Leishmania parasites are transmitted to vertebrate hosts by female phlebotomine sand flies as they bloodfeed by lacerating the upper capillaries of the dermis with their barbed mouthparts. In the sand fly midgut secreted proteophosphoglycans from Leishmania form a biological plug known as the promastigote secretory gel (PSG), which blocks the gut and facilitates the regurgitation of infective parasites. The interaction between the wound created by the sand fly bite and PSG is not known. Here we nanoinjected a sand fly egested dose of PSG into BALB/c mouse skin that lead to the differential expression of 7,907 transcripts. These transcripts were transiently up-regulated during the first 6 hours post-wound and enriched for pathways involved in inflammation, cell proliferation, fibrosis, epithelial cell differentiation and wound remodelling. We found that PSG significantly accelerated wound healing in vitro and in mice; which was associated with an early up-regulation of transcripts involved in inflammation (IL-1β, IL-6, IL-10, TNFα) and inflammatory cell recruitment (CCL2, CCL3, CCL4, CXCL2), followed 6 days later by enhanced expression of transcripts associated with epithelial cell proliferation, fibroplasia and fibrosis (FGFR2, EGF, EGFR, IGF1). Dermal expression of IGF1 was enhanced following an infected sand fly bite and was acutely responsive to the deposition of PSG but not the inoculation of parasites or sand fly saliva. Antibody blockade of IGF1 ablated the gel's ability to promote wound closure in mouse ears and significantly reduced the virulence of Leishmania mexicana infection delivered by an individual sand fly bite. Dermal macrophages recruited to air-pouches on the backs of mice revealed that IGF1 was pivotal to the PSG's ability to promote macrophage alternative activation and Leishmania infection. Our data demonstrate that through the regurgitation of PSG Leishmania exploit the wound healing response of the host to the vector bite by promoting the action of IGF1 to drive the alternative activation of macrophages.

Department of Clinical Research Faculty of Infectious Tropical Diseases London School of Hygiene and Tropical Medicine London United Kingdom

Department of Disease Control London School of Hygiene and Tropical Medicine London United Kingdom

Department of Immunology and Infection Faculty of Infectious Tropical Diseases London School of Hygiene and Tropical Medicine London United Kingdom

Department of Medicine Section of Immunology Imperial College London St Mary's Campus London United Kingdom

Department of Parasitology Faculty of Science Charles University Prague Czech Republic

Division of Biomedical and Life Sciences Lancaster University Lancaster United Kingdom

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Sacks D, Kamhawi S (2001) Molecular aspects of parasite-vector and vector-host interactions in leishmaniasis. Annu Rev Microbiol 55: 453–483. doi: 10.1146/annurev.micro.55.1.453 PubMed DOI

Lestinova T, Rohousova I, Sima M, de Oliveira CI, Volf P (2017) Insights into the sand fly saliva: Blood-feeding and immune interactions between sand flies, hosts, and PubMed DOI PMC

Titus RG, Ribeiro JM (1988) Salivary gland lysates from the sand PubMed

Kamhawi S, Belkaid Y, Modi G, Rowton E, Sacks DL (2000) Protection against cutaneous leishmaniasis resulting from bites of uninfected sand flies. Science 290: 1351–1354. PubMed

Oliveira F, Lawyer PG, Kamhawi S, Valenzuela JG (2008) Immunity to distinct sand fly salivary proteins primes the anti- PubMed DOI PMC

Rogers ME, Ilg T, Nikolaev AV, Ferguson MA, Bates PA (2004) Transmission of cutaneous leishmaniasis by sand flies is enhanced by regurgitation of fPPG. Nature 430: 463–467. doi: 10.1038/nature02675 PubMed DOI PMC

Bates PA (2007) Transmission of PubMed DOI PMC

Stierhof YD, Bates PA, Jacobson RL, Rogers ME, Schlein Y, et al. (1999) Filamentous proteophosphoglycan secreted by PubMed DOI

Rogers ME, Chance ML, Bates PA (2002) The role of promastigote secretory gel in the origin and transmission of the infective stage of PubMed

Rogers ME, Bates PA (2007) PubMed DOI PMC

Kimblin N, Peters N, Debrabant A, Secundino N, Egen J, et al. (2008) Quantification of the infectious dose of PubMed DOI PMC

