The IRAK family is made up of 4 members—IRAK1, IRAK2, IRAK3 (also known as IRAK-M), and IRAK4 [20]. All members share similar functional and structural domains, including a N-terminal death domain (DD), a proline/serine/threonine (ProST) domain, and a kinase or pseudokinease domain (KD or PKD) [21]. Except for IRAK4, the rest of the IRAK members contain an additional C-terminal domain (CD), which is required for activation of tumor necrosis factor receptor-associated factor 6 (TRAF6) [21]. DD plays an important role in TLR and IL-1R signaling by interacting with the myeloid differentiation primary response protein 88 (MyD88). Whereas the ProST domain, which contains 2 peptide sequences rich in proline (P), glutamic acid (E), serine (S), and threonine (T) (the so-called PEST sequence), is responsible for hyperphosphorylation and degradation of IRAK1 [22, 23].

IRAKs were first functionally described as key mediators in coordinating multiple IL-1 signaling pathways and in facilitating production of pro-inflammatory cytokines, but later, IRAKs were also found to be implicated in signal transduction through TLRs [21]. Each IRAK family member exerts a different role in modulating TLRs-/IL-1R-associated downstream responses. Following activation of TLRs-/IL-1R, MyD88 and IRAK proteins are recruited to form the receptor complex [22, 23], in which IRAK4, as an upstream kinase, activates IRAK1 and IRAK2 through phosphorylation. Phosphorylated IRAK1 and 2 can cause some common or different functional effects. In the early phase, both IRAK1 and IRAK2 have the same function, which is associated with an acute inflammatory response; while in the late phase, IRAK2 is believed to be involved in chronic inflammatory responses [24].

Activation of IRAKs (1, 2, and 4) by TLRs/IL-1R leads to stimulation of various downstream signaling pathways, especially the nuclear factor kappa B (NF-$\kappa$B)/mitogen-activated protein kinase (MAPK) pathway (NF-$\kappa$B/MAPK) [25], the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway (JAK/STAT) [26], and the NLR protein 3 (NLRP3)/Caspase-4/-5/-11 pathway (NLRP3/Caspase-4/-5/-11) [18], which eventually stimulates production and secretion of pro-inflammatory cytokines (Fig. 1). In contrast, IRAK3 negatively modulates TLRs/IL-1R (or TIR) domain signaling by inhibiting the IRAK1-2-4 complex, reducing production and secretion of inflammatory cytokines, eventually resulting in immunosuppression [27, 28].

Fig. 1.

IRAK1-mediated signaling pathways in cardiovascular and immune cells. Injured CMs, ECs, and VSMCs release DAMPs, which then bind to TLRs/IL-1R on immune cells and cardiovascular cells (CMs, ECs, and VSMCs) and activate downstream inflammatory pathways including: (a) IRAK1 $>$ NF-$\kappa$B and MAPK; (b) IRAK1 $>$ JAK and STAT; and (c) IRAK1 $>$ NLRP3 $>$ Caspase 4/5/11, presumably leading to foam cell formation and cardiac cell apoptosis. The schematic cell refers to an immune (e.g., macrophage) cell or a cardiovascular cell (e.g., CM, ECC, and VSCMC). Abbreviations: DAMPs, damage associated molecular patterns; HMGB1, high mobility group box 1; HSP 60/70/72, heat shock protein 60, 70, 72; I/R, ischemia/reperfusion; IL-1/-6/-10/-18/-27, interleukin-6,-10,-18, -27; IL-1R, interleukin-1 receptor; IRAK1/4, interleukin receptor associate kinase 1/4; MAPK, mitogen-activated protein kinase; MyD88, myeloid differentiation primary response 88; NF-$\kappa$B, nuclear factor kappa-light-chain-enhancer of activated B cells; NLRP3, nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3; STAT, signal transducers and activators of transcription; TLRs, toll-like-receptors; TRAF6, tumor necrosis factor receptor-associated factor 6 (The figure was originally created by the authors using BioRender.com online software).

