-2-OM showed lectin-like activity and could strongly hemagglu -tinate human intact erythrocytes with or without the presence of Ca2＋. The diverse activities of HDPs implied that they may play different roles in host defense reactions of O. margaretae.

Host defense peptide-mimicking peptide polymer-based antibacterial hydrogel enables efficient healing of MRSA-infected wounds

Wound infections are a compelling health issue caused by the invasion and proliferation of pathogens in wound sites.Antibioticloaded hydrogels are widely used to achieve anti-infectious wound healing.However,due to the quick emergence of drugresistant bacteria,such as methicillin-resistant Staphylococcus aureus(MRSA),wound infection has been a formidable challenge to human health.To address MRSA-infected wounds,an antibacterial peptide polymer-loaded hyaluronic acid(HA)hydrogel(Gel-HA@P)is prepared.The peptide polymer is designed to mimic host defense peptides as the antibiotic alternative showing potent antibacterial activity,low susceptibility to drug resistance and good stability against proteolysis.HA is biocompatible and biodegradable hydrogel substrate as a primary constituent of the extracellular matrix and suitable for cell migration and wound healing.Gel-HA@P shows potent activity against MRSA in vitro and in vivo,low toxicity during the treatment and promotes the wound healing in vivo.This design has proven to be an effective and antibiotic-free strategy to enable the healing of MRSA-infected wounds.

Host defense peptide-mimickingβ-peptide polymer displaying strong antibacterial activity against cariogenic Streptococcus mutans

Cariogenic Streptococcus mutans(S.mutans)is a leading cause of bacterial-induced oral diseases.Current strategies to kill bacteria based on Host defense peptide(HDP)mimicking polymers hold promise to treat oral bacterial infection.Here,we explore the impact of hydrophobic subunit and chain length variation on the antibacterial and antibiofilm activity ofβ-peptide polymers.The physicochemical and biological prop-erties,such as the toxicity,the antibacterial activity,and the effect on bacterial transcription ofβ-peptide polymers,were systematically investigated with numerous techniques.The results exhibited that the op-timalβ-peptide polymer has low toxicity towards human periodontal ligament fibroblasts,andβ-peptide polymers(especially P3)have more excellent antibacterial activity against S.mutans than metronidazole.In addition,β-peptide polymers inhibited the reversible and irreversible bacterial adhesion during the formation of biofilms.The polymer can promote biofilm dispersion by decreasing the hydrophobicity of bacterial cells after adhering to cell surfaces.Analysis of the transcriptome for S.mutans treated withβ-peptide polymers demonstrated thatβ-peptide polymers could reduce the cariogenicity of S.mutans by impacting the transcription of the energy and acid metabolism-related genes.β-peptide polymers are promising antimicrobial agents in clinical dentistry due to their high antibacterial efficiency and low tox-icity.

Tissue expression and developmental regulation of chicken cathelicidin antimicrobial peptides

Cathelicidins are a major family of antimicrobial peptides present in vertebrate animals with potent microbicidal and immunomodulatory activities. Four cathelicidins, namely fowlicidins 1 to 3 and cathelicidin B1, have been identified in chickens. As a first step to understand their role in early innate host defense of chickens, we examined the tissue and developmental expression patterns of all four cathelicidins. Real-time PCR revealed an abundant expression of four cathelicidins throughout the gastrointestinal, respiratory, and urogenital tracts as well as in all primary and secondary immune organs of chickens. Fowlicidins 1 to 3 exhibited a similar tissue expression pattern with the highest expression in the bone marrow and lung, while cathelicidin B1 was synthesized most abundantly in the bursa of Fabricius. Additionally, a tissue-specific regulatory pattern was evident for all four cathelicidins during the first 28 days after hatching. The expression of fowlicidins 1 to 3 showed an age-dependent increase both in the cecal tonsil and lung, whereas all four cathelicidins were peaked in the bursa on day 4 after hatching, with a gradual decline by day 28. An abrupt augmentation in the expression of fowlicidins 1 to 3 was also observed in the cecum on day 28, while the highest expression of cathelicidin B1 was seen in both the lung and cecal tonsil on day 14. Collectively, the presence of cathelicidins in a broad range of tissues and their largely enhanced expression during development are suggestive of their potential important role in early host defense and disease resistance of chickens.