Rogers M, Kropf P, Choi BS, Dillon R, Podinovskaia M, et al. (2009) Proteophosophoglycans regurgitated by PubMed DOI PMC

Rogers M, Corware K, Müller I, Bates P (2010) PubMed DOI

Ilg T (2000) Proteophosphoglycans of PubMed

Niu Y, Miao M, Cao X, Song F, Ji X, Dong J, Lu S. (2014) Infiltration of macrophages and their phenotype in the healing process of full-thickness wound in rat. Zhonghua Shao Shang Za Zhi 30:109–115. PubMed

Murray HW, Xiang Z, Ma X. (2006) Responses to PubMed

Bauer S, Müller T, Hamm S. (2009) Pattern recognition by Toll-like receptors. Adv Exp Med Biol 653:15–34. PubMed

Tuon FF, Amato VS, Bacha HA, Almusawi T, Duarte MI, Amato Neto V. (2008) Toll-like receptors and leishmaniasis. Infect Immun 76:866–872. doi: 10.1128/IAI.01090-07 PubMed DOI PMC

Anthony RM, Rutitzky LI, Urban JF Jr, Stadecker MJ, Gause WC. (2007) Protective immune mechanisms in helminth infection. Nat Rev Immunol 7:975–987. doi: 10.1038/nri2199 PubMed DOI PMC

Byers DE, Holtzman MJ. (2011) Alternatively activated macrophages and airway disease. Chest 140:768–774. doi: 10.1378/chest.10-2132 PubMed DOI PMC

Wanasen N, Soong L. (2008) L-arginine metabolism and its impact on host immunity against PubMed DOI PMC

Raes G, Van den Bergh R, De Baetselier P, Ghassabeh GH, Scotton C, et al. (2005) Arginase-1 and Ym1 are markers for murine, but not human, alternatively activated myeloid cells. J Immunol 174: 6561–6562. PubMed

Kropf P, Fuentes JM, Fähnrich E, Arpa L, Herath S et al. (2005) Arginase and polyamine synthesis are key factors in the regulation of experimental leishmaniasis PubMed DOI

Roberts SC, Tancer MJ, Polinsky MR, Gibson KM, Heby O, et al. (2004) Arginase plays a pivotal role in polyamine precursor metabolism in PubMed DOI

Yizengaw E, Getahun M, Tajebe F, Cruz Cervera E, Adem E, et al. (2016) Visceral Leishmaniasis Patients Display Altered Composition and Maturity of Neutrophils as well as Impaired Neutrophil Effector Functions. Front Immunol 7: 517 doi: 10.3389/fimmu.2016.00517 PubMed DOI PMC

Mortazavi H, Sadeghipour P, Taslimi Y, Habibzadeh S, Zali F, et al. (2016) Comparing acute and chronic human cutaneous leishmaniasis caused by PubMed DOI

Peters NC, Egen JG, Secundino N, Debrabant A, Kimblin N, et al. (2008) PubMed DOI PMC

Sakthianandeswaren A, Elso CM, Simpson K, Curtis JM, Kumar B, et al. (2005) The wound repair response controls outcome to cutaneous leishmaniasis. Proc Natl Acad Sci USA 102: 15551–15556. doi: 10.1073/pnas.0505630102 PubMed DOI PMC

Bertho AL, Santiago MA, Coutinho SG (1994) An experimental model of the production of metastases in murine cutaneous leishmaniasis. J Parasitol 80: 93–99. PubMed

Wortmann GW, Aronson NE, Miller RS, Blazes D, Oster CN (2000) Cutaneous leishmaniasis following local trauma: a clinical pearl. Clin Infect Dis 31: 199–201. doi: 10.1086/313924 PubMed DOI

Späth GF, Beverley SM. (2001) A lipophosphoglycan-independent method for isolation of infective PubMed DOI

Telleria EL, Sant'Anna MR, Ortigão-Farias JR, Pitaluga AN, Dillon VM, et al. (2012) Caspar-like gene depletion reduces PubMed DOI PMC

de La Llave E, Lecoeur H, Besse A, Milon G, Prina E, et al. (2011) A combined luciferase imaging and reverse transcription polymerase chain reaction assay for the study of PubMed DOI

Schroeder A, Mueller O, Stocker S, Salowsky R, Leiber M, et al. (2006) The RIN: an RNA integrity number for assigning integrity values to RNA measurements. BMC Mol Biol 7: 3 doi: 10.1186/1471-2199-7-3 PubMed DOI PMC