IRAK1, the first-discovered and most-studied member of the IRAK family, is known to play an important role in mediating IL-1-associated immune and inflammatory responses by activating NF-$\kappa$B pathway [29]. It was found that human IRAK1 is ubiquitously expressed in almost every organ, whereas mouse IRAK1 appears to be manifested mainly in the liver, kidneys, and testis [29].

IRAK1 comprises 3 splice variants (1b, 1c, and 1s) as well as a full-length form and each variant has its unique biological function. For example, IRAK1b remains active and extremely stable after being activated by IL-1 signaling, thus, leading to the sustainable activity of NF-$\kappa$B [30]. IRAK1c, which is functionally similar to IRAK3, is a dominant-negative regulator in TIR domain-mediated immune and inflammatory responses [31]. IRAK1c is dominantly expressed in human brain and may be mainly involved in neuronal responses to infection and tissue damage [30, 32]. Lastly, IRAK1s appears to be expressed only in murine mammals as an inactive kinase. Although its biological function remains unclear, overexpression of IRAK1s stimulates the downstream NF-$\kappa$B and c-Jun N-terminal kinase (JNK) pathway via binding to endogenous IRAK1 [33]. Nevertheless, existence of IRAK1 splice variants with different biological functions and capacities demonstrate the diverse impacts of the IRAK1 subfamily on innate immune and inflammatory responses. The different expression of IRAK1 variants between mice and humans may signify their common and also special requirements for immune and inflammatory responses between the two species [34], which requires further investigation.

The involvement of TIR domain-associated IRAK1 $>$ JAK $>$ STAT and IRAK1 $>$ NLRP3 $>$ Caspase signaling in cardiac cells is largely unknown and published studies using cardiovascular cells are very limited. Although most of the published data in this area is in non-cardiac cells, such as macrophages, lymphocytes, and microglial cells (Fig. 1) [35, 36], the findings are likely similar in cardiovascular cells. Huang et al. [37] reported that IRAK1 is essential for STAT3 activation and subsequent expression of interleukin-6 and 10 (IL-6, IL-10) because IRAK1-deficient cells showed impaired IL-10 production. It is possible that IRAK1 directly binds to the IL-10 promoter upon lipopolysaccharide (LPS) challenge or modulates STAT3 activity via TRAF6, increasing production of inflammatory cytokines [38]. IRAK1 also participates in STAT1 activation in IL-1R signaling [38]. In IRAK1-deficient macrophages, higher levels of STAT1/2 activities and IL27 production increase following IFN-$\beta$ stimulation [39]. Since IL-10 and IL-27 are anti-inflammatory cytokines, IRAK1-mediated IL-1R to STAT signaling may play a role in the late stage tissue repair and remodeling, while IRAK1-mediated production of pro-inflammatory cytokines in NF-$\kappa$B/MAPK signaling may engage in the early stage of infection and inflammation [35, 36].

NLRs are the only cytoplasmic receptors that recognize pathogen-derived intracellular invaders (i.e., PAMPs) and non-infectious danger signals (i.e., DAMPs from injured cardiac cells [40]) among the four members of PRR [TLRs, NLRs, C-type lectin receptors (CLRs), and RIG-1 like receptors (RLR)] [41, 42] (also see Fig. 1). Activation of NLRs by TIR domain signaling promotes the formation of inflammasomes, which then activate Caspase-1/4/5/11, leading to the production of the pro-inflammatory cytokines IL-1$\beta$, and IL-18 [40, 42]. In this signaling response, NLRP3 appears to be the most important factor and involves a wide range of inflammation and autoimmune-induced disorders in patients [40]. Emerging evidence indicates that IRAK1 plays an essential role in the rapid activation of NLRP3, thus, promoting TIR domain-associated immune responses [41].

