Innate immune modulation in transplantation: mechanisms, challenges, and opportunities

Organ transplantation is characterized by a sequence of steps that involve operative trauma, organ preservation, and ischemia-reperfusion injury in the transplant recipient. During this process, the release of damage-associated molecular patterns (DAMPs) promotes the activation of innate immune cells via engagement of the toll-like receptor (TLR) system, the complement system, and coagulation cascade. Different classes of effector responses are then carried out by specialized populations of macrophages, dendritic cells, and T and B lymphocytes; these play a central role in the orchestration and regulation of the inflammatory response and modulation of the ensuing adaptive immune response to transplant allografts. Organ function and rejection of human allografts have traditionally been studied through the lens of adaptive immunity; however, an increasing body of work has provided a more comprehensive picture of the pivotal role of innate regulation of adaptive immune responses in transplant and the potential therapeutic implications. Herein we review literature that examines the repercussions of inflammatory injury to transplantable organs. We highlight novel concepts in the pathophysiology and mechanisms involved in innate control of adaptive immunity and rejection. Furthermore, we discuss existing evidence on novel therapies aimed at innate immunomodulation and how this could be harnessed in the transplant setting.


Innate immune modulation in transplantation: mechanisms, challenges, and opportunities Introduction
The immune system consists, broadly, of two intricately interconnected arms: the innate and the adaptive.The innate arm is responsible for a rapid and broad response to invading pathogens, as well as to diverse physical and metabolic insults (1).This response involves the activation of a complex network of cellular and soluble mediators of inflammation that play a crucial role in maintaining immune surveillance at critical tissue sites.However, the failure to resolve the response can lead to persistent inflammation, tissue damage, and fibrosis (2,3).The innate immune system uses pattern recognition receptors (PRRs) to detect conserved pathogenic motifs, such as lipid and carbohydrate moieties.PRRs are expressed on various immune and non-immune cells and can sense pathogen-associated molecular patterns and danger signals.Toll-like receptors (TLRs) are the most extensively studied PRRs, found on the cell surface and endosomes of cells, and they recognize bacterial lipopolysaccharides and viral nucleic acids.Intracellular signaling pathways are initiated upon PRR activation, resulting in the mobilization of inflammatory and immune responses (4,5).The innate immune system's hard-wired responsiveness provides a critical first line of defense against invading pathogens and tissue damage, ensuring a rapid and effective response to potential threats.
In contrast, the adaptive arm is triggered by the presence of specific antigens, and it involves a highly targeted and specific response.This response is characterized by clonal expansion and affinity maturation of T and B cells, resulting in the generation of high-affinity receptors for antigenic peptides (6).The innate arm plays a vital role in initiating and enhancing the adaptive response by providing signals that trigger the recruitment and activation of cells of the adaptive immune system.The coordinated and integrated interplay between the innate and adaptive arms is essential for the acquisition of effective and long-lasting immunity against invading pathogens (7).The innate arm provides a rapid and non-specific response, while the adaptive arm provides a highly specific and targeted response.Therefore, the ability of the immune system to generate a successful defense against invading pathogens depends on the functional interdependence between these two arms of the immune system (8).
The innate pathway has been extensively studied in the context of transplantation.Understanding the role of these innate pathways in graft dysfunction and rejection is of paramount importance.By exploring the intricate interplay between the innate immune system and the transplantation process, we can identify novel therapeutic targets for promoting graft acceptance.
This review aims to provide a comprehensive analysis of the complement and TLR systems in the context of transplantation.We will examine the mechanisms underlying their activation, their contribution to graft dysfunction and rejection, and the potential for targeted manipulation of specific components to steer the immune response towards graft protection and acceptance.In doing so, we hope to deepen our understanding of the complex immune interactions that underlie innate immune responses in transplantation and to provide valuable insights into the development of new immunomodulatory strategies for improving transplant outcomes.