Bolstad BM, Irizarry RA, Astrand M, Speed TP. (2003) A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 19: 185–193. PubMed

Jain N, Thatte J, Braciale T, Ley K, O'Connell M, et al. (2003) Local-pooled-error test for identifying differentially expressed genes with a small number of replicated microarrays. Bioinformatics 19: 1945–1951. PubMed

Benjamini Y, Hochberg Y. (1990) More powerful procedures for multiple significance testing. Stat Med 9: 811–818. PubMed

Prina E, Roux E, Mattei D, Milon G. (2007) PubMed DOI

Hellemans J, Mortier G, De Paepe A, Speleman F, Vandesompele J. (2007) qBase relative quantification framework and software for management and automated analysis of realtime quantitative PCR data. Genome Biol 8: R19 doi: 10.1186/gb-2007-8-2-r19 PubMed DOI PMC

Lecoeur H, de La Llave E, Osorio Y Fortéa J, Goyard S, Kiefer-Biasizzo H, et al. (2010) Sorting of PubMed DOI

Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, et al. (2002) Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol 3: RESEARCH0034. PubMed PMC

Andersen CL, Jensen JL, Orntoft TF. (2004) Normalization of real-time quantitative reverse transcription-PCR data: a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets. Cancer Res 64: 5245–5250. doi: 10.1158/0008-5472.CAN-04-0496 PubMed DOI

Liang CC, Park AY, Guan JL. (2007) PubMed DOI

Osorio y Fortea J, de La Llave E, Regnault B, Coppee JY, Milon G, et al. (2009) Transcriptional signatures of BALB/c mouse macrophages housing multiplying PubMed DOI PMC

Krenn HW, Aspöck H. (2012) Form, function and evolution of the mouthparts of blood-feeding Arthropoda. Arthropod Struct Dev 41:101–118. doi: 10.1016/j.asd.2011.12.001 PubMed DOI

Delavary MB, van der Veer WM, van Egmond M, Niessen FB, Beelen RH. (2011) Macrophages in skin injury and repair. Immunobiol 216: 753–762. PubMed

Sato Y, Ohshima T, Kondo T. (1999) Regulatory role of endogenous interleukin-10 in cutaneous inflammatory response of murine wound healing. Biochem Biophys Res Commun 265: 194–199. doi: 10.1006/bbrc.1999.1455 PubMed DOI

Moore KW, O'Garra A, de Waal Malefyt R, Vieira P, Mosmann TR. (1993) Interleukin-10. Annu Rev Immunol 11: 165–190. doi: 10.1146/annurev.iy.11.040193.001121 PubMed DOI

Belkaid Y, Hoffmann KF, Mendez S, Kamhawi S, Udey MC, et al. (2001) The role of interleukin (IL)-10 in the persistence of PubMed PMC

Ajuebor MN, Das AM, Virag L, Szabo C, Perretti M. (1999) Regulation of macrophage inflammatory protein-1 alpha expression and function by endogenous interleukin-10 in a model of acute inflammation. Biochem Biophys Res Commun 255: 279–282. doi: 10.1006/bbrc.1999.0196 PubMed DOI

Rheinwald JG, Green H. (1977) Epidermal growth factor and the multiplication of cultured human epidermal keratinocytes. Nature 265: 421–424. PubMed

Tokumaru S, Higashiyama S, Endo T, Nakagawa T, Miyagawa JI, et al. (2000) Ectodomain shedding of epidermal growth factor receptor ligands is required for keratinocyte migration in cutaneous wound healing. J Cell Biol 151: 209–220. PubMed PMC

Frank S, Hubner G, Breier G, Longaker MT, Greenhalgh DG, et al. (1995) Regulation of vascular endothelial growth factor expression in cultured keratinocytes. Implications for normal and impaired wound healing. J Biol Chem 270: 12607–12613. PubMed

Gille J, Khalik M, Konig V, Kaufmann R. (1998) Hepatocyte growth factor/scatter factor (HGF/SF) induces vascular permeability factor (VPF/VEGF) expression by cultured keratinocytes. J Invest Dermatol 111: 1160–1165. doi: 10.1046/j.1523-1747.1998.00418.x PubMed DOI