The first known role of IRAK1 is the mediation of signal transduction from TLRs to inflammatory cytokines via the NF-$\kappa$B and MAPK pathways (Fig. 1). TLRs belongs to the PRR family and are Type I single-spanning glycoprotein [24]. TLRs are mainly expressed in immune cells [e.g., macrophages, mast cells, dendritic cells, nature killer (NK) cells, T cells, and B cells], but they were also identified in cardiomyocytes, epithelial cells, ECs, and vascular smooth muscle cells (VSMCs) [43]. As to cellular distribution, TLR1-2, TLR4-6, and TLR11 are found on the cell surface while TLR3 and TLR7-9 are located on the membranes of intracellular organelles, such as endosomes and lysosomes [44]. In the heart of mouse and human, TLR4 is probably the most abundant TLR among all 13 TLR family members [45] and it is the only LPS receptor that is associated with an immediate inflammatory response in heart following myocardial ischemia and sepsis [3], implying its unique biological role in modulating cardiac responses to both infection- and non-infection-induced injury.

TLRs can be recognized and activated by two types of ligands, PAMPs and DAMPs as mentioned above. PAMPs refer to infectious pathogens, such as PS, phospholipids, extracellular matrix, peptide, and nucleic acids released from bacteria; while DAMPs denote non-infectious endogenous molecules, such as heat shock proteins 60/70/72 (HSP60, 70, and 72) and high mobility group box 1 (HMGB1) protein released from, for example, injured cardiomyocytes [3, 46]. Studies indicate that all TLRs contain an extracellular domain with leucine-rich repeat (LRR) motifs specific for recognizing TLR ligands, whereas all IL-1Rs contain three immunoglobulin-like domains (Ig-like domains) specific for recognizing IL-1 ligands [24]. Since TLRs and IL-1Rs share the same homologous TIR domain, IRAK1 could participate in both TLR- and IL-1R-mediated signaling, suggesting its biological importance [24].

Activation of TLRs/IL-1R causes a series of intracellular reactions in the downstream signaling cascades, including recruitment of the TIR domain adaptors such as MyD88 [47], formation of the IRAK1/4 and MyD88 complexes, IRAK1 phosphorylation by IRAK4, detachment of hyper-phosphorylated IRAK1 [17], activation of TRAF6 [48]/transforming growth factor-$\beta$-activated kinase 1 (TAK1), phosphorylation/degradation of inhibitory KappaB (I kappa B or I$\kappa$B) protein by I$\kappa$B kinases (IKKs), and the nuclear translocation of NF-$\kappa$B. The final consequence of the above steps will ultimately lead to transcription and production of pro-inflammatory cytokines such as IL-1$\beta$, IL-6, and tumor necrosis factor $\alpha$ (TNF$\alpha$) [48]. Alongside, activation of TLRs/IL-1R also stimulates the MAPK signaling via TAK1 pathway [49]. TAK1 then leads to the phosphorylation of JNK, p38, and, ultimately, cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB), ending up with the activation of activator protein 1 (AP-1). The AP-1 transcription factor thereafter elicits transcription and production of pro-inflammatory cytokines similar to NF-$\kappa$B [50] (also see Fig. 1).

Atherosclerosis (often denoted as arteriosclerosis), which is characterized by lipid-laden foam cell accumulation in the arterial wall, is recognized as a chronic inflammatory disease [51]. As stated above, activation of IRAK1 enhances production of IL-10 [37] and patients with atherosclerosis often have an elevated level of IL-10, implying that the IRAK1-mediated innate immune response may play an important role in the development of atherosclerosis [37]. Numerous in vitro and in vivo studies listed below support these findings.

