Small Leucine-Rich Proteoglycans in Renal Inflammation: Two Sides of the Coin

Abstract

It is now well-established that members of the small leucine-rich proteoglycan (SLRP) family act in their soluble form, released proteolytically from the extracellular matrix (ECM), as danger-associated molecular patterns (DAMPs). By interacting with Toll-like receptors (TLRs) and the inflammasome, the two SLRPs, biglycan and decorin, autonomously trigger sterile inflammation. Recent data indicate that these SLRPs, besides their conventional role as pro-inflammatory DAMPs, additionally trigger anti-inflammatory signaling pathways to tightly control inflammation. This is brought about by selective employment of TLRs, their co-receptors, various adaptor molecules, and through crosstalk between SLRP-, reactive oxygen species (ROS)-, and sphingolipid-signaling. In this review, the complexity of SLRP signaling in immune and kidney resident cells and its relevance for renal inflammation is discussed. We propose that the dichotomy in SLRP signaling (pro- and anti-inflammatory) allows for fine-tuning the inflammatory response, which is decisive for the outcome of inflammatory kidney diseases.

Introduction

Chronic inflammation and fibrosis are common features of various kidney diseases such as chronic glomerulo- and interstitial nephritis, diabetic nephropathy (DN), and crystal nephropathies.¹ Numerous triggers of renal inflammation have been described so far ranging from infectious agents to sterile inducers such as toxins, ischemia, trauma, or extra- and intracellular-derived components.^2–4 Over the last decades, extensive work in areas of innate immunity, sterile inflammation, and cell–cell communication provided good evidence that a fine balance of and crosstalk between various mediators released during injury determines the outcome of renal inflammation.^5–8 The end result of renal inflammation ranges from complete resolution to ongoing generation of danger-associated molecular patterns (DAMPs), extracellular matrix (ECM) accumulation with scar formation, loss of highly differentiated renal cells, finally leading to organ fibrosis and loss of function.^1,4,9,10 DAMPs are endogenous molecules, which are recognized by different pattern recognition receptors to alert the innate immune system through various signaling pathways.⁴

ECM accumulation plays a prominent role in the evolution of renal disease, as it might lead to a deranged matrix composition and structure finally to scar formation. It also acts as a meshwork for different molecular mediators, such as enzymes, growth factors, and cytokines.^11,12 Among various ECM components, the small leucine-rich proteoglycans (SLRPs) appear to play a crucial role in renal inflammation and fibrogenesis.^13–17

The SLRP family consists of 18 distinct proteoglycans that are characterized by their relatively small molecular mass protein cores of 36–42 kDa and the presence of leucine-rich repeats (LRRs) within their structure.^18–20 The SLRPs are organized in five classes depending on evolutionary conservation, protein and genomic homology, and chromosomal organization.¹⁸ The class I SLRPs decorin and biglycan, as well as lumican and fibromodulin belonging to class II, are the best characterized members of the SLRP family.¹⁸

During synthesis, SLRPs are glycosylated and posttranslationally modified in the Golgi apparatus and secreted by exocytosis directly into the extracellular environment.²¹ In the normal kidney, decorin, biglycan, lumican, and fibromodulin proteins are present mainly in the tubulointerstitium, and only trace amounts are detected in the glomerulus. Details regarding the discrepancy between the mRNA and protein expression of SLRPs in the kidney have been extensively discussed in previous publications.^13,17,22,23 When bound to the ECM, SLRPs interact with various types of collagen and regulate fibril growth and organization, cell-matrix interactions, ECM assembly, and tissue function.²⁴ Moreover, matrix-bound SLRPs are able to sequester signaling mediators in the ECM, thus interfering with different signaling cascades.^25,26

Upon proteolytic digestion or due to ECM saturation, SLRPs are released and act as soluble mediators.^4,27–30 For details regarding numerous proteases involved in SLRP fragmentation, please refer to more topical reviews.^17,19,30,31 Soluble SLRPs engage specific receptors and signaling pathways in both immune and resident renal cells leading to either resolution or aggravation of disease.¹⁹ Based on their interactions with Toll-like receptors (TLRs) and the inflammasome, biglycan and decorin act as ECM-derived DAMPs and promote the inflammatory response.⁴