Components of the innate immune system and their role in transplant Complement
The complement system is a vital component of the innate immune system.It is triggered by pattern recognition molecules (PRMs) that recognize specific patterns on the surface of invasive pathogens or damaged cells.The classical and lectin complement activation pathways converge on C3, the pivotal component of complement, while the alternative pathway generates a C3 convertase from C3b and factor B (fB).The resulting cleavage of C3 generates complement effectors, including the membrane attack complex (MAC), C3a, and C5a, which recruit the adaptive immune system and mediate inflammation (Figure 1) (9, 10).
Allograft rejection is a complex process that involves both innate and adaptive immune responses, and complement activation has been widely implicated in the initiation and progression of allograft injury during ischemia reperfusion injury (IRI) and in the pathophysiology of rejection (10-12).Complement activation during IRI is triggered by locally released CL-11, which activates C3a/C3aR interaction leading to glomerular and tubular injury and stimulates secondary epithelial cell chemokine production that contributes to local inflammation (13,14).
The role of complement activation in acute rejection is more complex.The inflammatory response triggered by complement activation in a transplanted organ is mediated via generation of biologically active complement fragments.These fragments mediate chemotaxis and activate neutrophils and macrophages, resulting in a cascade of biological effects that depend on the site and trigger of complement activation (15).Once the MAC is formed, cells are induced to produce IL-1α and IL-8, further increasing local tissue inflammation (16).
Complement activation is detrimental in itself, as it damages the transplanted organ, but it also leads to recruitment of the adaptive immune system by enhancing T-cell activity.C5a has been shown to be a strong chemoattractant, and C5aR deficiency has demonstrated a protective effect in mouse models of IRI (17).Understanding the varying roles of complement within the transplant process is critical to predicting transplant outcomes and developing strategies to minimize the risk of allograft rejection.

Toll-like receptors
IRI is a multifaceted process that entails the activation of diverse pathways such as toll-like receptor (TLR) signaling, extracellular signaling molecules, alterations in gene expression, production of reactive oxygen species (ROS), regulation of cell death, activation of innate and adaptive immune factors, and the triggering of hypoxia-inducible factors (18).In general, this process constitutes a classic positive feedback loop in which damaged cells at the transplantation site release redox-sensitive damage-associated molecular patterns (DAMPs) that promote local accumulation of recipient-derived monocytes and polymorphonuclear leukocytes (PMNs), such as neutrophils (19).These cells sustain immune cascades and amplify the destruction of foreign tissue.This self-amplified cytotoxic cascade is initiated by the release of DNA/RNA complexes and/or acetylation of the non-histone chromatin-associated protein such as high mobility group box 1 (HMGB1), a representative danger signal.Additionally, degradation of the extracellular matrix can release molecules such as heparan sulfate, hyaluronan, fibrinogen, fibronectin A domain, or tenascin C, which can further amplify the response to tissue damage (20).
Acute kidney injury (AKI) is a complex disorder characterized by a rapid decline in renal function resulting from various etiologies.Neutrophils and macrophages are among the earliest effector cells involved in AKI, where they are found to persist for several days.The contribution of these cells to the pathophysiology of IRI results from their adherence to endothelium, release of ROS and proteases, and their ability to synthesize and secrete cytokines/chemokines that recruit other effector cells to the organ, thereby amplifying local inflammation (21)(22)(23).
Zhang et al. investigated the inactivation of neutrophils using mAbs against high mobility group box 1 (HMGB1) in a mouse model of AKI.They found that the treatment alleviated IRIinduced renal dysfunction by suppressing the activation of the HMGB1-Toll-like receptor 4 (TLR4)-interleukin (IL)-23-IL-17A signaling axis.This resulted in the reduction of neutrophilmediated inflammation, chemokine expression, oxidative stress, and apoptosis (24).Neutrophil trafficking is also dependent on the crosstalk between donor non-classical and classical monocytes in a lung transplant IRI model (25).
Depleting neutrophils entirely may not be clinically feasible, given their critical role in pathogen clearance and immune surveillance.Instead, selectively blocking the recruitment of circulating leukocytes to the inflammatory site may be a better approach.In a recent study, researchers used mAb treatment to block the vascular surface marker CD321, a marker for the transmigration of circulating leukocytes into the inflamed tissue.This blockade attenuated damage responses by hepatic IRI, evaluated by serum liver enzymes, inflammatory cytokines, and hepatocyte cell death (26).

