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Toll like receptors and acute allograft rejection
Transplant Immunology 17 (2006) 11 – 15 www.elsevier.com/locate/trim
Review
Toll like receptors and acute allograft rejection Daniel R. Goldstein ⁎ Section of Cardiovascular Medicine, Department of Internal Medicine Yale University School of Medicine, 333 Cedar St, 3 FMP PO BOX 208017, New Haven, CT, USA Received 29 August 2006; accepted 13 September 2006
Abstract Toll like receptors (TLR) are critical innate immune receptors expressed on a variety of cells including dendritic cells that are activated by the presence of invading pathogens. However, there is emerging evidence that TLR signaling participates in inflammation that may occur in the absence of overt infection. In solid organ transplantation there is increasing evidence, both in experimental and human studies, that TLR activation is involved in the innate immune recognition of allografts. Further investigation of how innate immunity impacts solid organ transplantation may lead to improved therapies for transplant recipients. © 2006 Elsevier B.V. All rights reserved. Keywords: Dendritic cell; Toll like receptor; Innate immunity
Contents 1. 2.
Introduction . . . . . . . . . . . . . . . TLR immunobiology . . . . . . . . . . 2.1. TLRs and their ligands . . . . . 2.2. TLR signaling pathways. . . . . 2.3. Interaction of TLRs and immune 3. TLRs and acute allograft rejection . . . 3.1. Rodent studies . . . . . . . . . . 3.2. Human studies. . . . . . . . . . 4. Conclusion . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . .
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1. Introduction The initiation of a primary immune response is the result of a balanced interaction of the innate and adaptive immune systems [1]. The innate system acts as the first line of host defense against microbial invasion and consists of different receptors and signaling pathways [2,3]. The first mammalian homologs of the TLRs were discovered nearly 10 years ago [4]. These receptors have remained highly conserved throughout evolution
⁎ Tel.: +1 203 785 3271; fax: +1 203 785 7567. E-mail address: [email protected]. 0966-3274/$ - see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.trim.2006.09.012
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and have been found to key for initiating an innate immune response [5]. The past 50 years has seen unprecedented success in the development of solid organ transplantation as a therapeutic modality for end stage organ disease. This success has been due to multiple factors including improved donor organ harvest and surgical implantation techniques, improved peri-operative management and the development of more tailored immunosuppressive medications. The latter have primarily inhibited components of adaptive immunity, notably T cell function part of the adaptive immune system. However, there is emerging evidence that innate immunity plays an important role in the immune recognition of solid organ allografts.
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D.R. Goldstein / Transplant Immunology 17 (2006) 11–15
2. TLR immunobiology 2.1. TLRs and their ligands TLRs are germ-line encoded innate immune receptors that the host employs to detect the presence of invading pathogens [6,2]. TLRs are expressed on a variety of cell types including antigen presenting cells (APC), epithelial and endothelial cells [3]. To date, there are at least thirteen TLRs discovered in mammals that can detect multiple pathogen associated molecular pattern (PAMP). These include bacterial lipoproteins and lipoteichoic acid (via TLR 2), double stranded RNA (TLR 3), lipopolysaccharide (TLR 4), flagellin (TLR 5), single stranded RNA (TLR 7 and 8), unmethylated CpG (TLR 9), DNA viruses (TLR 9) and protozoal profilin protein (TLR 11) [7,3,8]. Interestingly, TLRs, 1, 2, 4, 5 and 6 are expressed on the cell surface and appear important for bacterial recognition [3]. In contrast, TLRs, 3, 7, and 9 are vital for viral recognition by detecting nucleic acids, which are not necessarily unique to the invading pathogen [3]. However, these TLRs are contained within the late endosomal compartment within the cell, which prevents them being activated by host nucleic acids. Thus, TLRs are an integral part of the host pattern recognition strategy in response to infection. 2.2. TLR signaling pathways TLRs consist extracellulary of leucine rich repeats with a cytoplasmic Toll/interleukin (IL)-1 receptor (TIR) domain that they share with the IL-1 receptor family [2]. Consistent with this homology, TLRs and IL-1Rs share an adaptor protein myeloid differentiation factor 88 (MyD88), which also contains a TIR domain in addition to a death domain [9]. After activation of TLRs MyD88 is recruited to the TLR receptor complex that also includes IL-1 receptor associated kinase (IRAK) and tumor necrosis factor (TNF)-receptor associated factor 6 (TRAF 6) [10]. IRAK and TRAF 6 dissociate from this complex and then initiate the unmasking of the nuclear localization domain of NFκB, causing its translocation to the nucleus where it induces the expression of multiple proinflammatory genes. However, studies over the last 5 years have indicated that other TLR signal adaptors exist. All TLRs, except TLR 3, can signal via MyD88. TLR 2 and 4 can signal via TIRAP (or Mal) although this pathway is MyD88 dependent [7,11–13]. Very recently, it has shown that TIRAP contains a binding domain that allows localization to the plasma membrane upon TLR 4 activation [14]. TIRAP then facilitates the recruitment of MyD88 allowing subsequent TLR signal transduction. TLR 3 and 4 can signal via Trif, which leads to the production of type one interferons via the IRF-3 transcription factor [15–17]. However, type 1 interferons can be produced by viral infections that activate TLR 7 and 9 but this is dependent on MyD88 signaling [18–20]. Other TLR signal adaptors include TRAM, which has been found to transduce a TLR 4-dependent MyD88independent signaling pathway that is important for a late NFκB response [21]. Lastly, other adaptors have been discovered although their specific roles have yet to be elucidated [22].