Knighton DR, Silver IA, Hunt TK. (1981) Regulation of wound-healing angiogenesis-effect of oxygen gradients and inspired oxygen concentration. Surgery 90: 262–270. PubMed

Singer AJ, Clark RA. (1999) Cutaneous wound healing. N Engl J Med 341: 738–746. doi: 10.1056/NEJM199909023411006 PubMed DOI

Albina JE, Mills CD, Barbul A, Thirkill CE, Henry WL Jr, Mastrofrancesco B, et al. (1988) Arginine metabolism in wounds. Am J Physiol (1988) 254:E459–67. doi: 10.1152/ajpendo.1988.254.4.E459 PubMed DOI

Albina JE, Mills CD, Henry WL Jr, Caldwell MD. Temporal expression of different pathways of 1-arginine metabolism in healing wounds. J Immunol (1990) 144:3877–80. PubMed

Iniesta V, Gómez-Nieto LC, Molano I, Mohedano A, Carcelén J, Mirón C, Alonso C, Corraliza I. (2002) Arginase I induction in macrophages, triggered by Th2-type cytokines, supports the growth of intracellular Leishmania parasites. Parasite Immunol 24:113–118. PubMed

Vendrame CM, Carvalho MD, Tempone AG, Goto H. (2014) Insulin-like growth factor-I induces arginase activity in PubMed DOI PMC

Pauleau AL, Rutschman R, Lang R, Pernis A, Watowich SS, Murray PJ. (2004) Enhancer-mediated control of macrophage-specific arginase I expression. J Immunol 172:7565–7573. PubMed

El Kasmi KC, Qualls JE, Pesce JT, Smith AM, Thompson RW, Henao-Tamayo M, et al. (2008) Toll-like receptor-induced arginase 1 in macrophages thwarts effective immunity against intracellular pathogens. Nat Immunol 9:1399–1406. doi: 10.1038/ni.1671 PubMed DOI PMC

Goto H, Gomes CM, Corbett CE, Monteiro HP, Gidlund M. (1998) Insulin-like growth factor I is a growth-promoting factor for PubMed PMC

Vendrame CM, Carvalho MD, Rios FJ, Manuli ER, Petitto-Assis F, et al. (2007) Effect of insulin-like growth factor-I on PubMed DOI

Reis LC, Ramos-Sanchez EM, Goto H (2013) The interactions and essential effects of intrinsic insulin-like growth factor-I on PubMed DOI PMC

Gomes R, Oliveira F. (2012) The immune response to sand fly salivary proteins and its influence on PubMed DOI PMC

Solbach W, Laskay T. (2000) The host response to PubMed

Wiethe C, Debus A, Mohrs M, Steinkasserer A, Lutz M, Gessner A. (2008) Dendritic cell differentiation state and their interaction with NKT cells determine Th1/Th2 differentiation in the murine model of PubMed

Belkaid Y, Rouse BT. (2005) Natural regulatory T cells in infectious disease. Nat Immunol 6:353–360. doi: 10.1038/ni1181 PubMed DOI

Sacks D, Kamhawi S. (2001) Molecular aspects of parasite-vector and vector-host interactions in leishmaniasis. Annu Rev Microbiol 55: 453–483. doi: 10.1146/annurev.micro.55.1.453 PubMed DOI

Giraud E, Lecoeur H, Soubigou G, Coppée JY, Milon G, et al. (2012) Distinct transcriptional signatures of bone marrow-derived C57BL/6 and DBA/2 dendritic leucocytes hosting live PubMed DOI PMC

Leoni G, Neumann PA, Sumagin R, Denning TL, Nusrat A. (2015) Wound repair: role of immune-epithelial interactions. Mucosal Immunol. 8:959–968. doi: 10.1038/mi.2015.63 PubMed DOI PMC

Naiyer MM, Saha S, Hemke V, Roy S, Singh S, Musti KV, Saha B. (2013) Identification and characterization of a human IL-10 receptor antagonist. Hum Immunol 74:28–31. doi: 10.1016/j.humimm.2012.09.002 PubMed DOI

Lucas M, Zhang X, Prasanna V, Mosser DM. (2005) ERK activation following macrophage FcgammaR ligation leads to chromatin modifications at the IL-10 locus. J Immunol 175:469–477. PubMed