TLRs are transmembrane pattern recognition receptors (PRRs) expressed in both immune and non-immune cells either on the cell surface (TLR1, 2, 4, 5, and 6) or intracellularly on the endolysosomal membrane (TLR3, 7, 8, and 9).³² Inflammasomes are cytosolic PRRs, which form multiprotein complexes that can trigger the activation of the cysteine protease caspase-1, positioned upstream of interleukin (IL)-1β maturation.³³ There are several inflammasomes named after the protein forming the scaffold: NLR family CARD domain–containing protein 4 (NLRC4), NACHT, LRR, and PYD domains–containing protein (NLRP) 1, NLRP3, and absent in melanoma 2 (AIM2). While each of them requires a different stimulus, all of them promote caspase-1 activation.³⁴

It has become increasingly obvious that SLRPs besides acting as “classic DAMPs” may also induce anti-inflammatory responses.^28,35,36 Various decorin signaling receptors and pathways, not directly involved in the regulation of renal inflammation and fibrosis, are described in a comprehensive review focused on proteoglycan receptors.³⁷

A host of comprehensive reviews have been published on the importance of SLRPs in renal pathologies.^{1,14,17,19,26,30,38} In the current review, we aim to address the complexity of SLRPs signaling, which confers a dual role either to aggravating or to repairing tissue damage during kidney disease. Specific signaling pathways directly driven or influenced by SLRPs in immune and resident renal cells in correlation with the outcome of renal inflammation are discussed. Special attention was paid to pro- and anti-inflammatory effects of SLRP signaling in the kidney. As most of the data published up to now are centered on only a few representative SLRP members, we have focused our review on decorin, biglycan, lumican, and fibromodulin.

Soluble SLRP Levels and Renal Disease Progression

Over the last 20 years, research on proteoglycans has provided strong evidence that besides ECM-bound SLRPs, there are unsequestered forms that circulate in plasma or are secreted into urine during various inflammatory diseases.^1,17,19 In fact, the earliest hints regarding the release of soluble form of SLRPs were mainly coming from renal research.^14,17 Initially, the biological role of soluble SLRPs was not understood at all. Due to a lack of sensitive methods to isolate SLRPs from tissue, plasma, or urine, only decorin was detected in the urine of a patient with membranous nephropathy.^13,23

Another early suggestion for the presence of secreted SLRPs was provided by extensive studies in renal sections and urine from patients with DN.¹³ In this report, a clear discrepancy was observed in the glomerular mRNA and protein expression in patients with DN. In contrast to the mRNA overexpression for decorin, biglycan, lumican, and fibromodulin in glomeruli from patients with incipient, manifest, and advanced DN, the protein expression of these SLRPs was enhanced only at advanced stages disease.¹³ This strongly correlated with scar formation, which is due to enhanced accumulation of the ECM-bound SLRPs in the kidney.¹⁴ Accordingly, patients with DN have high plasma levels of decorin during the development of glomerulosclerosis.¹³ Thus, it is conceivable that before glomerular scar formation, large amounts of SLRPs are synthesized but not retained in the mesangial matrix and therefore released into the urine or bloodstream.^13,17,23

Once it became obvious that soluble biglycan and decorin act as signaling molecules,^15,27,39 more attention was paid to plasma and urinary SLRP levels during the progression of renal diseases. Accordingly, increased plasma levels of soluble biglycan in murine and human lupus nephritis (LN) correlate with the development of albuminuria, enhanced B-cell chemoattractant chemokine (C-X-C-motif) ligand (CXCL) 13 levels, and elevated numbers of renal B cells.¹⁶ In renal ischemia reperfusion injury (IRI), renal overexpression of biglycan correlates with kidney injury and the level of soluble biglycan in the circulation.^40,41 Not many data are available regarding the soluble forms of other SLRPs in correlation with renal damage. Screening plasma samples from patients with DN revealed the presence of soluble lumican.⁴² More details can be found in a comprehensive overview regarding SLRPs as biomarkers in renal diseases.¹⁷