Activation of TLRs is important for the development of Th1 immunity [23,24].The role of TLR driven immune function and the initiation of Th2 immunity is less well established. There are reports that have provided evidence that activation of specific TLR ligands (e.g., TLR 2 or TLR 5) may induce Th2 immunity under specific experimental conditions [25,26] In addition to proinflammatory pathways, TLR signaling also initiates negative regulatory signals that are thought to help resolve the host inflammatory responses. Such regulatory molecules include IRAK-M, SOCS1, A20, MyD88 short, which appear to negatively regulate TLR immune responses [27–31]. The role of TLR activation in the initiation of Th17 T cell immunity is yet to be elucidated [32]. 2.3. Interaction of TLRs and immune regulatory pathways Emerging evidence has suggested that TLRs and regulatory T cells (T regs) may act in concert to control immune responses. As TLRs are generally activated in response to foreign antigens and hence enhance host immunity, it is rational to expect the suppressive actions of T regs to be impaired during TLR activation. Pasare and Medzhitov elucidated this point by demonstrating that the ability of T regs to suppress cytotoxic T cells in vitro was abrogated if TLRs were activated [33]. This effect was mediated by IL-6 secreted by DCs that allowed reactive T cells to become insensitive to the suppressive effects of T regs. In contrast, Caramalho and colleagues demonstrated that T reg activity is enhanced after stimulation with LPS (a TLR 4 agonist) [34]. An explanation for these apparently contradictory results is that in the work by Pasare the dose of LPS was significantly lower than in the Caramalho study. Thus, it is possible that the lower dose of LPS mimics the initial contact with a pathogen or other foreign antigen (e.g., allograft), resulting in a situation where immunity needs to be enhanced and suppression reduced. In contrast, the higher level of LPS may represent an overwhelming inflammatory process (e.g., septic shock) where enhanced suppression would be beneficial. Other studies have indicated that T regs may express TLRs and their activation may modulate T reg function. Specifically, two studies provided evidence that TLR 2 activation on T regs increases their proliferation but transiently reduces their function. The concept here is that during infection T reg function needs to be transiently impaired so that enhanced immunity may allow elimination of the infection [35,36]. Yet increased inflammation needs to be controlled and hence the increasing numbers of T regs may lead to resolution of inflammation during elimination of the infection. However, other studies have indicated that TLR activation augments T reg function, which may occur by costimulation via heat shock proteins [37,38]. Thus, the impact of TLR immunity on the suppressive function of T regs remains to be fully elucidated. 3. TLRs and acute allograft rejection The implantation of solid organs is not a “natural event” in contrast to most infections. It is surgically induced, accompanied by local ischemia-reperfusion injury and a persistence of
D.R. Goldstein / Transplant Immunology 17 (2006) 11–15
antigenic tissue. Importantly, recent evidence has indicated that TLRs are critical for the initiation of ischemia reperfusion injury [39,40]. Further evidence that TLR signaling is important is ischemia-reperfusion injury comes from myocardial ischemia models that have shown that both TLR 2 and 4 signaling are important for infarct size and subsequent left ventricular dysfunction [41,42]. This type of injury in the setting of solid organ transplantation may release putative endogenous ligands that may activate DCs. This is supported by several reports that “endogenous” innate immune ligands exist that may be released during sterile inflammation. Such putative ligands include heat shock proteins (HSP), uric acid, oligimers of hyaluronan, fibrinogen and chromatin [43–49]. Some of these ligands have been found to signal via TLRs, predominantly TLR 4 [43–47]. These studies remain controversial since the effect of confounding LPS may not have been completely excluded [50]. 