Dietary modulation of endogenous host defense peptide synthesis as an alternative approach to in-feed antibiotics

Traditionally, antibiotics are included in animal feed at subtherapeutic levels for growth promotion and disease prevention.However, recent links between in-feed antibiotics and a rise in antibiotic-resistant pathogens have led to a ban of all antibiotics in livestock production by the European Union in January 2006 and a removal of medically important antibiotics in animal feeds in the United States in January 2017.An urgent need arises for antibiotic alternatives capable of maintaining animal health and productivity without triggering antimicrobial resistance.Host defense peptides(HDP) are a critical component of the animal innate immune system with direct antimicrobial and immunomodulatory activities.While in-feed supplementation of recombinant or synthetic HDP appears to be effective in maintaining animal performance and alleviating clinical symptoms in the context of disease, dietary modulation of the synthesis of endogenous host defense peptides has emerged as a cost-effective,antibiotic-alternative approach to disease control and prevention.Several different classes of smallmolecule compounds have been found capable of promoting HDP synthesis.Among the most efficacious compounds are butyrate and vitamin D.Moreover, butyrate and vitamin D synergize with each other in enhancing HDP synthesis.This review will focus on the regulation of HDP synthesis by butyrate and vitamin D in humans, chickens, pigs, and cattle and argue for potential application of HDP-inducing compounds in antibiotic-free livestock production.

Breaking or following the membrane-targeting mechanism:Exploring the antibacterial mechanism of host defense peptide mimicking poly(2-oxazoline)s

Peptides exert important biological functions but their application is hindered by their susceptibility to proteolysis and poor stability in vivo.Thus,functional peptide mimics have drawn a great deal of attention to address this challenge.Poly(2-oxazoline)s,a class of biocompatible and proteolysis-resistant polymer,can work as host defense peptide mimics without following the general membrane-targeting mechanism as shown in our previous work.This observation encouraged us to figure out if poly(2-oxazoline)s are special and break the general membrane-targeting mechanism of host defense peptides and their mimics.In this study,we aimed at the connection between structure and antibacterial mechanism of poly(2-oxazoline)s.A new γ-aminobutyric acid(GABA)-pendent poly(2-oxazoline)was synthesized and investigated to compare with glycine-pendent poly(2-oxazoline)in our previous study,with the former polymer has two extra CH2 groups in the sidechain to increase the hydrophobicity and amphiphilicity.Membrane depolarization assay suggested that incorporating two more CH2 groups into the sidechain of poly(2-oxazoline)resulted in a mechanism switch from DNA-targeting to membrane-targeting,which was supported by the slow time-kill kinetics and slightly distorted and sunken membrane morphology.Besides,GABA-pendent poly(2-oxazoline)showed potent activity against methicillin-resistant S.aureus and low hemolysis on human red blood cells.Moreover,repeated use of the antimicrobial poly(2-oxazoline)did not stimulate bacteria to obtain resistance,which was an obvious advantage of membrane-targeting antimicrobial agents.

宿主防御肽在免疫调节中的作用与机制

宿主防御肽(HDPs)具有广泛的抗微生物及免疫调节活性,其最初作为一种直接的抗菌药物被广泛研究,而它们对于免疫系统的调节功能也随着研究的深入而逐步被认知。HDPs通过对先天免疫和适应性免疫的调控,实现了对病原微生物的清除,以维持机体稳态。目前,HDPs作为一种重要的免疫调节药物,在人类疾病治疗及畜牧生产中的应用开展了大量试验,但仍面临着许多挑战。本文总结了HDPs在先天免疫、适应性免疫和炎症调节中的功能和机制,并结合它们的应用和目前存在的问题阐述了HDPs的研究进展。