The IL-6 pathway
As a critical regulator of inflammation, IL-6 plays a pivotal role in the pathophysiology of transplant rejection (27).It is noteworthy that upregulation of IL-6 during brain death promotes a proinflammatory state even before organ procurement (28,29), and during cold preservation, there is a detectable increase in graft IL-6 levels (30).This increase in IL-6 then upregulates adhesion molecules, inflammatory cytokines such as IL-17 and IFN-γ, and molecules that regulate migration in endothelial cells (31).Importantly, a growing body of evidence suggests that IL-6 blockade can mitigate the pro-inflammatory environment associated with brain death and IRI (32).Evidence from experimental and human studies links IL-6 to allograft injury, with blockade of the IL-6/IL-6R signaling pathway shown to mitigate these effects (33,34).In mouse skin transplantation models, IL-6 promotes T cell alloreactivity and impairs Treg function (35).In addition, inhibition of IL-6 signaling reduces the production of donor-specific antibodies and modulates immune regulatory and effector cells (36, 37).Studies in murine models demonstrate that IL-6 plays a critical role in the pathogenesis of acute and chronic allograft rejection, and blocking IL-6 reduces/prevents rejection and fibrosis (38-41).In human transplant recipients, higher levels of IL-6 are consistently associated with worse outcomes, including a higher risk for acute rejection (42-44).

Coagulation cascade
The coagulation system, through the extrinsic and intrinsic pathways, activates factor X. Initiated by vascular injury, the extrinsic pathway catalyzes thrombin production, vital for stimulating platelets and vascular cells.The intrinsic pathway is autoactivated when circulating factor XII contacts negatively charged surfaces, under the regulation of antithrombin, tissue factor pathway inhibitor, and activated protein C. Importantly, coagulation interacts closely with the innate immune system and the complement cascade, influencing thrombogenesis and fibrinolysis (45).
Thrombin and tissue factor interact with protease-activated receptors on immune cells, releasing inflammatory mediators crucial for transplantation and IRI.In delayed graft function (DGF), ischemia/reperfusion-triggered coagulation activation, fibrin deposition, and robust expression of pro-inflammatory agents primes adaptive alloimmune responses and leads to allograft dysfunction (46,47).
Mechanistically, this is explained by infiltrating dendritic cells expressing protease-activated receptor 1 (PAR-1), which influences cytokine gene expression and enhances T helper-1 bias in DGF (48).Interstitial coagulation activation, indicative of renal graft rejection, involves fibrin stimulation of fibrosis-contributing cells, and post-transplantation complications like microvascular thrombi, endothelial dysfunction, and fibrin deposition can lead to graft dysfunction, particularly in highly sensitized recipients (49,50).
The fibrinolytic system, responsible for removing fibrin, prevents vessel occlusion and is significant in transplantation (51).The cause of organ donors' death can affect fibrinolysis activation, with injured donors exhibiting enhanced fibrinolytic activity (52).DGF correlates with increased urokinase (uPA) and urokinase-type plasminogen activator receptor (uPAR) expression, which impacts graft function and renal filtration.Chronic renal graft failure is characterized by fibrin deposition due to a disrupted plasminogen/plasmin system inhibiting fibrinolysis (53, 54).

Therapeutic strategies
Many current therapies in transplantation target the adaptive immune system in order to dampen the immune response to the new organ.The adaptive immune system and innate immune system are intimately linked, and the various components of the innate immune system may serve as novel therapeutic targets (Figure 2).