3.1. Rodent studies We postulated that the acute antigen independent injury that occurs during donor organ implantation releases innate immune ligands, although the nature of such ligands remains to be elucidated. This is supported by circumstantial evidence that certain innate immune ligands are released during acute allograft rejection both from clinical and experimental studies [51]. Since there is suggestive evidence that endogenous innate immune ligands can induce DC maturation via TLRs we sought to investigate whether TLRs are important for acute allograft rejection. We hypothesized that TLR signaling on either the donor or recipient DC during solid organ transplantation would lead to DC maturation and consequent alloimmune priming. We initially tested this hypothesis in a murine minor mismatched skin allograft model and determined that acute allograft rejection was dependent on TLR driven MyD88 signaling [52]. Furthermore, we showed that MyD88 signaling was important for DC maturation, CD8 alloimmune priming and consequent Th1 dependent alloimmunity in response to transplantation [52]. It is not completely clear from our work which TLRs are involved although TLR 2 had a modest effect and TLR 4 did not appear to be important, in agreement with a later report [53]. Therefore, our work provides evidence that TLR-dependent signaling initiates alloimmunity, indicating that this host defense mechanism is utilized in allograft recognition. In a subsequent study, we found that MyD88 signaling was no longer critical for the rejection of fully allogeneic skin or cardiac allografts [54]. However, MyD88 signaling was important for maximal Th1 alloimmune priming by donor DCs [54]. In both the minor mismatch and fully allogeneic models Th2 immune function remained preserved in the absence of MyD88, suggesting that innate Th2 dependent mechanism may be important for acute allograft rejection. 3.2. Human studies The first study to investigate the role of TLRs in human transplantation occurred in lung transplant recipients. This is
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particularly relevant since lung allografts have persistent exposure to environment pathogens and inhaled toxins. Prior studies have shown that two specific singly-nucleotide polymorphisms (SNPs) Asp299Gly and Thr399Ile are associated with blunted TLR 4 dependent immune responses (e.g., inhaled responses to endotoxin) in humans [10]. Using genetic PCR based assays to genotype donor and recipients, one study demonstrated that recipients that were heterozygote for either of the TLR 4 SNPs manifest a reduced biopsy proven acute rejection rate compared to wild type patients up to 3 years post transplantation [55,56]. The overall rate of biopsy-proven acute rejection was 44% in heterozygotes and 71% in wild types (p = 0.02) with a trend towards reduced chronic rejection in the heterozygotes [56]. Interestingly, TLR 4 mutations in the donor allograft had no impact on acute allograft rejection. A study in a cohort of renal transplant recipients found that the same heterozygote TLR 4 SNPs were associated with a reduced rate of acute rejection (7.4% vs. 26.1%, p = 0.02) over a 95-month follow up period [57]. Additionally, TLR4 mutations in renal transplant recipients were associated with reduced atherosclerotic events in agreement with a prior study [58] but an increased incidence of bacterial infections. Finally, there is evidence that increase TLR expression correlates with endothelial dysfunction in cardiac transplant recipients [59]. Thus, the above studies provide clinical evidence that TLR signaling at the very least participates in acute allograft rejection. What is not yet known and clearly requires further study is whether immunosuppressive regimens can be tailored to recipient TLR genotypes. 4. Conclusion There is increasing evidence from several human and rodent studies that TLRs are important in the innate immune recognition of allografts. However, it is likely that these receptors are not the only innate receptors employed by the host during innate allorecognition. Future studies are warranted to elucidate the nature of these receptors, the ligands that activate them and TLRs and degree of redundancy that occurs between them during allograft rejection. References [1] Banchereau J, Steinman RM. Dendritic cells and the control of immunity. Nature 1998;392(6673):245. [2] Akira S, Takeda K. Toll-like receptor signalling. Nat Rev Immunol 2004;4 (7):499. [3] Iwasaki A, Medzhitov R. Toll-like receptor control of the adaptive immune responses. Nat Immunol 2004;5(10):987. [4] Medzhitov R, Preston-Hurlburt P, Janeway Jr CA. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity. Nature 1997;388(6640):394. [5] Hoffmann JA, R, JM. Drosophila innate immunity: an evolutionary perspective. Nat Immunol 2002;3(2):121. [6] Akira S, Uematsu S, Takeuchi O. Pathogen recognition and innate immunity. Cell 2006;124(4):783. [7] Yarovinsky F, Zhang D, Andersen JF, Bannenberg GJ, Serhan EN, Hayden MS, et al. TLR11 activation of dendritic cells by a protozoan profilin-like protein. Science 2005;308(5728):1626.
