Evaluating the Effects of Imiquimod on Paths of TLRs and Inflammatory Cytokines Signaling in Infected Macrophages with Leishmania major in Vitro and in Vivo
-
Parisa Ebrahimisadr
,
Fatemeh Ghaffarifar
,
Javad Jabari
,
John Horton
,
Zohreh Sharifi
,
Abdolhossein Dalimi
,
Mohammad Saaid Dayer
Abstract
Background: Leishmania major is an obligate and intracellular pathogen and the macrophages are the cell hosts for L. major. Imiquimod stimulates macrophages to secrete different cytokines via the expression of TLRs.
Methods: This study was carried out in the Department of Parasitology, Faculty of Medical Sciences, Tarbiat Modares University, in 2018. The effect of imiquimod was investigated on non-infected and infected macrophages with L. major on the expression of Toll-like receptors (TLRs) and inflammatory cytokines. TLRs play an important role in enhancing the proceeding of phagocytosis and killing parasites. Moreover, the cytokines such as TNFα, IL6, and IL1, are often identified in inflammatory conditions as interfering targets in treatment. Healthy macrophages and macrophages infected with Leishmania major parasites were affected by different concentrations of imiquimod, after that the expression of TLR genes (TLR1, TLR2, TLR3, TLR4, TLR7 and TLR9) and cytokines were evaluated by real time RT-PCR. For experiments in laboratory animals, infected BALB/c mice were exposed to imiquimod and then isolated peritoneal macrophages.
Results: The expression of TLR2 decreased in non-infected macrophages were affected by the imiquimod. The expression level of TLR7 in healthy macrophages, decreased and the difference with control group was significant. Imiquimod increases the expression of inflammatory cytokines and IL12 in mouse macrophages and also decrease the expression of IL10.
Conclusion: This suggests that imiquimod may improve the therapeutic effects in infected mice with Leishmania major. Imiquimod causes that TLR2 decreased expression but TLR7 and TLR9 increased expression. Imiquimod as TLR7 agonist, enhance the recovery of leishmaniasis.
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22. Ebrahimisadr P, Ghaffarifar F, JABARI J, et al. Therapeutic and preventive effects of morphine against Leishmania major and evaluation the expression of TLRs and cytokines in infected macrophages in vitro and in BALB/c mice. Preprint article.
23. Jabari J, Ghaffarifar F, Horton J, et al. Evaluation of Morphine with Imiquimod as Opioid Growth Factor Receptor or Nalmefene as Opioid Blocking Drug on Leishmaniasis Caused by Leishmania major in Vitro. Iran J Parasitol. 2019; 14(3):394-403.
24. Pandey SP, Doyen N, Mishra GC, et al. TLR9- deficiency reduces TLR1, TLR2 and TLR3 expressions in Leishmania major-infected macrophages. Exp Parasitol. 2015; 154:82-86.
25. Tauer JT, Abdullah S, Rauch F. Effect of AntiTGF-β Treatment in a Mouse Model of Severe Osteogenesis Imperfecta. J Bone Miner Res. 2019; 34(2):207-214.
26. Ribeiro Gomes FL, Moniz De Souza MC, Alexandre Moreira MS, et al. Neutrophils activate macrophages for intracellular killing of Leishmania major through recruitment of TLR4 by neutrophil elastase. J Immunol. 2007; 179:3988–3994.
27. Ribeiro Gomes FL, Otero AC, Gomes NA, et al. Macrophage interactions with neutrophils regulate Leishmania major infection. J Immunol. 2004; 172:4454-4462.
28. Dong L, Uzonna JE. The early interaction of Leishmania with macrophages and dendritic cells and its influence on the host immune response. Front Cell Infect Microbiol. 2012; 2: 83.
29. Hemmi H, Kaisho T, Takeuchi O, et al. Small anti-viral compounds activate immune cells via the TLR7 MyD88-dependent signaling pathway. Nat Immunol. 2002; 3:196–200.
30. Scho¨n M, Bong AB, Drewniok C, et al. Tumor-selective induction of apoptosis and the small- molecule immune res-ponsemodifier imiquimod. J Natl Cancer Inst. 2003; 95:1138–1149.