Complement inhibition
There is increasing experimental and clinical evidence indicating that complement blockade can be an effective therapeutic target for improving outcomes in organ transplantation (55).A range of therapeutic agents, including monoclonal antibodies, small molecules, and small interfering RNA (siRNA) agents, have been developed to inhibit complement cascade activation.For example, the administration of a soluble complement receptor-1 antagonist (sCR1) can inactivate the C3 and C5 convertases, resulting in reduced neutrophil migration into grafts and reduced posttransplantation reperfusion edema in animal models of lung allotransplantation (56).In clinical trials, sCR1 has been shown to effectively inhibit complement activation, and treating recipient mice with a C5aR antagonist before transplantation has been shown to improve graft survival and reduce monocyte/macrophage infiltration (57,58).
Eculizumab, a humanized monoclonal antibody that binds to C5 and inhibits its cleavage to C5a, has been approved by the FDA to block complement cascade activation and constitutes another approach to attenuating IRI in organ transplantation (59, 60).Clinical case reports have shown promising results for eculizumab in patients with atypical hemolytic uremic syndrome undergoing kidney transplantation (61).Another potential strategy to inhibit complement cascade activation is to silence C3 using siRNA.In a mouse model of kidney IRI, the systemic administration of C3-specific siRNA reduced renal injury and mouse mortality by diminishing renal C3 synthesis, as C3 is the central component of complement cascade activation where all three pathways converge (62, 63).Silencing the C5a receptor (C5aR) gene using siRNA before the induction of ischemia resulted in reduced expression of the proinflammatory cytokine TNF-α and chemokines MIP-2 and KC, resulting in reductions in neutrophil influx and cell necrosis in the kidneys (64).
In a preclinical context, C1 inhibitors (C1 INH), which are approved for hereditary angioedema treatment, can prevent the activation of both the classical and lectin pathways (65).Research using a brain-dead rat model has shown that C1 INH can mitigate complement activation resulting from brain death (66).Our preliminary studies using a mouse model of renal IRI showed that pre-treating animals with C1 INH led to enhanced renal function, improved survival rates, and decreased C5a release, C3b deposition, as well as neutrophil/macrophage infiltration in the grafts.Furthermore, we observed a significant reduction in tissue fibrosis and TGF-β1 levels 30 and 90 days post-ischemia (67).Utilizing a nonhuman primate kidney transplant model, we demonstrated that managing brain-dead donors with recombinant human C1 INH and heparin substantially diminished complement pathway activation, attenuated systemic inflammatory response, and crucially, averted DGF in all instances compared to controls (68).A subsequent study suggested that C1 INH, when administered systemically to either the brain-dead donor or recipient only, could decrease complement deposition on biopsy and prevent DGF (69).Lastly, in a pig kidney auto-transplant model, C1 INH treatment expedited the recovery of glomerular function and significantly lessened long-term graft fibrosis (70,71).
Clinical trials pertaining to kidney transplantation have thus far explored the use of C1 and C5 inhibitors as well as a C3 inhibitor.The primary objective of these trials was to avert early graft failure due to DGF, and to prevent both early and late rejection episodes, particularly antibody-mediated rejection (ABMR).Currently, only one phase I/II trial (NCT02134314) comparing the use of the C1esterase with a placebo has been completed.Although there were no significant differences between groups in the incidence of DGF, the duration of dialysis was notably shortened in the C1-esterase inhibitor group (72).Furthermore, in a follow-up study conducted 3.5 years post-transplantation, a significantly improved estimated glomerular filtration rate (eGFR) was found in the C1-esterase inhibitor group (73).
Recent alternative anti-complement therapies have shown promise by directly administering complement inhibitors to donor kidneys ex vivo before transplantation.This method focuses on local kidney complement inhibition, minimizing the impact on the recipient's systemic complement pool.Mirococept, derived from human CR1, was tested in the EMPIRIKAL trial to evaluate its effect on DGF duration.The initial trial demonstrated the feasibility and safety of ex vivo Mirococept delivery to kidney allografts and highlighted the need for dose calibration (74).Subsequent pig kidney studies showed strong localization to the treated kidney's tubular epithelium and capillaries, with minimal recipient release.The EMPIRIKAL trial is now revising its human study protocol based on the optimal higher dose achieved in pigs (EMPIRIKAL-2).