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[8] Medzhitov R. Toll-like receptors and innate immunity. Nat Rev Immunol 2001;1(2):135. [9] Barton GM, Medzhitov R. Toll-like receptor signaling pathways. Science 2003;300(5625):1524. [10] Cook C, Pisetsky D, Schwartz D. Toll-like receptors in the pathogeneis of human disease. Nat Immunol 2004;5(10):975. [11] Horng T, Barton GM, Medzhitov R. TIRAP: an adapter molecule in the Toll signaling pathway. [see comments]. Nat Immunol 2001;2(9):835. [12] Horng T, Barton GM, Flavell RA, Medzhitov R. The adaptor molecule TIRAP provides signalling specificity for Toll-like receptors. Nature 2002;420(6913):329. [13] Fitzgerald KA, Palsson-McDermott EM, Bowie AG, et al. Mal (MyD88adapter-like) is required for Toll-like receptor-4 signal transduction. Nature 2001;413(6851):78. [14] Kagan JC, Medzhitov R. Phosphoinositide-mediated adaptor recruitment controls toll-like receptor signaling. Cell 2006;125(5):943. [15] Hoebe K, Du X, Georgel P, et al. Identification of Lps2 as a key transducer of MyD88-independent TIR signalling. Nature 2003;424 (6950):743. [16] Hoebe K, Janssen EM, Kim SO, et al. Upregulation of costimulatory molecules induced by lipopolysaccharide and double-stranded RNA occurs by Trif-dependent and Trif-independent pathways. Nat Immunol 2003;4(12):1223. [17] Yamamoto M, Sato S, Hemmi H, et al. Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway. Science 2003;301(5633):640. [18] Lund JM, Alexopoulou L, Sato A, et al. Recognition of single-stranded RNA viruses by Toll-like receptor 7. Proc Natl Acad Sci U S A 2004;101 (15):5598. [19] Lund J, Sato A, Akira S, Medzhitov R, Iwasaki A. Toll-like receptor 9mediated recognition of herpes simplex virus-2 by plasmacytoid dendritic cells. J Exp Med 2003;198(3):513. [20] Heil F, Hemmi H, Hochrein H, et al. Species-specific recognition of single-stranded rna via toll-like receptor 7 and 8. Science 2004;303 (5663):1526. [21] Yamamoto M, Sato S, Hemmi H, et al. TRAM is specifically involved in the toll-like receptor 4-mediated MyD88-independent signaling pathway. Nat Immunol 2003;4(11):1144. [22] O'Neill LAJ, Fitzgerald KA, Bowie AG. The toll-IL-1 receptor adaptor family grows to five members. Trends Immunol 2003;24(6):286. [23] Schnare M, Barton GM, Holt AC, Takeda K, Akira S, Medzhitov R. Tolllike receptors control activation of adaptive immune responses. Nat Immunol 2001;2(10):947. [24] Jankovic D, Kullberg MC, Hieny S, Caspar P, Collazo CM, Sher A. In the absence of IL-12, CD4(+) T cell responses to intracellular pathogens fail to default to a Th2 pattern and are host protective in an IL-10(−/−) setting. Immunity 2002;16(3):429. [25] Redecke V, Hacker H, Datta SK, et al. Cutting edge: activation of toll-like receptor 2 induces a Th2 immune response and promotes experimental asthma. J Immunol 2004;172(5):2739. [26] Supajatura V, Ushio H, Nakao A, et al. Differential responses of mast cell Toll-like receptors 2 and 4 in allergy and innate immunity. J Clin Invest 2002;109(10):1351. [27] Kobayashi K, Hernandez LD, Galan JE, Janeway Jr CA, Medzhitov R, Flavell RA. IRAK-M is a negative regulator of toll-like receptor signaling. Cell 2002;110(2):191. [28] Kinjyo I, Hanada T, Inagaki-Ohara K, et al. SOCS1/JAB is a negative regulator of LPS-induced macrophage activation. Immunity 2002;17 (5):583. [29] Burns K, Janssens S, Brissoni B, Olivos N, Beyaert R, Tschopp J. Inhibition of interleukin 1 receptor/toll-like receptor signaling through the alternatively spliced, short form of MyD88 is due to its failure to recruit IRAK-4. J Exp Med 2003;197(2):263. [30] Wald D, Qin J, Zhao Z, et al. SIGIRR, a negative regulator of toll-like receptor-interleukin 1 receptor signaling. Nat Immunol 2003;4(9):920. [31] Boone DL, Turer EE, Lee EG, et al. The ubiquitin-modifying enzyme A20 is required for termination of toll-like receptor responses. Nature 2004;5(10):1052.
