Knockout of MicroRNA-155 Ameliorates the Th17/Th9 Immune Response and Promotes Wound Healing

Abstract

MiRNAs are integral for maintaining immune homeostasis and self-tolerance. In this study, qPCR analyses were performed to determine which miRNAs play an important role in wound healing. Next, an experiment in a model of wound healing was performed, and histology, mRNA expression and T-cell subpopulations in wound tissue were analyzed. The accelerated experiments were performed by local injection of either rIL-17A and/or rIL-9 after wound healing. In vitro, the differentiation of Th17/Th9 in miR-155^+/+ or miR-155^−/− mice was investigated, and the target genes of miR155 were analyzed. From our findings, miR-155^−/− in mice promoted wound healing and weakened T cell-mediated inflammation, especially in IL-17/IL-9, and less severe skin fibrosis developed in the mice. rIL-17A and/or rIL-9 could exacerbate inflammatory injury and delay wound healing. We also demonstrated that miR-155^−/− led to a defect in the differentiation of Th17/Th9 in vitro, and this effect of IL-17/IL-9 might be related to the expression of C-maf, which is a target gene of miR155. MiR-155 regulated IL-17/IL-9-related inflammation in wound healing and might be a potential therapeutic target to attenuate the inflammatory response in wound tissue and promote the closure of wound injuries.

Keywords

wound healing / T cells / miR155 / C-maf / IL-17A / IL-9

References

[1] WilgusTA. Immune cells in the healing skin wound: influential players at each stage of repair. Pharmacol Res, 2008, 58(2): 112-116

[2] RoyS, KhannaS, RinkC, et al.. Characterization of the acute temporal changes in excisional murine cutaneous wound inflammation by screening of the wound-edge transcriptome. Physiol Genomics, 2008, 34(2): 162-184

[3] WernerS, GroseR. Regulation of wound healing by growth factors and cytokines. Physiol Rev, 2003, 83(3): 835-870

[4] SehmT, SachseC, FrenzelC, et al.. miR-196 is an essential early-stage regulator of tail regeneration, upstream of key spinal cord patterning events. Dev Biol, 2009, 334(2): 468-480

[5] BiswasS, RoyS, BanerjeeJ, et al.. Hypoxia inducible microRNA 210 attenuates keratinocyte proliferation and impairs closure in a murine model of ischemic wounds. Proc Natl Acad Sci U S A, 2010, 107(15): 6976-6981

[6] WangT, ZhangL, ShiC, et al.. TGF-beta-induced miR-21 negatively regulates the antiproliferative activity but has no effect on EMT of TGF-beta in HaCaT cells. Int J Biochem Cell Biol, 2012, 44(2): 366-376

[7] BerteroT, GastaldiC, Bourget-PonzioI, et al.. miR-483-3p controls proliferation in wounded epithelial cells. Faseb J, 2011, 25(9): 3092-3105

[8] YangX, WangJ, GuoSL, et al.. miR-21 promotes keratinocyte migration and re-epithelialization during wound healing. Int J Biol Sci, 2011, 7(5): 685-690

[9] Martinez-NunezRT, LouafiF, FriedmannPS, et al.. MicroRNA-155 modulates the pathogen binding ability of dendritic cells (DCs) by down-regulation of DC-specific intercellular adhesion molecule-3 grabbing non-integrin (DC-SIGN). J Biol Chem, 2009, 284(24): 16334-16342

[10] DivekarAA, DubeyS, GangalumPR, et al.. Dicer insufficiency and microRNA-155 overexpression in lupus regulatory T cells: an apparent paradox in the setting of an inflammatory milieu. J Immunol, 2011, 186(2): 924-930

[11] TakagiT, NaitoY, MizushimaK, et al.. Increased expression of microRNA in the inflamed colonic mucosa of patients with active ulcerative colitis. J Gastroenterol Hepatol, 2010, 25(Suppl1): S129-33

[12] StanczykJ, PedrioliDM, BrentanoF, et al.. Altered expression of MicroRNA in synovial fibroblasts and synovial tissue in rheumatoid arthritis. Arthritis Rheum, 2008, 58: 1001-1009