Osorio EY, Travi BL, daCruz AM, Saldarriaga OA, Medina AA, Melby PC. (2014). Growth factor and Th2 cytokine signalling pathways converge at STAT6 to promote arginase expression in progressive experimental visceral leishmaniasis. PLoS Pathog 10:e1004165 doi: 10.1371/journal.ppat.1004165 PubMed DOI PMC

Tonkin J, Sampson L, Gallego-Colon E, Barberi L, et al. (2015) Monocyte/Macrophage-derived IGF-1 Orchestrates Murine Skeletal Muscle Regeneration and Modulates Autocrine Polarization. Mol Ther 23: 1189–1200. doi: 10.1038/mt.2015.66 PubMed DOI PMC

Yin Y, Hua H, Li M, Liu S, Kong Q, et al. (2016) mTORC2 promotes type I insulin-like growth factor receptor and insulin receptor activation through the tyrosine kinase activity of mTOR. Cell Research. 26: 46–65. doi: 10.1038/cr.2015.133 PubMed DOI PMC

Chen F, Liu Z, Wu W, Rozo C, Bowdridge S, et al. (2012) An essential role for TH2-type responses in limiting acute tissue damage during experimental helminth infection. Nat Med 18: 260–266. doi: 10.1038/nm.2628 PubMed DOI PMC

Prates DB, Araújo-Santos T, Luz NF, Andrade BB, França-Costa J, et al. (2011) PubMed DOI

van Zandbergen G, Klinger M, Mueller A, Dannenberg S, Gebert A, et al. (2004) Cutting edge: neutrophil granulocyte serves as a vector for PubMed

Peters NC, Kimblin N, Secundino N, Kamhawi S, Lawyer P, et al. (2009) Vector transmission of leishmania abrogates vaccine-induced protective immunity. PLoS Pathog 5: e1000484 doi: 10.1371/journal.ppat.1000484 PubMed DOI PMC

Atayde VD, Aslan H, Townsend S, Hassani K, Kamhawi S, et al. (2015) Exosome Secretion by the Parasitic Protozoan PubMed DOI PMC

Cho KA, Suh JW, Lee KH, Kang JL, Woo SY. (2012) IL-17 and IL-22 enhance skin inflammation by stimulating the secretion of IL-1beta by keratinocytes via the ROS-NLRP3-caspase-1 pathway. Int Immunol 24: 147–158. doi: 10.1093/intimm/dxr110 PubMed DOI

Boaventura VS, Santos CS, Cardoso CR, de Andrade J, Dos Santos WL, et al. (2010) Human mucosal leishmaniasis: neutrophils infiltrate areas of tissue damage that express high levels of Th17-related cytokines. Eur J Immunol 40: 2830–2836. doi: 10.1002/eji.200940115 PubMed DOI

Rodero MP, Hodgson SS, Hollier B, Combadiere C, Khosrotehrani K. (2013) Reduced Il17a expression distinguishes a Ly6c(lo)MHCII(hi) macrophage population promoting wound healing. J Invest Dermatol 133:783–792. doi: 10.1038/jid.2012.368 PubMed DOI

Pitta MG, Romano A, Cabantous S, Henri S, Hammad A, et al. (2009) IL-17 and IL-22 are associated with protection against human kala azar caused by PubMed DOI PMC

Lopez Kostka S, Dinges S, Griewank K, Iwakura Y, Udey MC, et al. (2009) IL-17 promotes progression of cutaneous leishmaniasis in susceptible mice. J Immunol 182: 3039–3046. doi: 10.4049/jimmunol.0713598 PubMed DOI PMC

Braune J, Weyer U, Hobusch C, Mauer J, Brüning JC, Bechmann I, Gericke M. (2017). IL-6 Regulates M2 Polarization and Local Proliferation of Adipose Tissue Macrophages in Obesity. J Immunol. 198: 2927–2934. doi: 10.4049/jimmunol.1600476 PubMed DOI

Mauer J, Chaurasia B, Goldau J, Vogt MC, Ruud J, et al. (2014) Signaling by IL-6 promotes alternative activation of macrophages to limit endotoxemia and obesity-associated resistance to insulin. Nat Immunol. 15: 423–430. doi: 10.1038/ni.2865 PubMed DOI PMC

The First Non-LRV RNA Virus in Leishmania

Promastigote secretory gel from natural and unnatural sand fly vectors exacerbate Leishmania major and Leishmania tropica cutaneous leishmaniasis in mice