Email Alerts
RSS
Volume 38 Issue 10
Oct.  2022
Turn off MathJax
Article Contents
Article Navigation >  CHINESE JOURNAL OF BURNS AND WOUNDS  > 2022  >  38(10): 905-913
Song W,Li Z,Zhu SJ,et al.Application of three-dimensional bioprinting ink containing platelet-rich plasma derived from human umbilical cord blood in the treatment of full-thickness skin defects in nude mice[J].Chin J Burns Wounds,2022,38(10):905-913.DOI: 10.3760/cma.j.cn501225-20220618-00243.
Citation: Song W,Li Z,Zhu SJ,et al.Application of three-dimensional bioprinting ink containing platelet-rich plasma derived from human umbilical cord blood in the treatment of full-thickness skin defects in nude mice[J].Chin J Burns Wounds,2022,38(10):905-913.DOI: 10.3760/cma.j.cn501225-20220618-00243.
shu

Application of three-dimensional bioprinting ink containing platelet-rich plasma derived from human umbilical cord blood in the treatment of full-thickness skin defects in nude mice

doi: 10.3760/cma.j.cn501225-20220618-00243

  • Research Center for Wound Repair and Tissue Regeneration, Medical Innovation Research Department, the PLA General Hospital, Beijing 100048, China

Funds:

National Key Research and Development Program of China 2017YFC1103303

Youth Science Foundation Project of National Natural Science Foundation of China 32000969

Shanghai Wang Zhengguo Foundation for Traumatic Medicine Growth Factor Rejuvenation Plan SZYZ-TR-03