[13] Nazari-JahantighM, WeiY, NoelsH, et al.. MicroRNA-155 promotes atherosclerosis by repressing Bcl6 in macrophages. J Clin Invest, 2012, 122(11): 4190-4202

[14] KollsJK, LindenA. Interleukin-17 family members and inflammation. Immunity, 2004, 21(4): 467-476

[15] NogralesKE, KruegerJG. Anti-cytokine therapies for psoriasis. Exp Cell Res, 2011, 317: 1293-1300

[16] RoderoMP, HodgsonSS, HollierB, et al.. Reduced Il17a expression distinguishes a Ly6c(lo)MHCII(hi) macrophage population promoting wound healing. J Invest Dermatol, 2013, 133(3): 783-792

[17] LiH, RostamiA. IL-9: basic biology, signaling pathways in CD4+ T cells and implications for autoimmunity. J Neuroimmune Pharmacol, 2010, 5(2): 198-129

[18] TemannUA, LaouarY, EynonEE, et al.. IL9 leads to airway inflammation by inducing IL13 expression in airway epithelial cells. Int Immunol, 2007, 19(1): 1-10

[19] O’ConnellRM, TaganovKD, BoldinMP, et al.. MicroRNA-155 is induced during the macrophage inflammatory response. Proc Natl Acad Sci U S A, 2007, 104(5): 1604-1609

[20] RutzS, NoubadeR, EidenschenkC, et al.. Transcription factor c-Maf mediates the TGF-beta-dependent suppression of IL-22 production in T(H)17 cells, 2011, 12(12): 1238-1245

[21] ChicaybamL, SodreAL, CurzioBA, et al.. An efficient low cost method for gene transfer to T lymphocytes. PLoS One, 2013, 8(3): e60298

[22] SunY, YeP, WuJ, et al.. Inhibition of intimal hyperplasia in murine aortic allografts by the oral administration of the transforming growth factor-beta receptor I kinase inhibitor SD-208. J Heart Lung Transplant, 2014, 33(6): 654-661

[23] DivekarAA, DubeyS, GangalumPR, et al.. Dicer insufficiency and microRNA-155 overexpression in lupus regulatory T cells: an apparent paradox in the setting of an inflammatory milieu. J Immunol, 2011, 186(2): 924-930

[24] KollsJK, LindenA. Interleukin-17 family members and inflammation. Immunity, 2004, 21: 467-476

[25] RodriguezA, VigoritoE, ClareS, et al.. Requirement of bic/microRNA-155 for normal immune function, 2007, 316(5824): 608-611

[26] ElyamanW, BradshawEM, UyttenhoveC, et al.. IL-9 induces differentiation of TH17 cells and enhances function of FoxP3+ natural regulatory T cells, 2009, 106(31): 12885-12890

[27] BanerjeeJ, ChanYC, SenCK. MicroRNAs in skin and wound healing. Physiol Genomics, 2011, 43(10): 543-556

[28] SandM, GambichlerT, SandD, et al.. MicroRNAs and the skin: Tiny players in the body’s largest organ. J Dermatol Sci, 2009, 53(3): 169-175

[29] BerteroT, GastaldiC, Bourget-PonzioI, et al.. miR-483-3p controls proliferation in wounded epithelial cells. FASEB J, 2011, 25(9): 3092-3105

[30] YangX, WangJ, GuoSL, et al.. miR-21 promotes keratinocyte migration and re-epithelialization during wound healing. Int J Biol Sci, 2011, 7(5): 685-690

[31] LiZ, HassanMQ, JafferjiM, et al.. Biological functions of miR-29b contribute to positive regulation of osteoblast differentiation. J Biol Chem, 2009, 284(23): 15676-15684

[32] van RooijE, SutherlandLB, ThatcherJE, et al.. Dysregulation of microRNAs after myocardial infarction reveals a role of miR-29 in cardiac fibrosis. Proc Natl Acad Sci U S A, 2008, 105(35): 13027-13032

[33] YangL, LiuJ, BaiX, et al.. Acute downregulation of miR-155 at wound sites leads to a reduced fibrosis through attenuating inflammatory response. Biochem Bioph Res Co, 2014, 453(1): 153-159

