Email Alerts
RSS
Volume 37 Issue 11
Nov.  2021
Turn off MathJax
Article Contents
Article Navigation >  CHINESE JOURNAL OF BURNS AND WOUNDS  > 2021  >  37(11): 1024-1035
Zhang QR,Yang X,Li Z,et al.Effects of reactive oxygen species-responsive antibacterial microneedles on the full-thickness skin defect wounds with bacterial colonization in diabetic mice[J].Chin J Burns,2021,37(11):1024-1035.DOI: 10.3760/cma.j.cn501120-20210831-00299.
Citation: Zhang QR,Yang X,Li Z,et al.Effects of reactive oxygen species-responsive antibacterial microneedles on the full-thickness skin defect wounds with bacterial colonization in diabetic mice[J].Chin J Burns,2021,37(11):1024-1035.DOI: 10.3760/cma.j.cn501120-20210831-00299.
shu

Effects of reactive oxygen species-responsive antibacterial microneedles on the full-thickness skin defect wounds with bacterial colonization in diabetic mice

doi: 10.3760/cma.j.cn501120-20210831-00299
  • 1.

    Department of Burns and Plastic Surgery, Affiliated Hospital of Nantong University, Nantong 226001, China

  • 2.

    State Key Laboratory of Trauma, Burns and Combined Injury, Institute of Burn Research, the First Affiliated Hospital of Army Medical University (the Third Military Medical University), Chongqing Key Laboratory for Disease Proteomics, Chongqing 400038, China

Funds:

Natural Science Foundation of Chongqing Municipality of China cstc2021jcyj-msxmX0513