More Information
    Abstract
  •   Objective  To investigate the printability and cytocompatibility of sodium alginate-gelatin (AG) bioink containing platelet-rich plasma derived from human umbilical cord blood (HUCB-PRP), named HUCB-PRP-AG bioink, and the effect of the three-dimensionally printed tissue with the bioink on full-thickness skin defect wounds in nude mice.  Methods  The method of experimental research was used. HUCB-PRP-AG bioinks with 2.5%, 5.0%, and 10.0% of HUCB-PRP by volume were prepared and named 1P-AG, 2P-AG, and 4P-AG, respectively. The appearances of AG, 1P-AG, 2P-AG, and 4P-AG at room temperature were observed, and their viscosity and storage/loss modulus were measured by a rotational rheometer. The above four bioinks were used for three-dimensional bioprinting respectively, and the appearances of the printed tissue were observed (the printed tissue was subsequently cross-linked and used). The four kinds of bioprinted tissue were respectively co-cultured with human umbilical vein endothelial cells (HUVECs) in Transwell chambers with HUVEC special medium for 24 h, and the cell proliferation level was detected by cell counting kit 8 (n=3). The four kinds of bioprinted tissue were respectively cultured in Dulbecco's modified eagle medium for 12, 24, and 48 h, which were dried and weighed, and the degradation rate was calculated (n=3). The expression of vascular endothelial growth factor (VEGF) in the culture supernatant of 1P-AG, 2P-AG, or 4P-AG cultured in phosphate buffer solution at 0.5, 24.0, and 48.0 h was detected by enzyme-linked immunosorbent assay (n=5). Sixteen female BALB/c-NU nude mice aged 6-8 weeks were selected to establish a full-thickness skin defect wound model on the back and were divided into conventional control group with wounds being covered with medical hydrocolloid dressing alone, HUCB-PRP group with additional HUCB-PRP dripping to the wounds, AG group additionally covered with AG printed tissue, and 4P-AG group additionally covered with 4P-AG printed tissue, respectively (with 4 nude mice in each group). The wound healing of 3 nude mice in each group was observed on post injury day (PID) 4, 8, and 14, and the wound healing rate was calculated. The wound tissue of the remaining nude mouse in each group was collected on PID 8, the histopathological changes were observed after hematoxylin and eosin staining, and the CD31-positive new blood vessels were observed after immunohistochemical staining. Data were statistically analyzed with analysis of variance for repeated measurement, least significant difference test, and Bonferroni correction.  Results  At room temperature, AG, 1P-AG, 2P-AG, and 4P-AG were semi-transparent liquid, and AG was light yellow, while 1P-AG, 2P-AG, and 4P-AG were light red but the color successively deepened. The viscosity of AG, 1P-AG, 2P-AG, and 4P-AG decreased with the increase of shear rate at the temperature of 10 ℃ and shear rate of 0.1-10.0 s-1; the storage moduli of the four bioinks were greater than the loss moduli at the temperature of 10 ℃ and angular frequency range of 1-100 rad/s. Both the resolution and morphology of the printed tissue of four bioinks were similar. The proliferation levels of HUVECs co-cultured with 1P-AG, 2P-AG, and 4P-AG printed tissue for 24 h were 0.885±0.030, 1.126±0.032, and 1.156±0.045, respectively, which were significantly higher than 0.712±0.019 of HUVECs co-cultured with AG printed tissue (P<0.01). The proliferation levels of HUVECs co-cultured with 2P-AG and 4P-AG printed tissue for 24 h were significantly higher than the level of HUVECs co-cultured with 1P-AG printed tissue (P<0.01). The degradation rates of 1P-AG, 2P-AG, and 4P-AG printed tissue were significantly higher than those of AG printed tissue at 12, 24, and 48 h of culture (P<0.01). The degradation rates of 2P-AG and 4P-AG printed tissue at 24 and 48 h of culture were significantly higher than those of 1P-AG printed tissue (P<0.01). The degradation rate of 4P-AG printed tissue at 12 h of culture was significantly higher than that of 1P-AG printed tissue (P<0.01), and the degradation rates of 4P-AG printed tissue at 24 and 48 h of culture were significantly higher than those of 2P-AG printed tissue (P<0.01). At 0.5, 24.0, and 48.0 h of culture, the expressions of VEGF in the culture supernatant of 2P-AG printed tissue were significantly higher than those of 1P-AG printed tissue (P<0.01), and the expressions of VEGF in the culture supernatant of 1P-AG and 2P-AG printed tissue were significantly lower than those of 4P-AG printed tissue (P<0.01). The wounds of nude mice in conventional control group and HUCB-PRP group were dry and smaller on PID 8 compared with those on PID 4, and the wounds of nude mice in HUCB-PRP group were smaller with no scabs on PID 14 compared with those in conventional control group. The printed tissue on the wound of nude mice in AG and 4P-AG groups was significantly degraded with no obvious exudation being observed on the wounds on PID 4, the wounds were significantly epithelialized and smaller on PID 8, and there was no scab on the wound on PID 14. The wounds of nude mice in 4P-AG group were completely epithelialized on PID 14. Compared with those in conventional control group, the wound healing rate of nude mice in AG group was significantly decreased on PID 4 (P<0.05), and the wound healing rates of nude mice in HUCB-PRP group and 4P-AG group at all time points after injury and in AG group on PID 8 and 14 were significantly increased (P<0.01). Compared with those in HUCB-PRP group, the wound healing rates of nude mice were significantly decreased on PID 4 and 8 in AG group and on PID 4 in 4P-AG group (P<0.01), while the wound healing rates of nude mice were significantly increased on PID 14 in AG group and on PID 8 and 14 in 4P-AG group (P<0.01). The wound healing rate of nude mice in 4P-AG group was significantly higher than that in AG group at all time points after injury (P<0.01). On PID 8, a large number of inflammatory cells infiltration, a small amount of new microvessels, and a small amount of CD31-positive new blood vessels were observed in the wounds of nude mice in conventional control group; a large number of inflammatory cells infiltration, abundant new microvessels, and quite a lot CD31-positive new blood vessels were observed in the wounds of nude mice in HUCB-PRP group; light inflammatory inflammation, a small amount of new microvessels, and a small amount of CD31-positive new blood vessels were observed in the wounds of nude mice in AG group; light inflammatory inflammation, a large number of new microvessels, and a large number of CD31-positive new blood vessels were observed in the wounds of nude mice in 4P-AG group.  Conclusions  HUCB-PRP-AG bioink has good printability and cytocompatibility, and its three-dimensionally printed tissue can promote vascularization of full-thickness skin defect wounds in nude mice and accelerate wound healing.