TLR inhibition
Inhibitors and antagonists that target TLR signaling or downstream components have the potential to reduce IRI and improve allograft survival, and several studies on animal models suggest that blocking antibodies against TLR pathway molecules can be an effective therapeutic strategy.For example, the administration of an anti-HMGB1 neutralizing antibody reduced kidney IRI in mice, as shown by lower levels of inflammation and apoptosis in tubular epithelial cells, and reduced infiltration of neutrophils and macrophages (75).In addition, in a model of cardiac transplantation, neutralizing HMGB1 resulted in a reduction of inflammatory CD11b + Ly6C high myeloid cells both at the level of the allograft and the spleen (76).
There have been numerous attempts to block the TLR pathway as a strategy to attenuate ischemia-reperfusion injury (IRI) and enhance allograft survival.One such attempt involves the use of eritoran, a TLR4 antagonist that has been tested in transplantation (77).Eritoran is a synthetic structural analog of the lipid A portion of lipopolysaccharide (LPS) that binds to the MD2-TLR4 receptor and terminates MD2/TLR4-mediated signaling.This action blocks LPS from binding at the cell surface and subsequently inhibits the pro-inflammatory signaling cascade that follows (78).In a rat transplantation model, eritoran treatment induced less monocyte infiltration; lower levels of TNF-α, IL-1β, IL-6, and MCP1; and prolonged survival.There is also evidence to suggest that eritoran may block the HMGB1-TLR4 interaction, which could explain its ability to attenuate IRI.Tissue injury and ischemia activates the innate immune system, which is comprised of intricately interconnected pathways that influence the activation of the adaptive immune system, which can ultimately lead to graft dysfunction, alloreactivity, and rejection in organ transplantation.Molecules within each of these pathways may be targeted to suppress those reactions and promote transplant organ function and acceptance by the host immune system.In a mouse model of liver IRI, eritoran was shown to inhibit the TLR4-dependent release of HMGB1, indicating a potential mechanism by which eritoran exerts its protective effects (79).
Another TLR-4 inhibitor, TAK-242, is a small compound that mitigates the release of inflammatory cytokines triggered by pathogens, primarily by impeding TLR-4-mediated signaling pathways.Furthermore, it exhibits suppressive effects on the synthesis of nitric oxide (NO) or tumor necrosis factor (TNF)alpha, specifically when induced by the TLR4-specific ligand LPS (80).A study investigated the effects of TAK-242 on liver transplant viability in a model of swine Maastricht-category-III cardiac death.TAK-242 was administered prior to induction of cardiac death with mRNA and protein levels of TLR4 signaling pathway evaluated at different time points after induction of cardiac death.The results showed that the mRNA and protein levels of important immune and inflammatory response molecules such as TLR4, NF-ϰB, MCP-1, TNF-α, IL-6, ICAM-1, and MPO, increased significantly after cardiac death.However, infusion of TAK-242 one hour before induction blocked the increase of many of these molecules; though notably, the increase of TLR4 levels was not affected by the TAK-242 infusion (81).

Other potential targets in the innate immune system IL-6 inhibitors
Tocilizumab, a potent antagonist of the interleukin-6 receptor, has shown remarkable efficacy in reducing HLA antibodies in highly sensitized patients awaiting kidney transplantation and those with antibody-mediated rejection.In 2015, Vo et al. conducted a phase I/II open-label trial (NCT01594424) with ten unresponsive end-stage renal disease patients.The six-month treatment regimen with monthly tocilizumab and IVIg administration resulted in a reduction of DSA levels, enabling five patients to undergo kidney transplantation.Biopsies revealed no evidence of antibody-mediated rejection (AMR) six months post-transplant, but two patients who stopped tocilizumab experienced mild AMR a year later.The study suggested that targeting the IL-6 pathway could offer an alternative for desensitization (82).
A subsequent study by this group evaluated the use of tocilizumab as a rescue therapy for DSA-positive chronic antibody-mediated rejection (cAMR) and transplant glomerulopathy in patients who had previously failed standard treatments.Monthly administration of tocilizumab yielded an 80% graft survival rate at six years, decreased DSA levels, and stabilized renal function without significant adverse events.Four patients did experience graft loss after discontinuing tocilizumab, suggesting rebound IL-6 pathway signaling may contribute to generation of an alloimmune response (83).
Furthermore, tocilizumab has been studied as a first-line therapy for cAMR in kidney transplantation, resulting in GFR stabilization and a significant decrease in DSA with improved pathology observed at six months.IL-6 blockade may also be a potential treatment option for regulating T-cell alloimmune response in kidney transplantation.A recent randomizedcontrolled trial found that kidney transplant recipients who received tocilizumab had increased T-reg frequency and a blunted T-effector cytokine response (84).A multicenter phase II clinical trial investigating the efficacy of tocilizumab in cardiac transplantation is currently under way to evaluate the impact of tocilizumab on acute and chronic cardiac allograft rejection (NCT03644667).