Nantong Science and Technology Project JC2021178, HS2020006

More Information
  • Corresponding author: Yu Yunlong, Email: yuyunlong666@gmail.com; Zhang Yi, Email: 198zy@163.com
  • Received Date: 2021-08-31
    Abstract
  •     Objective   To study the effects of reactive oxygen species (ROS)-responsive antibacterial microneedles (MNs) on the full-thickness skin defect wounds with bacterial colonization in diabetic mice.    Methods   Experimental research methods were adopted. The ROS-responsive crosslinker N1-(4-boronobenzyl)-N3-(4-boronophenyl)-N1, N1, N3, N3-tetramethylpropane-1,3-diaminium (TSPBA) was first synthesized, and then the polyvinyl alcohol (PVA)-TSPBA MNs, PVA-ε-polylysine (ε-PL)-TSPBA MNs, PVA-TSPBA-sodium hyaluronate (SH) MNs, and PVA-ε-PL-TSPBA-SH MNs were prepared by mixing corresponding ingredients, respectively. The PVA-TSPBA MNs were placed in pure phosphate buffer solution (PBS) and PBS containing hydrogen peroxide, respectively. The degradation of MNs immersed for 0 (immediately), 3, 7, and 10 days was observed to indicate their ROS responsiveness. The standard strains of Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) cultured in Luria-Bertani medium containing hydrogen peroxide were divided according to the random number table (the same grouping method below) into blank control group (without any treatment, the same below) and 0 g/L ε-PL group, 1.0 g/L ε-PL group, 5.0 g/L ε-PL group, and 10.0 g/L ε-PL group with which PVA-ε-PL-TSPBA MNs containing the corresponding concentration of ε-PL were co-cultured, respectively. Bacterial growth was observed after 24 h of culture, and the relative survival rate of bacteria was calculated (n=3). The mouse fibroblast cell line 3T3 cells at logarithmic growth stage (the same growth stage below) were divided into blank control group and 0 g/L ε-PL group, 1.0 g /L ε-PL group, 5.0 g /L ε-PL group, and 10.0 g /L ε-PL group in which cells were cultured in medium with the extract from PVA-ε-PL-TSPBA MNs containing the corresponding concentration of ε-PL, respectively. Cell growth was observed after 24 h of culture by optical microscopy, and the relative survival rate of cells was detected and calculated by cell counting kit 8 (CCK-8) assay to indicate the cytotoxicity (n=6). Both PVA-TSPBA MNs and PVA-TSPBA-SH MNs were taken, the morphology of the two kinds of MNs was observed by optical microscopy, and the mechanical properties of the two kinds of MNs were tested by microcomputer controlled electronic universal testing machine (denoted as critical force, n=6). Six male BALB/c mice aged 6-8 weeks (the same gender and age below) were divided into PVA-TSPBA group and PVA-TSPBA-SH group, with 3 mice in each group. After pressing the skin on the back of mice vertically with the corresponding MNs for 1 minute, the skin condition was observed at 0, 10, and 20 min after pressing. Another batch of 3T3 cells were divided into blank control group, 0 g/L ε-PL group and simple 5.0 g/L ε-PL group which were cultured with the extract of PVA-ε-PL-TSPBA MNs containing the corresponding concentration of ε-PL, and 5.0 g/L ε-PL+SH group which were cultured with the extract of PVA-ε-PL-TSPBA-SH MNs with 5.0 g/L ε-PL. The CCK-8 assay was performed to detect and calculate the relative survival rate of cells cultured for 24, 48, and 72 h to indicate the cell proliferation activity (n=6). Eighteen BALB/c mice were induced into diabetic mice model by high-sugar and high-fat diet combined with streptozotocin injection and then divided into sterile dressing group, 0 g/L ε-PL+SH group, and 5.0 g/L ε-PL+SH group, with 6 mice in each group. A full-thickness skin defect wound was made on the back of each mouse, and S. aureus solution was added to make a full-thickness skin defect wound with bacterial colonization model for diabetic mouse. The wounds of mice in 0 g/L ε-PL+SH group and 5.0 g/L ε-PL+SH group were covered with PVA-ε-PL-TSPBA-SH