31. Sacks D, Noben-Trauth N. The immunology of susceptibility and resistance to Leishmania major in mice. Nat Rev Immunol. 2002; 2:845–58.
32. Medzhitov R, Janeway CA. Innate immunity: the virtues of a nonclonal system of recognition. Cell. 1997; 91 (3): 295–298.
33. Akira S, Takeda K. Toll-like receptor signaling. Nat Rev Immunol. 2004; 4 (7):499–511.
34. Flandin JF, Chano F, Descoteaux A. RNA interference reveals a role for TLR2 and TLR3 in therecognition of Leishmania donovani promastigotes by interferon primed macrophages. Eur J Immunol. 2006; 36: 411–420.
35. Kropf P, Freudenberg MA, Modolell M, et al. Toll-like receptor 4 contributes to efficient control of infection with the protozoan parasite Leishmania major. Infect Immun. 2004; 72:1920-1928.
36. Kropf P, Freudenberg N, Kalis C, et al. Infection of receptor C57BL/10ScCr and C57BL/10ScNCr mice with Leishmania major reveals a role for Toll Like 4 in the control of parasite replication. J Leukoc Biol. 2004;76: 48-57.
37. Ribeiro-Gomes FL, Moniz-de-Souza MCA, Alexandre-Moreira MS, et al. Neutrophils activate macrophages for intracellular killing of Leishmania major through recruitment of TLR4 by neutrophil elastase. J Immunol. 2007; 179: 3988-3994.
38. Whitaker SM, Colmenares M, Pestana KG, et al. Leishmania pifanoi proteoglycolipid complex P8 induces macrophage cytokine production through Toll-Like Receptor 4. Infect Immun. 2008; 76: 2149-2156.
39. Zhang P, Yang M, Chen C, et al. Toll-Like Receptor 4 (TLR4)/Opioid Receptor Pathway Crosstalk and Impact on Opioid Analgesia, Immune Function, and Gastrointestinal Motility. Front Immunol. 2020; 11:1455.
40. Schleicher U, Liese J, Knippertz I, et al. NK cell activation in visceral leishmaniasis requires TLR9, myeloid DCs, and IL-12, but is independent of plasmacytoid DCs. J Exp Med. 2007; 204: 893–906.
41. Singal P, Singh PP. Leishmania donovani amastigote component induced colony- stimulating factor production by macrophages: modulation by morphine. Microbes Infect. 2005;7 (2):148–156.
42. O’Garra A, Vieira P. TH1 cells control themselves by producing interleukin-10. Nat Rev Immunol. 2007; 7(6):425–8.
43. Belkaid Y, Hoffmann KF, Mendez S, et al. The role of interleukin (IL)-10 in the persistence of Leishmania major in the skin after healing and the therapeutic potential of Anti–IL-10 receptor antibody for sterile cure. J Exp Med. 2001; 194(10):1497–1506.
2. Sundar S, More DK, Singh MK, et al. Failure of pentavalent antimony in visceral leishmaniasis in India: report from the center of the Indian epidemic. Clin Infect Dis. 2000; 31(4): 1104-1107.
3. Beheshti N, Soflaei S, Shakibaie M, et al. Efficacy of biogenic selenium nanoparticles against Leishmania major: in vitro and in vivo studies. J Trace Elem Med Biol. 2013; 27(3): 203-207.
4. Ghaffarifar F, Jorjani O, Sharifi Z, et al. Enhancement of immune response induced by DNA vaccine cocktail expressing complete LACK and TSA genes against Leishmania major. APMIS. 2013; 121:290-298.
5. Ghaffarifar F. Plasmid DNA vaccines: where are we now. Drugs Today (Barc). 2018; 54:315-333.
6. Ghaffarifar F, Heydari F, Dalimi A, et al. Evaluation of apoptotic and antileishmanial activities of Artemisinin on promastigotes and BALB/C mice infected with Leishmania major. Iran J Parasitol. 2015; 10: 258-67.