The source of circulating SLRPs is not completely clarified, yet. However, it is conceivable that SLRPs are released from the kidney due to enhanced de novo synthesis of SLRPs triggered, e.g., by transforming growth factor-β (TGF-β) and proinflammatory cytokines.^15,27 Thus, rapidly increasing abundance of newly produced SLRPs may exceed the capacity of the ECM to sequester these molecules. In addition, ECM-bound SLRPs can be released from the ECM by various proteolytic enzymes to further enhance the pool of soluble SLRPs.^17,43,44

The sparse data regarding correlation of renal, plasma, and urinary SLRP levels with the degree of kidney damage is due to complex methods for SLRP isolation and detection. Another important obstacle is the lack of specific antibodies to SLRP neoepitopes as indicator of proteolytic digestion. As only unsequestered SLRPs are capable of inducing signaling events,^27,45 the detection of SLRP fragments in plasma instead of tissue samples might be a more promising approach to correlate progression of renal disease with SLRP levels for diagnostic purposes. Thus, there is still an unmet need to simplify detection of soluble SLRPs in tissue, plasma, and urine.

Signaling of Soluble SLRPs in Immune Cells

The realization that soluble biglycan and decorin act as signaling molecules^20,31,46 represented a paradigm shift in our understanding of SLRP-dependent regulation of renal disease.¹⁴ Soluble biglycan and decorin are recognized as ECM-derived DAMPs.^27,28 In macrophages, by signaling through TLR2, TLR4, and the inflammasome, they act as proinflammatory molecules triggering autonomously sterile or potentiating pathogen-dependent inflammation.^27,28,45,47 However, recent data suggest that SLRPs in crosstalk with other signal transduction pathways are under certain conditions also capable of dampening the inflammatory response.^17,28 In this section, an extensive research emphasizing the signaling properties of soluble SLRPs in immune cells is summarized.

Proinflammatory Signaling of Soluble SLRPs in Immune Cells: Ramifications for Renal Inflammation

Proinflammatory Effects of Biglycan in Immune Cells

The initial notion that biglycan regulates behavior of innate immune cells was based on the observation that in tubulointerstitial inflammation biglycan is overexpressed in tubules before macrophage infiltration.¹⁵ Indeed, in macrophages, soluble biglycan has been shown to form a complex with the LRR-containing TLR2 and TLR4 and their co-receptors cluster of differentiation (CD) 14 and lymphocyte antigen 96 (MD2).²⁷ Downstream, this leads to activation of the mitogen-activated protein kinase (MAPK) p38, Erk, and NF-κB.²⁷ It is of note that the affinity binding of biglycan to TLR4/MD2 complex is comparable with the binding of lipopolysaccharide (LPS).⁴¹ This together with the fact that biglycan, in contrast to LPS, also interacts with TLR2 might explain the strong proinflammatory effects of this proteoglycan. Although the roles of TLR2 and TLR4 in biglycan-triggered signaling have been described in great detail, more insights regarding the involvement of the TLR2/4 co-receptors still have to be revealed.