MNs with the corresponding concentration of ε-PL, and the wounds of mice in the 3 groups were all covered with sterile surgical dressings. The wound healing was observed on post injury day (PID) 0, 3, 7, and 12, and the wound healing rate on PID 3, 7, and 12 was calculated. On PID 12, the skin tissue of the wound and the wound margin were stained with hematoxylin and eosin to observe the growth of new epithelium and the infiltration of inflammatory cells. Data were statistically analyzed with one-way analysis of variance, analysis of variance for repeated measurement, Mann-Whitney U test, and Bonferroni test.    Results   With the extension of the immersion time, the PVA-TSPBA MNs in PBS containing hydrogen peroxide gradually dissolved and completely degraded after 10 days of immersion. The PVA-TSPBA MNs in pure PBS only swelled but did not dissolve. After 24 h of culture, there was no growth of S. aureus in 5.0 g/L ε-PL group or 10.0 g/L ε-PL group, and there was no growth of E. coli in 10.0 g/L ε-PL group. The relative survival rate of S. aureus was significantly lower in 1.0 g/L ε-PL group, 5.0 g/L ε-PL group, and 10.0 g/L ε-PL group than in blank control group (P<0.05 or P<0.01). The relative survival rate ofE. coli was significantly lower in 5.0 g/L ε-PL group and 10.0 g/L ε-PL group than in blank control group (P<0.01). After 24 h of culture, the cells in blank control group, 0 g/L ε-PL group, 1.0 g/L ε-PL group, 5.0 g/L ε-PL group, and 10.0 g/L ε-PL group all grew well, and the relative survival rate of cells was similar among the groups (P>0.05). The needle bodies of PVA-TSPBA MNs and PVA-TSPBA-SH MNs were both quadrangular pyramid-shaped and neatly arranged, and the needle bodies of PVA-TSPBA-SH MNs was more three-dimensional and more angular. The critical force of PVA-TSPBA-SH MNs was significantly higher than that of PVA-TSPBA MNs (Z=3.317, P<0.01). The MNs in PVA-TSPBA+SH group penetrated the skin of mice at 0 min after pressing, and the pinholes partially disappeared after 10 min and completely disappeared after 20 min, while the MNs in PVA-TSPBA group failed to penetrate the skin of mice. After 24, 48, and 72 h of culture, the proliferation activity of the cells in 5.0 g/L ε-PL+SH group was significantly higher than that of blank control group (P<0.05 or P<0.01). In sterile dressing group, the wounds of mice healed slowly and exuded more. The wound healing speed of mice in 0 g/L ε-PL+SH group was similar to that of sterile dressing group in the early stage but was faster than that of sterile dressing group in the later stage, with moderate exudation. The wound healing of mice in 5.0 g/L ε-PL+SH group was faster than that in the other two groups, with less exudation. The wound healing rates of mice in 5.0 g/L ε-PL+SH group were (40.6±4.2)%, (64.3±4.1)%, and (95.8±2.4)% on PID 3, 7, and 12, which were significantly higher than (20.4±2.7)%, (38.9±2.2)%, and (59.1±6.2)% in sterile dressing group and (21.6±2.6)%, (44.0±1.7)%, and (82.2±5.3)% in 0 g/L ε-PL+SH group (P<0.01). The wound healing rates of mice in 0 g/L ε-PL+SH group on PID 7 and 12 were significantly higher than those in sterile dressing group (P<0.05 or P<0.01). On PID 12, the wounds of mice in 5.0 g/L ε-PL+SH group were almost completely epithelialized with less inflammatory cell infiltration, the wounds of mice in 0 g/L ε-PL+SH group were partially epithelialized with a large number of inflammatory cell infiltration, and no obvious epithelialization but a large number of inflammatory cell infiltration was found in the wounds of mice in sterile dressing group.    Conclusions   The composite MNs prepared by TSPBA, PVA, ε-PL, and SH can successfully penetrate mouse skin and slowly respond to ROS in the wound to resolve and release antibacterial substances, inhibit bacterial colonization, and promote the repair of full-thickness skin defect wounds with bacterial colonization in diabetic mice.