抗炎宿主防御肽治疗脓毒症机制研究进展

脓毒症是一种感染性疾病。近年来脓毒症已成为重症监护病房患者病死的主要原因,且其发病率和致死率呈现逐年上升趋势。尽管对于脓毒症的治疗有了一定的进展,如应用器官功能技术与抗感染治疗,但会产生如治疗周期长、治疗效果差且易产生耐药等问题,没有实质性地改善脓毒症患者的治疗效果,因此急需开发治疗效果好且不易产生耐药问题的新型药物。抗炎宿主防御肽(抗炎肽)是一类具有抗炎活性的多肽,因其来源广泛便于合成、毒副作用低且不易产生耐药问题而受到广泛的关注。本文对抗炎肽的理化性质和治疗脓毒症的作用机制进行综述,以期为治疗脓毒症的药物开发提供新思路。

宿主防御肽调节动物肠道屏障功能的研究进展

肠道屏障功能主要是通过调节黏蛋白和紧密连接(TJ)蛋白的合成来实现的,这对维持动物肠道健康和生产性能至关重要。作为先天性免疫系统的重要组成部分,宿主防御肽(HDPs)在黏膜防御中发挥着重要作用。肠道中的HDPs缺乏与屏障功能障碍和稳态失调有关。HDPs通过直接诱导多种黏蛋白和TJ蛋白的表达来增强肠道屏障功能。HDPs的诱导化合物有改善动物肠道形态、生产性能和饲料转化率的功能。本文将从HDPs对黏蛋白和TJ蛋白的转录调控作用、肠道屏障的分子调控机制以及动物生产性能的影响等方面进行综合阐述。

宿主防御肽的生物学功能、表达特性及调控机制

宿主防御肽是机体先天性免疫防御系统的重要组成成分,具有抗菌、抗病毒、抗寄生虫、免疫调节等多种生物学功能。宿主防御肽在动物机体的表达量可受多种因素调控,因此,透彻研究宿主防御肽的表达特性,对通过诱导宿主防御肽表达增强机体免疫力、抵御病原微生物感染具有重要意义。本文综述了宿主防御肽的概念、分类、生物学功能、表达特性及调控机制,为通过饲粮调控宿主防御肽表达以替代饲用抗生素应用于畜牧生产提供重要理论基础。

β-防御素在感染和炎症过程中的作用

防御素(defensins)在整个进化过程中构成了动物体内最大的一组宿主防御肽,这些富含半胱氨酸的阳离子肽具有广谱的抗微生物活性,能有效杀灭细菌、酵母和病毒等微生物。因此,防御素是生物体内先天性免疫的一个主要组成部分。防御素在免疫调节中的功能已被广泛研究,文章综述了β-防御素的免疫调节活性及β-防御素在感染和炎症过程中作用的最新进展。

牙龈上皮细胞的抗菌防御作用

牙龈上皮细胞不仅具有屏障作用,还具有先天免疫抗菌功能,如牙龈上皮细胞表面受体的信号传递、分泌抗菌多肽、蛋白酶和细胞因子等。下面就Toll样受体、抗菌多肽或蛋白、细胞因子或趋化因子、蛋白酶等与牙龈上皮细胞关系的研究进展作一综述,旨在加强对牙龈上皮细胞防御机制的认识。

抗菌肽的生物学功能及应用前景的研究进展(英文)

抗微生物肽(简称抗菌肽)是天然免疫系统的重要组成成分。它是一类分子量小、两亲性、带正电、进化上保守的小分子肽类物质。细菌、真菌、植物、无脊椎动物、脊椎动物以及人类等几乎所有生物中均可找到其的存在。在多细胞生物中,抗菌肽通过发挥一系列膜杀伤作用展示出了广谱抗微生物效应,从而扮演着一个重要的机体防御角色。这些防御效应包括抗细菌效应、抗真菌效应、抗寄生虫效应、抗病毒效应以及抗肿瘤效应。除了这些防御效应以外,抗菌肽也参与了机体的一系列生理和病理生理过程,包括免疫调节、伤口愈合、生殖过程以及决定犬类动物毛色等。近年来,随着微生物耐药现象越来越普遍,人们面临着日益严重的挑战,而对抗菌肽各种生物学功能的深入阐释,给我们带来了一种新的控制感染的有效途径。抗菌肽的这些生物学功能以及其潜在应用前景将在这篇综述中予以讨论。