     

    • FullText(HTML)
  • loading
    • References(29)
  • [1]
    ChenS,ShiY,LuoY,et al.Layer-by-layer coated porous 3D printed hydroxyapatite composite scaffolds for controlled drug delivery[J].Colloids Surf B Biointerfaces,2019,179:121-127.DOI: 10.1016/j.colsurfb.2019.03.063.
    [2]
    da SilvaLP,ReisRL,CorreloVM,et al.Hydrogel-based strategies to advance therapies for chronic skin wounds[J].Annu Rev Biomed Eng,2019,21:145-169.DOI: 10.1146/annurev-bioeng-060418-052422.
    [3]
    HuH,XuFJ.Rational design and latest advances of polysaccharide-based hydrogels for wound healing[J].Biomater Sci,2020,8(8):2084-2101.DOI: 10.1039/d0bm00055h.
    [4]
    XiaS,WengT,JinR,et al.Curcumin-incorporated 3D bioprinting gelatin methacryloyl hydrogel reduces reactive oxygen species-induced adipose-derived stem cell apoptosis and improves implanting survival in diabetic wounds[J/OL].Burns Trauma,2022,10:tkac001[2022-06-18].https://pubmed.ncbi.nlm.nih.gov/35291229/.DOI: 10.1093/burnst/tkac001.
    [5]
    YaoB,WangR,WangY,et al.Biochemical and structural cues of 3D-printed matrix synergistically direct MSC differentiation for functional sweat gland regeneration[J].Sci Adv,2020,6(10):eaaz1094.DOI: 10.1126/sciadv.aaz1094.
    [6]
    张超,李曌,宋薇,等.含人脂肪来源蛋白复合物的三维生物打印墨水的创面修复效应初探[J].中华烧伤杂志,2021,37(11):1011-1023.DOI: 10.3760/cma.j.cn501120-20210813-00282.
    [7]
    SongW,YaoB,ZhuD,et al.3D-bioprinted microenvironments for sweat gland regeneration[J/OL].Burns Trauma,2022,10:tkab044[2022-06-18].https://pubmed.ncbi.nlm.nih.gov/35071651/.DOI: 10.1093/burnst/tkab044.
    [8]
    LopesSV,CollinsMN,ReisRL,et al.Vascularization approaches in tissue engineering: recent developments on evaluation tests and modulation[J].ACS Appl Bio Mater,2021,4(4):2941-2956.DOI: 10.1021/acsabm.1c00051.
    [9]
    BelderbosME,LevyO,MeyaardL,et al.Plasma-mediated immune suppression: a neonatal perspective[J].Pediatr Allergy Immunol,2013,24(2):102-113.DOI: 10.1111/pai.12023.
    [10]
    CoxST,DanbyR,HernandezD,et al.Functional characterisation and analysis of the soluble NKG2D ligand repertoire detected in umbilical cord blood plasma[J].Front Immunol,2018,9:1282.DOI: 10.3389/fimmu.2018.01282.
    [11]
    CoxST,MadrigalJA,SaudemontA.Three novel allelic variants of the RAET1E/ULBP4 gene in humans[J].Tissue Antigens,2012,80(4):390-392.DOI: 10.1111/j.1399-0039.2012.01933.x.
    [12]
    SambergM,StoneR2nd,NatesanS,et al.Platelet rich plasma hydrogels promote in vitro and in vivo angiogenic potential of adipose-derived stem cells[J].Acta Biomater,2019,87:76-87.DOI: 10.1016/j.actbio.2019.01.039.
    [13]
    SteinleA,LiP,MorrisDL,et al.Interactions of human NKG2D with its ligands MICA, MICB, and homologs of the mouse RAE-1 protein family[J].Immunogenetics,2001,53(4):279-287.DOI: 10.1007/s002510100325.
    [14]
    VersuraP,ProfazioV,BuzziM,et al.Efficacy of standardized and quality-controlled cord blood serum eye drop therapy in the healing of severe corneal epithelial damage in dry eye[J].Cornea,2013,32(4):412-418.DOI: 10.1097/ICO.0b013e3182580762.
    [15]
    EvertsP,OnishiK,JayaramP,et al.Platelet-rich plasma: new performance understandings and therapeutic considerations in 2020[J].Int J Mol Sci,2020,21(20):7794.DOI: 10.3390/ijms21207794.