     

    • FullText(HTML)
  • loading
    • References(40)
  • [1]
    WuYK,ChengNC,ChengCM.Biofilms in chronic wounds: pathogenesis and diagnosis[J].Trends Biotechnol,2019,37(5):505-517.DOI: 10.1016/j.tibtech.2018.10.011.
    [2]
    GriceEA,SnitkinES,YockeyLJ,et al.Longitudinal shift in diabetic wound microbiota correlates with prolonged skin defense response[J].Proc Natl Acad Sci U S A,2010,107(33):14799-14804.DOI: 10.1073/pnas.1004204107.
    [3]
    NishikawaT,EdelsteinD,DuXL,et al.Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage[J].Nature,2000,404(6779):787-790.DOI: 10.1038/35008121.
    [4]
    FerrucciL,FabbriE.Inflammageing: chronic inflammation in ageing, cardiovascular disease, and frailty[J].Nat Rev Cardiol,2018,15(9):505-522.DOI: 10.1038/s41569-018-0064-2.
    [5]
    OkinD,MedzhitovR.Evolution of inflammatory diseases[J].Curr Biol,2012,22(17):R733-740.DOI: 10.1016/j.cub.2012.07.029.
    [6]
    DuXL,MatsumuraT,EdelsteinD,et al.Inhibition of GAPDH activity by poly(ADP-ribose) polymerase activates three major pathways of hyperglycemic damage in endothelial cells[J].J Clin Invest,2003,112(7):1049-1057.DOI: 10.1172/JCI18127.
    [7]
    GerardC,RollinsBJ.Chemokines and disease[J].Nat Immunol,2001,2(2):108-115.DOI: 10.1038/84209.
    [8]
    FullertonJN,GilroyDW.Resolution of inflammation: a new therapeutic frontier[J].Nat Rev Drug Discov,2016,15(8):551-567.DOI: 10.1038/nrd.2016.39.
    [9]
    ChouhanD,DeyN,BhardwajN,et al.Emerging and innovative approaches for wound healing and skin regeneration: current status and advances[J].Biomaterials,2019,216:119267.DOI: 10.1016/j.biomaterials.2019.119267.
    [10]
    DuckworthPF,RowlandsRS,BarbourME,et al.A novel flow-system to establish experimental biofilms for modelling chronic wound infection and testing the efficacy of wound dressings[J].Microbiol Res,2018,215:141-147.DOI: 10.1016/j.micres.2018.07.009.
    [11]
    PrausnitzMR,GomaaY,LiW.Microneedle patch drug delivery in the gut[J].Nat Med,2019,25(10):1471-1472.DOI: 10.1038/s41591-019-0606-0.
    [12]
    LiW,TerryRN,TangJ,et al.Rapidly separable microneedle patch for the sustained release of a contraceptive[J].Nat Biomed Eng,2019,3(3):220-229.DOI: 10.1038/s41551-018-0337-4.
    [13]
    RamadonD,PermanaAD,CourtenayAJ,et al.Development, evaluation, and pharmacokinetic assessment of polymeric microarray patches for transdermal delivery of vancomycin hydrochloride[J].Mol Pharm,2020,17(9):3353-3368.DOI: 10.1021/acs.molpharmaceut.0c00431.
    [14]
    PermanaAD,MirM,UtomoE,et al.Bacterially sensitive nanoparticle-based dissolving microneedles of doxycycline for enhanced treatment of bacterial biofilm skin infection: a proof of concept study[J].Int J Pharm X,2020,2:100047.DOI: 10.1016/j.ijpx.2020.100047.
    [15]
    WangJQ,YeYQ,YuJR,et al. Core-shell microneedle gel for self-regulated insulin delivery[J].ACS Nano,2018,12(3):2466-2473.DOI: 10.1021/acsnano.7b08152.
    [16]
    ChenHH,ChengYH,TianJR,et al.Dissolved oxygen from microalgae-gel patch promotes chronic wound healing in diabetes[J].Sci Adv,2020,6(20):eaba4311.DOI: 10.1126/sciadv.aba4311.
    [17]
    WolcottRD,RumbaughKP,JamesG,et al.Biofilm maturity studies indicate sharp debridement opens a time-dependent therapeutic window[J].J Wound Care,2010,19(8):320-328.DOI: 10.12968/jowc.2010.19.8.77709.
    [18]
    FuXB. Wound healing center establishment and new technology application in improving the wound healing quality in China [J/OL]. Burns Trauma, 2020, 8: tkaa038[2021-08-31].https://academic.oup.com/burnstrauma/article/doi/ 10.1093/burnst/tkaa038/5939901.DOI: 10.1093/burnst/tkaa038.
    [19]
    MetcalfDG,BowlerPG.Clinical impact of an anti-biofilm Hydrofiber dressing in hard-to-heal wounds previously managed with traditional antimicrobial products and systemic antibiotics[J/OL].Burns Trauma,2020,8:tkaa004[2021-08-31].https://academic.oup.com/burnstrauma/article/doi/ 10.1093/burnst/tkaa004/5780075.DOI: 10.1093/burnst/tkaa004.
    [20]
    PermanaAD,McCruddenMTC,DonnellyRF.Enhanced intradermal delivery of nanosuspensions of antifilariasis drugs using dissolving microneedles: a proof of concept study [J].Pharmaceutics,2019,11(7):346.DOI: 10.3390/pharmaceutics11070346.
    [21]
    WaghuleT,SinghviG,DubeySK,et al.Microneedles: a smart approach and increasing potential for transdermal drug delivery system[J].Biomed Pharmacother,2019,109:1249-1258.DOI: 10.1016/j.biopha.2018.10.078.
    [22]
    AliR,MehtaP,ArshadMS,et al.Transdermal microneedles-a materials perspective[J].AAPS PharmSciTech,2019,21(1):12.DOI: 10.1208/s12249-019-1560-3.
    [23]
    Haj-AhmadR,KhanH,ArshadMS,et al.Microneedle coating techniques for transdermal drug delivery[J].Pharmaceutics,2015,7(4):486-502.DOI: 10.3390/pharmaceutics7040486.