There is growing evidence that soluble biglycan by selectively engaging a single TLR receptor and “choosing” among various TLR adaptor molecules stimulates different signaling branches, which in turn lead to different biological outcomes (Fig. 1).^36,48 Downstream of TLR2 and TLR4, the adaptor molecule myeloid differentiation primary response gene 88 (MyD88) is required.⁴⁹ TLR4 is capable of signaling via an alternative pathway utilizing TIR-domain-containing adapter-inducing interferon-β (TRIF).⁴⁹ In macrophages, exclusive signaling of biglycan through TLR4 and TRIF results in NADPH oxidase (NOX) 2 production (Fig. 1, upper panel). In contrast, by employing TLR4 and the MyD88 adaptor, biglycan triggers translocation of p47^phox from the cytoplasm to the plasma membrane, necessary for NOX2 activation (Fig. 2).³⁵ Through utilization of TLR2 and MyD88, biglycan induces heat shock protein (Hsp) 70 production and its binding to NOX2,³⁵ interaction known to facilitate the proteasomal degradation of NOX2 (Fig. 1, upper panel).⁵⁰ Recent data indicate that biglycan triggers erythropoietin (Epo) expression in the liver and kidney by signaling exclusively through TLR2.⁵¹ Signaling of biglycan via TLR4 and TRIF causes production of sphingosine kinase (SphK) 1, an important mediator in the sphingolipid-dependent signaling cascade.³⁶ Furthermore, soluble biglycan stimulates in a MyD88-dependent manner the production of tumor necrosis factor (TNF)-α, IL-1β, and of CXCL1, CXCL2, and chemokine (C-C motif) ligand (CCL)2 downstream of TLR2/4 and causes recruitment of neutrophils and macrophages in the kidney (Fig. 1, lower panel).^36,48 Simultaneously, biglycan induces CCL5 through TLR4/TRIF and attracts both T cells and macrophages.⁴⁸ Moreover, SphK1 potentiates the biglycan-driven CCL2 and CCL5 production and macrophages recruitment (Fig. 1, lower panel).³⁶ The mechanisms of biglycan-dependent selection of TLRs and their adaptors are still unknown. It is conceivable that several not yet identified co-receptors of biglycan/TLR2/4 interactions are required for signaling specificity.

Figure 1.

Mechanism of biglycan-mediated renal inflammation. (Upper panel) In its soluble form, biglycan triggers NOX2 and Hsp70 expression through TLR4/TRIF or TLR2/MyD88, respectively. NOX2 and Hsp70 proteins interact, and this facilitates the proteasomal degradation of NOX2. Following binding to TLR2 and TLR4 and involvement of NOX1- and NOX4-derived ROS, biglycan promotes the synthesis of Il1β gene and production of IL-1β. Furthermore, biglycan drives the production of CXCL13 via TLR2/4/NOXs and consequently the recruitment of B cells. (Lower panel) Biglycan induces the pro-IL-1β and TNF-α production via TLR2/4/MyD88 pathways. Through the same mechanism, biglycan generates CXCL1 and therefore neutrophils recruitment. Soluble biglycan clusters the TLR2 and TLR4 receptors with the P2X₇ purinergic receptor and leads to the NLRP3 inflammasome assembly, caspase-1 activation, and the cleavage of pro-IL-1β into mature IL-1β. In addition, in a SphK1-dependent manner, biglycan induces CCL5 via TLR4/TRIF and CCL2 through TLR2/4/MyD88 leading to the attraction of T cells and macrophages, respectively. Abbreviations: NOX, NADPH oxidase; Hsp70, heat shock protein 70; TLR, toll-like receptor; TRIF, Toll/IL-1R domain-containing adaptor inducing IFN-β; MyD88, myeloid differentiation primary response protein 88; ROS, reactive oxygen species; IL-1β, interleukin-1β; CXCL, chemokine (C-X-C-motif) ligand; TNF-α, tumor necrosis factor-α; P2X₇, purinergic receptor 7; NLRP3, NACHT, LRR, and PYD domains–containing protein-3; SphK1, sphingosine kinase 1; TLR, toll-like receptor; CCL; chemokine (C-C-motif) ligand; MΦ, macrophage.

Figure 2.