TNF and IL-1 receptor antagonists
The use of anti-inflammatory agents to counteract the harmful effects of proinflammatory cytokines has been extensively investigated, primarily in islet transplantation.Anti-cytokine strategies, such as TNF-alpha inhibition and IL-1 beta antagonism, have demonstrated promising results in improving islet cell function and promoting insulin-independence following transplantation.
TNF-a inhibitors have shown reduced inflammatory markers in animal models and long-lasting efficacy in treating autoimmune disorders (85, 86).The pharmacological characteristics of TNF inhibitors can vary considerably, with Etanercept (ETA) binding and neutralizing lymphotoxin-α3 and lymphotoxin-α2β1, while Infliximab (INF) does not possess these properties (87,88).A recent qualitative analysis of antiinflammatory agents in clinical islet transplantation summarized findings from multiple studies evaluating Etanercept monotherapy in allo-islet cell transplantation protocols (89).Although the studies differed in insulin independence (II) definitions and assessment time points, the overall results suggest that Etanercept may contribute to long-term maintenance of II.Despite some inconsistencies in outcomes, an Etanercept-based protocol could be associated with lower islet equivalent (IEQ) requirements (90, 91).
A single prospective, randomized study examined Infliximab as an adjunct to the Edmonton protocol, but no significant clinical benefits or substantial differences were observed in oral glucose tolerance test (OGTT) or intravenous glucose tolerance test (IVGTT) outcomes (92).
IL-1β, a pro-inflammatory cytokine, has been linked to β-cell destruction and diabetes development (93).Interleukin-1 receptor antagonists (IL-1RA) have been demonstrated to effectively counteract the effects of proinflammatory cytokines, including nitric oxide production, necrosis, apoptosis, glucosestimulated insulin secretion, and mitochondrial dysfunction (94).Several studies have shown that interleukin-1 receptor antagonist (IL-1RA) can improve pancreatic islet transplantation outcomes in mice, primates, and humans (95, 96).In two human trials, anakinra combined with other immunosuppressive agents during the first 1-2 weeks post-transplantation resulted in better engraftment without increased infection risk (97,98).
Limited data exist on IL-1 beta-directed therapies in transplantation outside the islet field.In renal transplantation, the safety and efficacy of combining IL-1 beta-directed therapy with standard immunosuppressive treatment were evaluated in three patients, with no significant complications except for minor infections (99).Case reports and series indicate that anakinra can be safely used alongside immunosuppressive agents in renal transplant recipients with IL-1 driven diseases, although one patient experienced neutropenia and decreased allograft function, with an unclear link to IL-1 blockade (100).

Coagulation cascade inhibition
The coagulation system, which regulates bleeding and clotting, also plays an important role in innate immunity; this, and its interaction with the fibrinolytic system influences IRI.Targeting coagulation factors is complicated by their presence within fibrin clots, often protecting them from inhibitors (45).Therapies such as rapamycin can mitigate fibrinolysis disruption by reducing plasminogen activator inhibitor type 1 (PAI-1) expression, slowing the progression of interstitial fibrosis (101).Furthermore, in porcine models of renal autotransplant, the use of Xa and dual Xa/IIa inhibitors has been shown to be beneficial to graft outcome (102, 103).In addition to inhibiting the complement system, C1 INH has also been shown to inhibit enzymes in the coagulation and fibrinolytic systems, including factor XII and plasmin, respectively (104).As discussed above, in pre-clinical models and ongoing clinical studies, C1 INH has demonstrated a beneficial effect on graft function.

Glucocorticoids
(GCs) play a crucial role in immunosuppression following renal transplantation by mitigating inflammatory responses and leukocyte infiltration.They also facilitate the resolution of inflammation through the suppression of vascular permeability, leukocyte distribution/trafficking, and modulation of cellular death/survival and differentiation processes (105, 106).While GCs were initially believed to exert their anti-inflammatory effects through the inhibition of proinflammatory cytokine-encoding gene regulators, recent studies have uncovered alternative mechanisms, including negative interference with inflammatory mediator synthesis, suppression of immune cell activation, and cooperation between the glucocorticoid receptor and transcription factors to induce antiinflammatory genes (107).
GCs are known to induce neutrophilic leukocytosis by promoting neutrophil maturation and mobilization, an effect that can be blocked by L-selectin adhesion protein inhibition.They also hinder the entire neutrophil activation process, including respiratory burst-related enzyme expression, chemotaxis, phagocytosis, and cytokine secretion.In neutrophils, GCs suppress transcription factors related to pro-and antiinflammatory genes, leading to upregulated expression of interleukin and pro-inflammatory leukotriene receptors and reduced apoptosis sensitivity, thus extending neutrophil lifespan.Furthermore, endogenous glucocorticoids under stress conditions affect natural killer (NK) cells by reducing their cytolytic activity and inducing pro-inflammatory cytokine synthesis through an epigenetic mechanism, enhancing the expression of IL-6 and INF-γ and promoting histone acetylation in their enhancer regions.
GCs also exhibit direct effects on innate immune cells, as demonstrated by in vitro studies on methylprednisolone-treated monocytes, which show increased anti-inflammatory cytokine expression and reduced antigen-presenting capabilities (108, 109).In vivo data from renal transplant recipients treated with methylprednisolone reveal alterations in monocyte populations and TLR4 downregulation on GC-treated monocytes, a critical component in monocyte activation during sepsis and the immune response to transplanted organs (110,111).Steroids also impact dendritic cell differentiation and maturation, with dexamethasone-exposed monocyte-derived dendritic cells displaying decreased expression of specific markers, impaired antigen-presenting cell function, and reduced cytokine secretion (112).
The anti-inflammatory effect of steroids in transplantation has been widely investigated in the deceased donor setting and initial studies have not consistently show benefits of donor methylprednisolone treatment in transplantation outcome (113).In addition, a large-scale, randomized, double-blind trials in kidney transplantation found that while methylprednisolone pretreatment improved inflammatory and apoptotic gene expression profiles, it did not impact delayed graft function or serum creatinine decline (114,115).Therefore, high-dose methylprednisolone before organ recovery is not recommended for kidney transplantation.
Although the trial's results were inconclusive, preliminary evidence suggests potential benefits of donor steroid treatment in liver transplantation, prompting continued research efforts into the routine use of steroids in combined hormonal resuscitation protocols (116).