    [24]
    XenikakisI,TzimtzimisM,TsongasK,et al.Fabrication and finite element analysis of stereolithographic 3D printed microneedles for transdermal delivery of model dyes across human skin in vitro[J].Eur J Pharm Sci,2019,137:104976.DOI: 10.1016/j.ejps.2019.104976.
    [25]
    RacV,LevićS,BalančB,et al.PVA cryogel as model hydrogel for iontophoretic transdermal drug delivery investigations. Comparison with PAA/PVA and PAA/PVP interpenetrating networks[J].Colloids Surf B Biointerfaces,2019,180:441-448.DOI: 10.1016/j.colsurfb.2019.05.017.
    [26]
    StrehlC,GaberT,MauriziL,et al.Effects of PVA coated nanoparticles on human immune cells[J].Int J Nanomedicine,2015,10:3429-3445.DOI: 10.2147/IJN.S75936.
    [27]
    NafeaEH,Poole-WarrenLA,MartensPJ.Bioactivity of permselective PVA hydrogels with mixed ECM analogues[J].J Biomed Mater Res A,2015,103(12):3727-3735.DOI: 10.1002/jbm.a.35510.
    [28]
    ZhaoH,HuangJ,LiY,et al.ROS-scavenging hydrogel to promote healing of bacteria infected diabetic wounds[J].Biomaterials,2020,258:120286.DOI: 10.1016/j.biomaterials.2020.120286.
    [29]
    HyldgaardM,MygindT,VadBS,et al.The antimicrobial mechanism of action of epsilon-poly-l-lysine[J].Appl Environ Microbiol,2014,80(24):7758-7770.DOI: 10.1128/AEM.02204-14.
    [30]
    SantiniMT,CamettiC,IndovinaPL,et al.Polylysine induces changes in membrane electrical properties of K562 cells[J].J Biomed Mater Res,1997,35(2):165-174.DOI: 10.1002/(sici)1097-4636(199705)35:2<165::aid-jbm4>3.0.co;2-l.
    [31]
    NiethammerP,GrabherC,LookAT,et al.A tissue-scale gradient of hydrogen peroxide mediates rapid wound detection in zebrafish[J].Nature,2009,459(7249):996-999.DOI: 10.1038/nature08119.
    [32]
    HecquetCM,AhmmedGU,VogelSM,et al.Role of TRPM2 channel in mediating H2O2-induced Ca2+ entry and endothelial hyperpermeability[J].Circ Res,2008,102(3):347-355.DOI: 10.1161/CIRCRESAHA.107.160176.
    [33]
    HuangLY,SzewczykG,SarnaT,et al.Potassium iodide potentiates broad-spectrum antimicrobial photodynamic inactivation using photofrin[J].ACS Infect Dis,2017,3(4):320-328.DOI: 10.1021/acsinfecdis.7b00004.
    [34]
    PeroglioM,GradS,MortisenD,et al.Injectable thermoreversible hyaluronan-based hydrogels for nucleus pulposus cell encapsulation[J].Eur Spine J,2012,21Suppl 6(Suppl 6):S839-849.DOI: 10.1007/s00586-011-1976-2.
    [35]
    Loureiro BorghettiR,de VargasKF,Pozatti MoureS,et al.Clinical and histologic evaluation of effects of hyaluronic acid in rat tongue[J].Oral Surg Oral Med Oral Pathol Oral Radiol,2012,113(4):488-494.DOI: 10.1016/j.tripleo.2011.04.023.
    [36]
    LiuS,JinMN,QuanYS,et al.Transdermal delivery of relatively high molecular weight drugs using novel self-dissolving microneedle arrays fabricated from hyaluronic acid and their characteristics and safety after application to the skin[J].Eur J Pharm Biopharm,2014,86(2):267-276.DOI: 10.1016/j.ejpb.2013.10.001.
    [37]
    MatsuoK,HirobeS,YokotaY,et al.Transcutaneous immunization using a dissolving microneedle array protects against tetanus, diphtheria, malaria, and influenza[J].J Control Release,2012,160(3):495-501.DOI: 10.1016/j.jconrel.2012.04.001.
    [38]
    LiuS,JinMN,QuanYS,et al.The development and characteristics of novel microneedle arrays fabricated from hyaluronic acid, and their application in the transdermal delivery of insulin[J].J Control Release,2012,161(3):933-941.DOI: 10.1016/j.jconrel.2012.05.030.
    [39]
    KatsumiH,LiuS,TanakaY,et al.Development of a novel self-dissolving microneedle array of alendronate, a nitrogen-containing bisphosphonate: evaluation of transdermal absorption, safety, and pharmacological effects after application in rats[J].J Pharm Sci,2012,101(9):3230-3238.DOI: 10.1002/jps.23136.
    [40]
    MönkäreJ,Reza NejadnikM,BaccoucheK,et al.IgG-loaded hyaluronan-based dissolving microneedles for intradermal protein delivery[J].J Control Release,2015,218:53-62.DOI: 10.1016/j.jconrel.2015.10.002.
    • Relative Articles
    • Supplements(0)
    • Cited By
  • 加载中

Catalog

    通讯作者: 陈斌, bchen63@163.com
    • 1. 

      沈阳化工大学材料科学与工程学院 沈阳 110142

    1. 本站搜索
    2. 百度学术搜索
    3. 万方数据库搜索
    4. CNKI搜索

    Figures(7)  / Tables(2)

    Get Citation
    PDF
    XML

    Article Metrics

    Article views (2687) PDF downloads(56) Cited by()
    Proportional views
    Related

    Copyright © Chinese Journal of Burns京ICP备07035254号-14

    E-mail：shaoshangzazhi@163.com

    Website：https://zhsszz.xml-journal.net/

    Supported by: Beijing Renhe Information Technology Co. Ltd    

    /

    DownLoad:  Full-Size Img  PowerPoint
    Return
    Return