[32] Weaver CT, Harrington LE, Mangan PR, Gavrieli M, Murphy KM. Th17: an effector CD4 T cell lineage with regulatory T cell ties. Immunity 2006;24(6):677. [33] Pasare C, Medzhitov R. Toll pathway-dependent blockade of CD4 + CD25 + T cell-mediated suppression by dendritic cells. Science 2003;299 (5609):1033. [34] Caramalho I, Lopes-Carvalho T, Ostler D, Zelenay S, Haury M, Demengeot J. Regulatory T cells selectively express toll-like receptors and are activated by lipopolysaccharide. J Exp Med 2003;197(4):403. [35] Netea MG, Sutmuller R, Hermann C, et al. Toll-like receptor 2 suppresses immunity against candida albicans through induction of IL-10 and regulatory T cells. J Immunol 2004;172(6):3712. [36] Liu H, Komai-Koma M, Xu D, Liew FY, et al. Toll-like receptor 2 signaling modulates the functions of CD4+ CD25+ regulatory T cells. Proc Natl Acad Sci U S A 2006;103(18):7048. [37] Peng G, Guo Z, Kiniwa Y, et al. Toll-like receptor 8-mediated reversal of CD4+ regulatory T cell function. Science 2005;309(5739):1380. [38] Zanin-Zhorov A, Cahalon L, Tal G, Margalit R, Lider O, Cohen IR. Heat shock protein 60 enhances CD4+ CD25+ regulatory T cell function via innate TLR2 signaling. J Clin Invest 2006;116(7):2022 [%R 10.1172/ JCI28423]. [39] Boros P, Bromberg JS. New cellular and molecular immune pathways in ischemia/reperfusion injury. Am J Transplant 2006;6(4):652. [40] Zhai Y, Shen X-d, O'Connell R, et al. Cutting edge: TLR4 activation mediates liver ischemia/reperfusion inflammatory response via IFN regulatory factor 3-dependent MyD88-independent pathway. J Immunol 2004;173(12):7115. [41] Oyama J-i, Blais Jr C, Liu X, et al. Reduced myocardial ischemiareperfusion injury in toll-like receptor 4-deficient mice. Circulation 2004;109(6):784. [42] Shishido T, Nozaki N, Yamaguchi S, et al. Toll-like receptor-2 modulates ventricular remodeling after myocardial infarction. Circulation 2003;108 (23):2905. [43] Guillot L, Balloy V, McCormack FX, Golenbock DT, Chignard M, SiTahar M. Cutting edge: the immunostimulatory activity of the lung surfactant protein-A involves toll-like receptor 4. J Immunol 2002;168 (12):5989. [44] Asea A, Kraeft SK, Kurt-Jones EA, et al. HSP70 stimulates cytokine production through a CD14-dependant pathway, demonstrating its dual role as a chaperone and cytokine. Nat Med 2000;6(4):435. [45] Johnson GB, Brunn GJ, Kodaira Y, Platt JL. Receptor-mediated monitoring of tissue well-being via detection of soluble heparan sulfate by toll-like receptor 4. J Immunol 2002;168(10):5233. [46] Ohashi K, Burkart V, Flohe S, Kolb H. Cutting edge: heat shock protein 60 is a putative endogenous ligand of the toll-like receptor-4 complex. J Immunol 2000;164(2):558. [47] Termeer C, Benedix F, Sleeman J, et al. Oligosaccharides of hyaluronan activate dendritic cells via toll-like receptor 4. J Exp Med 2002;195 (1):99. [48] Leadbetter EA, Rifkin IR, Hohlbaum AM, Beaudette BC, Shlomchik MJ, Marshak-Rothstein A. Chromatin-IgG complexes activate B cells by dual engagement of IgM and toll-like receptors. Nature 2002;416 (6881):603. [49] Rock KL, Hearn A, Chen CJ, YS. Natural endogenous adjuvants. Springer Semin Immunopathol 2005;26(3):231. [50] Gaston JS. Heat shock proteins and innate immunity.[comment]. Clin Exp Immunol 2002;127(1):1. [51] Pockley AG. Heat shock proteins, anti-heat shock protein reactivity and allograft rejection. Transplantation 2001;71(11):1503. [52] Goldstein DR, Tesar BM, Akira S, Lakkis FG. Critical role of the toll-like receptor signal adaptor protein MyD88 in acute allograft rejection. J Clin Invest 2003;111(10):1571. [53] Samstein B, Johnson GB, JL P. Toll-like receptor-4 and allograft responses. Transplantation 2004;15(77):475. [54] Tesar BM, Zhang J, Li Q, Goldstein DR. TH1 immune responses to fully MHC mismatched allografts are diminished in the absence of MyD88, a toll like receptor signal adaptor protein. Am J Transplant 2004;4 (9):1429.
D.R. Goldstein / Transplant Immunology 17 (2006) 11–15 [55] Palmer SM, Burch LH, Davis RD, et al. The role of innate immunity in acute allograft rejection after lung transplantation. Am J Respir Crit Care Med 2003;168(6):628. [56] Palmer SM, Burch LH, Trindade AJ, et al. Innate immunity influences long-term outcomes after human lung transplant. Am J Respir Crit Care Med 2005;171(7):780. [57] Ducloux D, Deschamps M, Yannaraki M, et al. Relevance of toll-like receptor-4 polymorphisms in renal transplantation. Kidney Int 2005;67 (6):2454.