Myeloid differentiation factor 88 (MyD88)
Myeloid differentiation factor 88 (MyD88) is a protein that plays a role in signal transduction through TLRs, prompting the production of pro-inflammatory cytokines that promote leukocyte recruitment.Specifically, MyD88 links the IL-1 receptor or other TLR family members to IL-1R-associated kinase (IRAK) kinases that leads to downstream events including NFkB activation and MAP kinase activation (117).Thus, it has a far-reaching effect on the inflammatory response of the immune system.
TJ-M2010-5 is a MyD88 inhibitor that targets and interacts with the MyD88 TIR domain, interfering with dimerization.This molecule has been shown in murine models to play a role in slowing liver fibrosis, and decreasing myocardial ischemic reperfusion injury (118), cerebral reperfusion injury (119), and hepatic reperfusion injury (120).Alleviation of hepatic reperfusion injury with this molecule has been particularly successful when coupled with hepatic hypothermic oxygenated perfusion (121).While these studies have been performed using animal models, they demonstrate some significant promise as a viable therapeutic option for the prevention or reduction of ischemia reperfusion injury in human transplantation.

Resveratrol
Resveratrol is a naturally-occurring phenol produced by plants that has been shown to play a role in inflammatory disease prevention and progression.Resveratrol is thought to be produced in response to oxidative stress, and several studies have shown it is able to regulate hepatic metabolism of lipids hence improving metabolic lipid homeostasis and slowing the progression of fatty liver disease (122).Resveratrol also functions as an anti-oxidant, and its main effect is linked to the reduction of reactive nitrogen species, direct elimination of free radicals, improvement in antioxidant enzymatic activity and promotion of synthesis of antioxidant.
Resveratrol interacts with several different immunomodulatory targets including Sirtuins.Sirtuins are NAD+-dependent histone deacetylases, and they are thought play a significant role in immunomodulatory responses and are of significant interest as targets to treat autoimmune disease (123).They are currently studied for their role in promoting organ transplant tolerance, in particular SIRT1 inhibition, which may affect Th17/Treg cell balance (124,125).
Overall Resveratrol has the potential to play a crucial role in prevention and treatment of liver disease such as anti-oxidant, steatosis and immune-modulator.Additional studies are ongoing with the goal of highlighting efficacy and safety.

Vitamin D
Vitamin D plays an important role in the regulation of the immune system, and its deficiency has been shown to play a role in immune dysregulation, particularly in the context of autoimmunity (126).Renal failure can lead to disruption of vitamin D metabolism.In fact, studies have shown that vitamin D deficiency may be a risk factor for rejection or decreased graft function (127)(128)(129).As such, studies have demonstrated that vitamin D supplementation may serve as a means to promote immunity and maintain graft function (130).

Conclusion
In order for the immune system to function appropriately, there must be a complex synergy between the innate and adaptive arms.The innate immune system is essential for the highly specialized adaptive immune system to function.Furthermore, modulation of this complex system is essential for successful organ transplantation, particularly to prevent ischemia reperfusion injury and rejection, and to promote graft function.
There is an abundance of research that highlights components of the innate immune system, their role in transplantation, and how those components may be harnessed for therapeutic benefit.
Ongoing studies are identifying novel therapeutic targets within the innate immune system that have the potential to greatly impact management of transplant patients.