By genotyping 4 loci on the IRAK1 gene on the X chromosome of 996 Caucasian patients with Type 2 diabetes (467 men and 529 women), the Diabetes Heart Study identified two major haplotypes, CTTT (82%) and TCCG (13%), in the IRAK1 gene. The TCCG haplotype is significantly correlated with an enhanced blood level of C-reactive protein (CRP, an acute inflammation marker) in women but not in men [52]. Since blood concentration of CRP has been used to assess the risk of coronary artery disease and predict MI and stroke [53], the presence or overexpression of the TCCG haplotype in the IRAK1 gene may promote pathogenesis of CVDs [52].

On the other hand, the ATP-binding cassette subfamily A member 1 (ABCA1), a crucial mediator of lipid efflux in the cell membrane, can be downregulated by IRAK1 through TLR4 signaling pathway following treatment of oxidized low-density lipoprotein (oxLDL) [54], suggesting IRAK1’s potential role in atherosclerotic development. This hypothesis is supported by a similar study, in which IRAK1 was found to increase lipid binding, uptake, and cholesterol efflux in foam cells [55] (also see Fig. 2). However, inhibition of IRAK1 (by an IRAK1 antagonist, such as IRAK1/4 inhibitor or its siRNA) significantly attenuated expression of cluster differentiation 36 (CD36) [55]. It is known that CD36 is a crucial macrophage scavenger receptor that is responsible for cellular cholesterol accumulation, oxLDL internalization, and foam cell formation [55]. Downregulation of CD36 may lead to beneficial effects in preventing or weakening the development of atherosclerosis. Furthermore, inhibition of IRAK1 also increases expression of ABCA1 and ATP-binding cassette subfamily G member 1 (ABCG1), which leads to increased cholesterol efflux from macrophages [55]. This process is accompanied by increased expression and activity of liver transcriptional X receptor alpha (LXR$\alpha$) and nuclear factor of activated T cell (NFATc2). LXR$\alpha$ serves as a cholesterol sensor and regulates ABCA1 and ABCG1 through NFATc2 [55]. IRAK1 plays a critical role in maintaining NFAT in a phosphorylated inactive state [56]. Suppression of IRAK1 in macrophages enhances binding of NFATc2 to the ABCA1 promoter and results in an increase of ABCA1 expression and cholesterol efflux [57].

Fig. 2.

Working hypothesis of IRAK1-induced formation of atheromatous foam cells in the vascular wall. It is known that foam cells are formed from monocyte-derived M${}_2$ macrophages (majority), synthetic VSMCs, or injured ECs. Once oxLDL binds to TLRs and CD36 receptors on macrophages, ECs, and VSMCs, it will activate the (a) IRAK1 $>$ STAT1; (b) IRAK1 $>$ NF-$\kappa$B inflammatory; and (c) IRAK1 $>$ NFATc2 and LXR$\alpha$ signaling pathways; together, they promote secretion of pro-inflammatory cytokines and cholesterol efflux and facilitate the formation of foam cells. Abbreviations: ABCA1 (ABCG1), ATP-binding cassette subfamily A (G) member 1; CD36, cluster of differentiation 36; IRAK1, interleukin receptor associate kinase 1; LXR$\alpha$, liver transcriptional X receptor alpha; NF-$\kappa$B, nuclear factor kappa-light-chain-enhancer of activated B cells; NFATc2, nuclear factor of activated T cell 2; oxLDL, oxidized low-density lipoprotein; STAT1, signal transducers and activators of transcription 1; TLRs, toll-like-receptors (The figure was originally created by the authors using BioRender.com online software).

IRAK1 also stimulates VSMC proliferation, which is a pivotal pathogenic process in the development of atherosclerosis [58]. Two signaling pathways are likely involved in IRAK1-associated VSMC proliferation: (a) the IL-1/IL-18 inflammatory pathway, in which activation of IRAK1 induces production of inflammatory cytokines such as IL-1 and IL-18 which, in turn, provokes VSMC proliferation during atherosclerosis progression [59]; and (b) the kinase pathway, in which IRAK1 facilitates VSMC proliferation and neointimal hyperplasia by activating protein kinase C (PKC) and extracellular signal regulated kinase (ERK) [60].