[34] van SolingenC, AraldiE, Chamorro-JorganesA, et al.. Improved repair of dermal wounds in mice lacking microRNA-155. J Cell Mol Med, 2014, 18(6): 1104-1112

[35] O’ConnellRM, TaganovKD, BoldinMP, et al.. MicroRNA-155 is induced during the macrophage inflammatory response. Proc Natl Acad Sci U S A, 2007, 104(5): 1604-1609

[36] AdolphVR, DiSantoSK, BleacherJC, et al.. The potential role of the lymphocyte in fetal wound healing. J Pediatr Surg, 1993, 28(10): 1316-1320

[37] Hopkinson-WoolleyJ, HughesD, GordonS, et al.. Macrophage recruitment during limb development and wound healing in the embryonic and foetal mouse. J Cell Sci, 1994, 107(Pt5): 1159-1167

[38] OlutoyeOO, YagerDR, CohenIK, et al.. Lower cytokine release by fetal porcine platelets: a possible explanation for reduced inflammation after fetal wounding. J Pediatr Surg, 1996, 31(1): 91-95

[39] DoviJV, HeLK, DiPietroLA. Accelerated wound closure in neutrophil-depleted mice. J Leukoc Biol, 2003, 73(4): 448-455

[40] GorenI, MullerE, SchiefelbeinD, et al.. Systemic anti-TNFalpha treatment restores diabetes-impaired skin repair in ob/ob mice by inactivation of macrophages. J Invest Dermatol, 2007, 127(9): 2259-2267

[41] ParkH, LiZ, YangXO, et al.. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17. Nat Immunol, 2005, 6(11): 1133-1141

[42] RoderoMP, HodgsonSS, HollierB, et al.. Reduced Il17a expression distinguishes a Ly6c(lo)MHCII(hi) macrophage population promoting wound healing. J Invest Dermatol, 2013, 133(3): 783-792

[43] AbdelilahS, LatifaK, EsraN, et al.. Functional expression of IL-9 receptor by human neutrophils from asthmatic donors: role in IL-8 release. J Immunol, 2001, 166(4): 2768-2774

[44] LittleFF, CruikshankWW, CenterDM. Il-9 stimulates release of chemotactic factors from human bronchial epithelial cells. Am J Respir Cell Mol Biol, 2001, 25(3): 347-352

[45] LouahedJ, ZhouY, MaloyWL, et al.. Interleukin 9 promotes influx and local maturation of eosinophils. Blood, 2001, 97(4): 1035-1042

[46] BauquetAT, JinH, PatersonAM, et al.. The costimulatory molecule ICOS regulates the expression of c-Maf and IL-21 in the development of follicular T helper cells and TH-17 cells. Nat Immunol, 2009, 10(2): 167-175

[47] LuLF, LindEF, GondekDC, et al.. Mast cells are essential intermediaries in regulatory T-cell tolerance. Nature, 2006, 442(7106): 997-1002

[48] SatoK, MiyoshiF, YokotaK, et al.. Marked induction of c-Maf protein during Th17 cell differentiation and its implication in memory Th cell development. J Biol Chem, 2011, 286(17): 14963-14971

[49] RutzS, NoubadeR, EidenschenkC, et al.. Transcription factor c-Maf mediates the TGF-beta-dependent suppression of IL-22 production in T(H)17 cells. Nat Immunol, 2011, 12(12): 1238-1245

[50] PoulinLF, RichardM, Le MoineA, et al.. Interleukin-9 promotes eosinophilic rejection of mouse heart allografts. Transplantation, 2003, 76(3): 572-577

[51] BilleterAT, HellmannJ, RobertsH, et al.. MicroRNA-155 potentiates the inflammatory response in hypothermia by suppressing IL-10 production. FASEB J, 2014, 28(12): 5322-5336

[52] QuinnSR, ManganNE, CaffreyBE, et al.. The role of Ets2 transcription factor in the induction of microRNA-155 (miR-155) by lipopolysaccharide and its targeting by interleukin-10. J Biol Chem, 2014, 289: 4316-4325