营养物质对动物内源性宿主防御肽表达的调节作用

宿主防御肽是动物体产生的一种多功能肽,不仅具有广谱的抗菌功能,还可作为先天性免疫调节因子,有效增强动物机体免疫力,阻止病原微生物的侵扰,对保证动物健康具有重要意义。宿主防御肽在动物体内的表达受到日粮中各种营养物质的调控,有研究者认为,以营养调控的方式促进宿主内源防御肽的表达有益于动物健康和生产性能的提高,是未来实现无抗、健康养殖的可能策略之一。本文综合论述了动物饲粮中多种营养物质对宿主内源防御肽表达的调控作用,为评价营养物质调节宿主免疫和对动物健康及生产的影响提供新的理论参考。

宿主防御肽在奶牛乳腺中的表达及应用

宿主防御肽是生物体内产生的一类具有抗菌作用的阳离子多肽,是生物体非特异性免疫功能的重要组成部分。宿主防御肽具有广谱、抗菌、耐热、无毒、不易产生耐药性等诸多特性,在畜牧生产中具有广阔的应用前景。本文主要对宿主防御肽在先天免疫系统中的作用、奶牛乳腺中的表达及乳房炎诊断与治疗应用的研究进展进行综述,为宿主防御肽作为抗生素替代品在畜牧生产中的研究与应用提供理论支持。

4种中药单体与丁酸钠诱导猪宿主防御肽表达的协同效果研究

【目的】研究中药单体黄腐酚、异丹叶大黄素、去氧紫草素和毛蕊异黄酮与丁酸钠诱导猪宿主防御肽(host defense peptides,HDPs)表达的协同效果。【方法】中药单体黄腐酚、异丹叶大黄素、去氧紫草素和毛蕊异黄酮与丁酸钠单独或共同处理猪肠上皮细胞IPEC-J2和猪肺泡巨噬细胞3D4/31,通过测定HDPs的基因表达、细胞因子基因表达,以及细胞对产肠毒素性大肠杆菌(ETEC)的抑菌作用,评估中药单体与丁酸钠协同诱导HDPs表达的效果。【结果】黄腐酚与丁酸钠共同处理对HDPs的表达不表现协同作用;异丹叶大黄素与丁酸钠共同处理可协同诱导IPEC-J2细胞pEP2C和pBD3基因的表达(P<0.01);去氧紫草素与丁酸钠共同处理可协同提高IPEC-J2细胞内pBD3和PG1-5基因的表达(P<0.05;P<0.01);毛蕊异黄酮与丁酸钠共同处理可协同促进IPEC-J2细胞内PG1-5、pEP2C、pBD2和pBD3基因(P<0.05;P<0.01)和3D4/31细胞内pBD3基因的表达(P<0.05)。中药单体与丁酸钠共同处理对细胞因子表达无显著协作效果(P>0.05)。除黄腐酚外,其他中药单体与丁酸钠单独或共同处理后均可提高细胞裂解产物对ETEC的抑菌能力,其中毛蕊异黄酮与丁酸钠处理对ETEC的抑制具有最佳协同效果。【结论】异丹叶大黄素、去氧紫草素和毛蕊异黄酮与丁酸钠在诱导HDPs表达、提高机体免疫和抗菌能力方面具有一定的协同作用。

Biofilm Disruption Utilizing α/β Chimeric Polypeptide Molecular Brushes

Gram-negative bacteria can cause serious infections and are well known problems in biomedical practices. Biofilms of gramnegative bacteria are notorious for their frequently encountered resistance toward antibiotics. We demonstrate that α/β chimeric polypeptide molecular brush (α/β CPMB) exerts potent activities against antibiotic-resistant gram-negative bacteria. MTT viability assay, bacterial colony counting, and live/dead staining all indicate that α/β CPMB not only inhibits biofilm formation of gram-negative Pseudomonas aeruginosa and Acinetobacter baumannii, but also effectively disrupts mature biofllms that are highly resistant to one of the most active antibiotics-colistin. The superior antibacterial performance of the α/β CPMB implies its potential topical applications in treating biofilms.