ECs, a major cell type involved in plaque formation in atherosclerosis, is also found to be regulated by IRAK1 [61]. Alfaidi et al. [62] revealed that a signaling adaptor, named non-catalytic region of tyrosine kinase 1 (Nck1), could interact with IRAK1 under shear stress and trigger NF-$\kappa$B-based secretion of proinflammatory cytokines. IRAK1 knockdown in ECs inhibits expression of NF-$\kappa$B and downstream endothelial adhesion molecules, such as vascular cell adhesion molecule-1 (VCAM-1) and intercellular cell adhesion molecule-1 (ICAM-1). These molecules are crucial factors for endothelial activation [63]. Another study found that carotid ligation (disturbing shear stress) in mice increases phosphorylation of IRAK1 and expression of Nck1 in ECs, supporting the human findings in which both Nck1 and IRAK1 expression levels are increased in ECs isolated from human carotid artery plaques [62]. Correlations among constitutive expression of IRAK1 in the peripheral blood mononuclear cells isolated from patients with atherosclerosis, elevated levels of the plasma IL-10, and the stimulation effect of IRAK1 on IL-1 [37, 64], imply direct involvement of IRAK1 in atherosclerotic development. Collectively, these data establish a link between disturbed blood flow and IRAK1-mediated inflammation, highlighting a unique role for IRAK1 as a specific Nck1 binding protein in mediating endothelial activation under atheroprone hemodynamics [62].

Growing evidence demonstrates that IRAK1 is also involved in the development of cardiac diseases, such as MI and HF (Table 1, Ref. [65, 66, 67, 68, 69]). Thomas et al. [65] reported that IRAK1 mediates LPS-induced myocardial dysfunction of contractile through TIR domain signaling and knockout of the Irak1 gene significantly reduces mortality of mice with HF. Because atherosclerotic-induced MI and bacterial-induced cardiac septic shock share common characteristics of myocardial dysfunction, pharmacological inhibition of IRAK1 may reduce cardiac inflammation and provide beneficial effects for patients with heart disease [3]. Similarly, studies using a mouse ischemia/reperfusion model found that TLR4 $>$ IRAK1 signaling is activated and HSP60 expression is increased in injured cardiomyocytes. Although the underlying mechanism is not well-known, it is probably related to activated signaling from HSP60 (released from injured cardiomyocytes) $>$ TLR2/4 $>$ IRAK1 $>$ apoptosis in cardiomyocytes [66, 70].

Recent studies suggest that small non-coding RNAs [e.g., microRNA (miRNA), small nuclear RNA (snRNA), small nucleolar RNA (snoRNA), and Piwi-interacting RNA (piRNA)] are also involved in pathogenesis through modulation of IRAK1 [71]. Among ~2000 miRNAs, miRNA-146a was first identified as a negative regulator targeting IRAK1 and TRAF6 and reduced the TLR-triggered innate immune response [72]. This finding was supported by a gain-of-function study using the same model of mouse MI. In this study, increased expression of miRNA-146a decreased cardiac infarct size (~50%) and increased cardiac function, which was believed to be due to direct inhibition of IRAK1 and TRAF6 by miRNA-146a [68]. Another study also found that miRNA-146a diminishes sepsis-induced cardiomyocyte apoptosis and infiltration of inflammatory monocyte cells by inhibiting the same molecule (i.e., IRAK1 and TRAF6) [67]. New studies found that IRAK1 can also be regulated by other miRNA, such as miRNA-142-3p. Su et al. [69] discovered that there is significant downregulation of miRNA-142-3p in porcine MI induced by coronary micro-embolization. Either up-regulation of miRNA-142-3p or down-regulation of IRAK1 appears to be able to reduce production of NF-$\kappa$B and pro-inflammation cytokines (IL-1$\beta$, IL-6) in cardiomyocytes [69], which could become potential pharmaceutical targets to treat patients with MI.