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[58] Kiechl S, Lorenz E, Reindl M, et al. Toll-like receptor 4 polymorphisms and atherogenesis. N Engl J Med 2002;347(3):185. [59] Methe H, Zimmer E, Grimm C, Nabauer Ma, Koglin J, et al. Evidence for a Role of toll-like receptor 4 in development of chronic allograft rejection after cardiac transplantation. Transplantation 1954;78(9):1324.
Review
Toll like receptors and acute allograft rejection Daniel R. Goldstein ⁎ Section of Cardiovascular Medicine, Department of Internal Medicine Yale University School of Medicine, 333 Cedar St, 3 FMP PO BOX 208017, New Haven, CT, USA Received 29 August 2006; accepted 13 September 2006
Abstract Toll like receptors (TLR) are critical innate immune receptors expressed on a variety of cells including dendritic cells that are activated by the presence of invading pathogens. However, there is emerging evidence that TLR signaling participates in inflammation that may occur in the absence of overt infection. In solid organ transplantation there is increasing evidence, both in experimental and human studies, that TLR activation is involved in the innate immune recognition of allografts. Further investigation of how innate immunity impacts solid organ transplantation may lead to improved therapies for transplant recipients. © 2006 Elsevier B.V. All rights reserved. Keywords: Dendritic cell; Toll like receptor; Innate immunity
Contents 1. 2.
Introduction . . . . . . . . . . . . . . . TLR immunobiology . . . . . . . . . . 2.1. TLRs and their ligands . . . . . 2.2. TLR signaling pathways. . . . . 2.3. Interaction of TLRs and immune 3. TLRs and acute allograft rejection . . . 3.1. Rodent studies . . . . . . . . . . 3.2. Human studies. . . . . . . . . . 4. Conclusion . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . .
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . regulatory pathways . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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1. Introduction The initiation of a primary immune response is the result of a balanced interaction of the innate and adaptive immune systems [1]. The innate system acts as the first line of host defense against microbial invasion and consists of different receptors and signaling pathways [2,3]. The first mammalian homologs of the TLRs were discovered nearly 10 years ago [4]. These receptors have remained highly conserved throughout evolution
⁎ Tel.: +1 203 785 3271; fax: +1 203 785 7567. E-mail address: [email protected]. 0966-3274/$ - see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.trim.2006.09.012
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and have been found to key for initiating an innate immune response [5]. The past 50 years has seen unprecedented success in the development of solid organ transplantation as a therapeutic modality for end stage organ disease. This success has been due to multiple factors including improved donor organ harvest and surgical implantation techniques, improved peri-operative management and the development of more tailored immunosuppressive medications. The latter have primarily inhibited components of adaptive immunity, notably T cell function part of the adaptive immune system. However, there is emerging evidence that innate immunity plays an important role in the immune recognition of solid organ allografts.
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2. TLR immunobiology 2.1. TLRs and their ligands TLRs are germ-line encoded innate immune receptors that the host employs to detect the presence of invading pathogens [6,2]. TLRs are expressed on a variety of cell types including antigen presenting cells (APC), epithelial and endothelial cells [3]. To date, there are at least thirteen TLRs discovered in mammals that can detect multiple pathogen associated molecular pattern (PAMP). These include bacterial lipoproteins and lipoteichoic acid (via TLR 2), double stranded RNA (TLR 3), lipopolysaccharide (TLR 4), flagellin (TLR 5), single stranded RNA (TLR 7 and 8), unmethylated CpG (TLR 9), DNA viruses (TLR 9) and protozoal profilin protein (TLR 11) [7,3,8]. Interestingly, TLRs, 1, 2, 4, 5 and 6 are expressed on the cell surface and appear important for bacterial recognition [3]. In contrast, TLRs, 3, 7, and 9 are vital for viral recognition by detecting nucleic acids, which are not necessarily unique to the invading pathogen [3]. However, these TLRs are contained within the late endosomal compartment within the cell, which prevents them being activated by host nucleic acids. Thus, TLRs are an integral part of the host pattern recognition strategy in response to infection. 2.2. TLR signaling pathways TLRs consist extracellulary of leucine rich repeats with a cytoplasmic Toll/interleukin (IL)-1 receptor (TIR) domain that they share with the IL-1 receptor family [2]. Consistent with this homology, TLRs and IL-1Rs share an adaptor protein myeloid differentiation factor 88 (MyD88), which also contains a TIR domain in addition to a death domain [9]. After activation of TLRs MyD88 is recruited to the TLR receptor complex that also includes IL-1 receptor associated kinase (IRAK) and tumor necrosis factor (TNF)-receptor associated factor 6 (TRAF 6) [10]. IRAK and TRAF 6 dissociate from this complex and then initiate the unmasking of the nuclear localization domain of NFκB, causing its translocation to the nucleus where it induces the expression of multiple proinflammatory genes. However, studies over the last 5 years have indicated that other TLR signal adaptors exist. All TLRs, except TLR 3, can signal via MyD88. TLR 2 and 4 can signal via TIRAP (or Mal) although this pathway is MyD88 dependent [7,11–13]. Very recently, it has shown that TIRAP contains a binding domain that allows localization to the plasma membrane upon TLR 4 activation [14]. TIRAP then facilitates the recruitment of MyD88 allowing subsequent TLR signal transduction. TLR 3 and 4 can signal via Trif, which leads to the production of type one interferons via the IRF-3 transcription factor [15–17]. However, type 1 interferons can be produced by viral infections that activate TLR 7 and 9 but this is dependent on MyD88 signaling [18–20]. Other TLR signal adaptors include TRAM, which has been found to transduce a TLR 4-dependent MyD88independent signaling pathway that is important for a late NFκB response [21]. Lastly, other adaptors have been discovered although their specific roles have yet to be elucidated [22].