7. Miranda-Verastegui C, Llanos-Cuentas A, Arevalo I, et al. Randomized, double-blind clinical trial of topical imiquimod 5% with parenteral meglumine antimoniate in the treatment of cutaneous leishmaniasis in Peru. Clin Infect Dis. 2005; 40(10):1395-403.
8. Dockrell D, Kinghorn G. Imiquimod and resiquimod as novel immunomodulators. J Antimicrob Chemother. 2001; 48: 751-755.
9. Zaki L, Ghaffarifar F, Sharifi Z, et al. Effect of Imiquimod on Tachyzoites of Toxoplasma gondii and Infected Macrophages in vitro and in BALB/c Mice. Front Cell Infect Microbiol. 2020; 10:387.
10. Sidky YA, Borden EC, Weeks CE, et al. Inhibition of murine tumor growth by an interferon-inducing imidazoquinolinamine. Cancer Res. 1992; 52(13): 3528-3533.
11. Stanley, M. Imiquimod and the imidazoquinolones: mechanism of action and therapeutic potential. Clin Exp Dermatol. 2002; 27(7): 571-577.
12. Miller RL, Gerster JF, Owens ML, et al. Review article imiquimod applied topically: a novel immune response modifier and new class of drug. Int J Immunopharmacol. 1999; 21(1): 1-14.
13. Buates S, Matlashewski G. Treatment of experimental leishmaniasis with the immunomodulators imiquimod and S-28463: efficacy and mode of action. J Infect Dis. 1999; 179(6): 1485-1494.
14. Geisse J, Caro I, Lindholm J, et al. Imiquimod 5% cream for the treatment of superficial basal cell carcinoma: results from two phase III, randomized, vehicle-controlled studies. J Am Acad Dermatol. 2004; 50(5): 722-733.
15. Akira S, Takeda K, Kaisho T. Toll-like receptors: critical proteins linking innate and acquired immunity. Nat Immunol. 2001; 2: 675-680.
16. Gazzinelli RT, Denkers EY. Protozoan encounters with Toll-like receptor signaling pathways: implications for host parasitism. Nat Rev Immunol. 2006; 6:895–906.
17. Pasare C, Medzhitov R. Toll-like receptors: linking innate and adaptive immunity. Microbes Infect. 2004; 6:1382–1387.
18. Fukao T, S Koyasu. PI3K and negative regulation of TLR signaling. Trends Immunol. 2003; 24(7): 358-363.
19. Murphy KM, Weaver C. Janeway's immuno biology. 9th Edition. 2018.
20. Kawai T, Akira S, TLR signaling. Cell Death Differ. 2006; 13(5): 816-825.
21. Colonna M. TLR pathways and IFN‐regulatory factors: To each its own. Eur J Immunol. 2007; 37(2): 306-309.
22. Ebrahimisadr P, Ghaffarifar F, JABARI J, et al. Therapeutic and preventive effects of morphine against Leishmania major and evaluation the expression of TLRs and cytokines in infected macrophages in vitro and in BALB/c mice. Preprint article.
23. Jabari J, Ghaffarifar F, Horton J, et al. Evaluation of Morphine with Imiquimod as Opioid Growth Factor Receptor or Nalmefene as Opioid Blocking Drug on Leishmaniasis Caused by Leishmania major in Vitro. Iran J Parasitol. 2019; 14(3):394-403.
24. Pandey SP, Doyen N, Mishra GC, et al. TLR9- deficiency reduces TLR1, TLR2 and TLR3 expressions in Leishmania major-infected macrophages. Exp Parasitol. 2015; 154:82-86.
25. Tauer JT, Abdullah S, Rauch F. Effect of AntiTGF-β Treatment in a Mouse Model of Severe Osteogenesis Imperfecta. J Bone Miner Res. 2019; 34(2):207-214.
26. Ribeiro Gomes FL, Moniz De Souza MC, Alexandre Moreira MS, et al. Neutrophils activate macrophages for intracellular killing of Leishmania major through recruitment of TLR4 by neutrophil elastase. J Immunol. 2007; 179:3988–3994.
27. Ribeiro Gomes FL, Otero AC, Gomes NA, et al. Macrophage interactions with neutrophils regulate Leishmania major infection. J Immunol. 2004; 172:4454-4462.