    [16]
    LjubimovAV,SaghizadehM.Progress in corneal wound healing[J].Prog Retin Eye Res,2015,49:17-45.DOI: 10.1016/j.preteyeres.2015.07.002.
    [17]
    NagarajaS,ChenL,DiPietroLA,et al.Computational analysis identifies putative prognostic biomarkers of pathological scarring in skin wounds[J].J Transl Med,2018,16(1):32.DOI: 10.1186/s12967-018-1406-x.
    [18]
    FréchetteJP,MartineauI,GagnonG.Platelet-rich plasmas: growth factor content and roles in wound healing[J].J Dent Res,2005,84(5):434-439.DOI: 10.1177/154405910508400507.
    [19]
    SirchiaG,RebullaP,MozziF,et al.A quality system for placental blood banking[J].Bone Marrow Transplant,1998,21 Suppl 3:S43-47.
    [20]
    BuzziM,VersuraP,GrigoloB,et al.Comparison of growth factor and interleukin content of adult peripheral blood and cord blood serum eye drops for cornea and ocular surface diseases[J].Transfus Apher Sci,2018,57(4):549-555.DOI: 10.1016/j.transci.2018.06.001.
    [21]
    RebullaP,PupellaS,SantodiroccoM,et al.Multicentre standardisation of a clinical grade procedure for the preparation of allogeneic platelet concentrates from umbilical cord blood[J].Blood Transfus,2016,14(1):73-79.DOI: 10.2450/2015.0122-15.
    [22]
    ReddyLVK,MuruganD,MullickM,et al.Recent approaches for angiogenesis in search of successful tissue engineering and regeneration[J].Curr Stem Cell Res Ther,2020,15(2):111-134.DOI: 10.2174/1574888X14666191104151928.
    [23]
    KoleskyDB,TrubyRL,GladmanAS,et al.3D bioprinting of vascularized, heterogeneous cell-laden tissue constructs[J].Adv Mater,2014,26(19):3124-3130.DOI: 10.1002/adma.201305506.
    [24]
    KobayashiY,SaitaY,TakakuT,et al.Platelet-rich plasma (PRP) accelerates murine patellar tendon healing through enhancement of angiogenesis and collagen synthesis[J].J Exp Orthop,2020,7(1):49.DOI: 10.1186/s40634-020-00267-1.
    [25]
    BerndtS,CarpentierG,TurziA,et al.Angiogenesis is differentially modulated by platelet-derived products[J].Biomedicines,2021,9(3):251.DOI: 10.3390/biomedicines9030251.
    [26]
    中国老年医学学会烧创伤分会.浓缩血小板制品在创面修复中应用的全国专家共识(2020版)[J].中华烧伤杂志,2020,36(11):993-1002.DOI: 10.3760/cma.j.cn501120-20200507-00256.
    [27]
    LiY,MouS,XiaoP,et al.Delayed two steps PRP injection strategy for the improvement of fat graft survival with superior angiogenesis[J].Sci Rep,2020,10(1):5231.DOI: 10.1038/s41598-020-61891-6.
    [28]
    MartinoMM,BriquezPS,RangaA,et al.Heparin-binding domain of fibrin(ogen) binds growth factors and promotes tissue repair when incorporated within a synthetic matrix[J].Proc Natl Acad Sci U S A,2013,110(12):4563-4568.DOI: 10.1073/pnas.1221602110.
    [29]
    ZhangX,YaoD,ZhaoW,et al.Engineering platelet-rich plasma based dual-network hydrogel as a bioactive wound dressing with potential clinical translational value[J]. Adv Funct Mater,2021,31(8):2009258.DOI: 10.1002/adfm.202009258.
    • 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(5)  / Tables(3)

    Get Citation
    PDF
    XML

    Article Metrics

    Article views (4886) PDF downloads(39) 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