Activation of TLRs is important for the development of Th1 immunity [23,24].The role of TLR driven immune function and the initiation of Th2 immunity is less well established. There are reports that have provided evidence that activation of specific TLR ligands (e.g., TLR 2 or TLR 5) may induce Th2 immunity under specific experimental conditions [25,26] In addition to proinflammatory pathways, TLR signaling also initiates negative regulatory signals that are thought to help resolve the host inflammatory responses. Such regulatory molecules include IRAK-M, SOCS1, A20, MyD88 short, which appear to negatively regulate TLR immune responses [27–31]. The role of TLR activation in the initiation of Th17 T cell immunity is yet to be elucidated [32]. 2.3. Interaction of TLRs and immune regulatory pathways Emerging evidence has suggested that TLRs and regulatory T cells (T regs) may act in concert to control immune responses. As TLRs are generally activated in response to foreign antigens and hence enhance host immunity, it is rational to expect the suppressive actions of T regs to be impaired during TLR activation. Pasare and Medzhitov elucidated this point by demonstrating that the ability of T regs to suppress cytotoxic T cells in vitro was abrogated if TLRs were activated [33]. This effect was mediated by IL-6 secreted by DCs that allowed reactive T cells to become insensitive to the suppressive effects of T regs. In contrast, Caramalho and colleagues demonstrated that T reg activity is enhanced after stimulation with LPS (a TLR 4 agonist) [34]. An explanation for these apparently contradictory results is that in the work by Pasare the dose of LPS was significantly lower than in the Caramalho study. Thus, it is possible that the lower dose of LPS mimics the initial contact with a pathogen or other foreign antigen (e.g., allograft), resulting in a situation where immunity needs to be enhanced and suppression reduced. In contrast, the higher level of LPS may represent an overwhelming inflammatory process (e.g., septic shock) where enhanced suppression would be beneficial. Other studies have indicated that T regs may express TLRs and their activation may modulate T reg function. Specifically, two studies provided evidence that TLR 2 activation on T regs increases their proliferation but transiently reduces their function. The concept here is that during infection T reg function needs to be transiently impaired so that enhanced immunity may allow elimination of the infection [35,36]. Yet increased inflammation needs to be controlled and hence the increasing numbers of T regs may lead to resolution of inflammation during elimination of the infection. However, other studies have indicated that TLR activation augments T reg function, which may occur by costimulation via heat shock proteins [37,38]. Thus, the impact of TLR immunity on the suppressive function of T regs remains to be fully elucidated. 3. TLRs and acute allograft rejection The implantation of solid organs is not a “natural event” in contrast to most infections. It is surgically induced, accompanied by local ischemia-reperfusion injury and a persistence of
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antigenic tissue. Importantly, recent evidence has indicated that TLRs are critical for the initiation of ischemia reperfusion injury [39,40]. Further evidence that TLR signaling is important is ischemia-reperfusion injury comes from myocardial ischemia models that have shown that both TLR 2 and 4 signaling are important for infarct size and subsequent left ventricular dysfunction [41,42]. This type of injury in the setting of solid organ transplantation may release putative endogenous ligands that may activate DCs. This is supported by several reports that “endogenous” innate immune ligands exist that may be released during sterile inflammation. Such putative ligands include heat shock proteins (HSP), uric acid, oligimers of hyaluronan, fibrinogen and chromatin [43–49]. Some of these ligands have been found to signal via TLRs, predominantly TLR 4 [43–47]. These studies remain controversial since the effect of confounding LPS may not have been completely excluded [50]. 