28. Dong L, Uzonna JE. The early interaction of Leishmania with macrophages and dendritic cells and its influence on the host immune response. Front Cell Infect Microbiol. 2012; 2: 83.
29. Hemmi H, Kaisho T, Takeuchi O, et al. Small anti-viral compounds activate immune cells via the TLR7 MyD88-dependent signaling pathway. Nat Immunol. 2002; 3:196–200.
30. Scho¨n M, Bong AB, Drewniok C, et al. Tumor-selective induction of apoptosis and the small- molecule immune res-ponsemodifier imiquimod. J Natl Cancer Inst. 2003; 95:1138–1149.
31. Sacks D, Noben-Trauth N. The immunology of susceptibility and resistance to Leishmania major in mice. Nat Rev Immunol. 2002; 2:845–58.
32. Medzhitov R, Janeway CA. Innate immunity: the virtues of a nonclonal system of recognition. Cell. 1997; 91 (3): 295–298.
33. Akira S, Takeda K. Toll-like receptor signaling. Nat Rev Immunol. 2004; 4 (7):499–511.
34. Flandin JF, Chano F, Descoteaux A. RNA interference reveals a role for TLR2 and TLR3 in therecognition of Leishmania donovani promastigotes by interferon primed macrophages. Eur J Immunol. 2006; 36: 411–420.
35. Kropf P, Freudenberg MA, Modolell M, et al. Toll-like receptor 4 contributes to efficient control of infection with the protozoan parasite Leishmania major. Infect Immun. 2004; 72:1920-1928.
36. Kropf P, Freudenberg N, Kalis C, et al. Infection of receptor C57BL/10ScCr and C57BL/10ScNCr mice with Leishmania major reveals a role for Toll Like 4 in the control of parasite replication. J Leukoc Biol. 2004;76: 48-57.
37. Ribeiro-Gomes FL, Moniz-de-Souza MCA, Alexandre-Moreira MS, et al. Neutrophils activate macrophages for intracellular killing of Leishmania major through recruitment of TLR4 by neutrophil elastase. J Immunol. 2007; 179: 3988-3994.
38. Whitaker SM, Colmenares M, Pestana KG, et al. Leishmania pifanoi proteoglycolipid complex P8 induces macrophage cytokine production through Toll-Like Receptor 4. Infect Immun. 2008; 76: 2149-2156.
39. Zhang P, Yang M, Chen C, et al. Toll-Like Receptor 4 (TLR4)/Opioid Receptor Pathway Crosstalk and Impact on Opioid Analgesia, Immune Function, and Gastrointestinal Motility. Front Immunol. 2020; 11:1455.
40. Schleicher U, Liese J, Knippertz I, et al. NK cell activation in visceral leishmaniasis requires TLR9, myeloid DCs, and IL-12, but is independent of plasmacytoid DCs. J Exp Med. 2007; 204: 893–906.
41. Singal P, Singh PP. Leishmania donovani amastigote component induced colony- stimulating factor production by macrophages: modulation by morphine. Microbes Infect. 2005;7 (2):148–156.
42. O’Garra A, Vieira P. TH1 cells control themselves by producing interleukin-10. Nat Rev Immunol. 2007; 7(6):425–8.
43. Belkaid Y, Hoffmann KF, Mendez S, et al. The role of interleukin (IL)-10 in the persistence of Leishmania major in the skin after healing and the therapeutic potential of Anti–IL-10 receptor antibody for sterile cure. J Exp Med. 2001; 194(10):1497–1506.
| Files | ||
| Issue | Vol 20 No 2 (2025) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijpa.v20i2.19021 | |
| Keywords | ||
| Leishmania major Imiquimod Macrophages Inflammatory Cytokines | ||
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How to Cite
1.
Ebrahimisadr P, Ghaffarifar F, Jabari J, Horton J, Sharifi Z, Dalimi A, Dayer MS. Evaluating the Effects of Imiquimod on Paths of TLRs and Inflammatory Cytokines Signaling in Infected Macrophages with Leishmania major in Vitro and in Vivo. Iran J Parasitol. 2025;20(2):178-192.