3.1. Rodent studies We postulated that the acute antigen independent injury that occurs during donor organ implantation releases innate immune ligands, although the nature of such ligands remains to be elucidated. This is supported by circumstantial evidence that certain innate immune ligands are released during acute allograft rejection both from clinical and experimental studies [51]. Since there is suggestive evidence that endogenous innate immune ligands can induce DC maturation via TLRs we sought to investigate whether TLRs are important for acute allograft rejection. We hypothesized that TLR signaling on either the donor or recipient DC during solid organ transplantation would lead to DC maturation and consequent alloimmune priming. We initially tested this hypothesis in a murine minor mismatched skin allograft model and determined that acute allograft rejection was dependent on TLR driven MyD88 signaling [52]. Furthermore, we showed that MyD88 signaling was important for DC maturation, CD8 alloimmune priming and consequent Th1 dependent alloimmunity in response to transplantation [52]. It is not completely clear from our work which TLRs are involved although TLR 2 had a modest effect and TLR 4 did not appear to be important, in agreement with a later report [53]. Therefore, our work provides evidence that TLR-dependent signaling initiates alloimmunity, indicating that this host defense mechanism is utilized in allograft recognition. In a subsequent study, we found that MyD88 signaling was no longer critical for the rejection of fully allogeneic skin or cardiac allografts [54]. However, MyD88 signaling was important for maximal Th1 alloimmune priming by donor DCs [54]. In both the minor mismatch and fully allogeneic models Th2 immune function remained preserved in the absence of MyD88, suggesting that innate Th2 dependent mechanism may be important for acute allograft rejection. 3.2. Human studies The first study to investigate the role of TLRs in human transplantation occurred in lung transplant recipients. This is
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particularly relevant since lung allografts have persistent exposure to environment pathogens and inhaled toxins. Prior studies have shown that two specific singly-nucleotide polymorphisms (SNPs) Asp299Gly and Thr399Ile are associated with blunted TLR 4 dependent immune responses (e.g., inhaled responses to endotoxin) in humans [10]. Using genetic PCR based assays to genotype donor and recipients, one study demonstrated that recipients that were heterozygote for either of the TLR 4 SNPs manifest a reduced biopsy proven acute rejection rate compared to wild type patients up to 3 years post transplantation [55,56]. The overall rate of biopsy-proven acute rejection was 44% in heterozygotes and 71% in wild types (p = 0.02) with a trend towards reduced chronic rejection in the heterozygotes [56]. Interestingly, TLR 4 mutations in the donor allograft had no impact on acute allograft rejection. A study in a cohort of renal transplant recipients found that the same heterozygote TLR 4 SNPs were associated with a reduced rate of acute rejection (7.4% vs. 26.1%, p = 0.02) over a 95-month follow up period [57]. Additionally, TLR4 mutations in renal transplant recipients were associated with reduced atherosclerotic events in agreement with a prior study [58] but an increased incidence of bacterial infections. Finally, there is evidence that increase TLR expression correlates with endothelial dysfunction in cardiac transplant recipients [59]. Thus, the above studies provide clinical evidence that TLR signaling at the very least participates in acute allograft rejection. What is not yet known and clearly requires further study is whether immunosuppressive regimens can be tailored to recipient TLR genotypes. 4. Conclusion There is increasing evidence from several human and rodent studies that TLRs are important in the innate immune recognition of allografts. However, it is likely that these receptors are not the only innate receptors employed by the host during innate allorecognition. Future studies are warranted to elucidate the nature of these receptors, the ligands that activate them and TLRs and degree of redundancy that occurs between them during allograft rejection. References [1] Banchereau J, Steinman RM. Dendritic cells and the control of immunity. Nature 1998;392(6673):245. [2] Akira S, Takeda K. Toll-like receptor signalling. Nat Rev Immunol 2004;4 (7):499. [3] Iwasaki A, Medzhitov R. Toll-like receptor control of the adaptive immune responses. Nat Immunol 2004;5(10):987. [4] Medzhitov R, Preston-Hurlburt P, Janeway Jr CA. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity. Nature 1997;388(6640):394. [5] Hoffmann JA, R, JM. Drosophila innate immunity: an evolutionary perspective. Nat Immunol 2002;3(2):121. [6] Akira S, Uematsu S, Takeuchi O. Pathogen recognition and innate immunity. Cell 2006;124(4):783. [7] Yarovinsky F, Zhang D, Andersen JF, Bannenberg GJ, Serhan EN, Hayden MS, et al. TLR11 activation of dendritic cells by a protozoan profilin-like protein. Science 2005;308(5728):1626.
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