SLRPs-induced anti-inflammatory mechanisms. Soluble biglycan induces the synthesis of the pro-inflammatory Il1β and Ccl2 genes through TLR2/4 and Ccl5 only via TLR4. In addition, biglycan triggers NOX2 expression and p47^phox translocation to the plasma membrane via TLR4/TRIF and TLR4/MyD88, respectively. Consequently, the NOX2 enzyme complex is assembled and activated in a Rac1-dependent manner. Biglycan-induced Il1β and Ccl2 genes synthesis is impaired by NOX2 activation. Soluble decorin triggers the transcription of the anti-inflammatory Il10 gene following binding to TLR2 and TLR4. Lumican binds LPS and presents it to the receptor, thereby potentiating the LPS/TLR4-triggered anti-inflammatory Il10 and Il4 genes. MMP-8 can cleave fibromodulin and release bound TGF-β, which in turn becomes active and leads to M2-macrophage polarization. Abbreviations: SLRPs, small leucine-rich proteoglycans; Il, interleukin; CCL, chemokine (C-C-motif) ligand; TLR, toll-like receptor; NOX, NADPH oxidase; TRIF, Toll/IL-1R domain–containing adaptor inducing IFN-β; MyD88, myeloid differentiation primary response protein 88; LPS, lipopolysaccharide; MMP-8, metalloproteinase-8; TGF-β, transforming growth factor-β.

Another level of complexity in biglycan signaling is created by the ability of biglycan to orchestrate the crosstalk between various receptors and pathways.^16,35,36,45 By clustering TLR2/4 with the P2X₇ purinergic receptor, biglycan triggers autonomously the maturation of IL-1β by activating the NLRP3 inflammasome and caspase-1 (Fig. 1, lower panel).^16,45

There is growing evidence for the crosstalk between biglycan- and reactive oxygen species (ROS)-signaling.^16,35,45,52 In this context, biglycan enhances Il1β synthesis and IL-1β production via the TLR2/4/MyD88 pathway and in a NOX1/4-dependent manner (Fig. 1, upper panel).³⁵ Furthermore, biglycan induces the B-cell chemoattractant CXCL13 in macrophages by interacting with TLR2/4 and depending on the NOXs-derived ROS generation (Fig. 1, upper panel).¹⁶ However, the exact mechanism of NOXs involvement in CXCL13 regulation is still not fully understood.

Up to now, biglycan signaling and its implications for the neutrophils,⁴⁸ macrophages,^{16,27,36,41,45,48} and T^16,53 and B lymphocytes¹⁶ recruitment have been primarily investigated in macrophages and dendritic cells.^4,53 A report, published before the identification of biglycan receptors, showing potentiating effects of biglycan on IL-7-dependent pre-B-cell proliferation,⁵⁴ suggests that there is a need to continue investigations addressing the direct influence of biglycan on immune cells other than macrophages.

With regard to biglycan signaling in renal resident cells, there is only one recent report indicating that biglycan stabilizes hypoxia-inducible factor (HIF)-2α by exclusively signaling through TLR2, thereby leading to enhanced Epo secretion from the kidney. In consequence, biglycan causes secondary polycythemia increasing hemoglobin concentration, red cell numbers, and total iron binding capacity.⁴⁹ This was quite an unexpected outcome of TLR2 and biglycan signaling, reaching far beyond the canonical context of innate immunity and inflammation.

Implications of Biglycan Signaling on Inflammatory Renal Diseases

Extensive in vivo data regarding biglycan signaling in the kidney show beneficial effects of biglycan deficiency, while animals overexpressing soluble biglycan display an inflammatory phenotype.^{16,35,41,45,51,55} Importantly, direct proof of the in vivo involvement of TLR2/4 in biglycan signaling was provided by mice transiently overexpressing soluble biglycan and are deficient of TLR2 or TLR4. Indeed, lack of TLR2 and/or TLR4 abolished biglycan-dependent production of proinflammatory mediators, reduced infiltration of mononuclear cells, and eliminated biglycan-mediated Epo production.^16,41,51

In experimental LN, biglycan overexpression triggered TLR2- and TLR4-dependent systemic and renal disease aggravation. Elevated levels of TNF-α and various other chemokines such as CCL2, CCL3, CCL5, CXCL13 led to enhanced recruitment of macrophages and T and B lymphocytes into the kidney, causing aggravation of organ damage.¹⁶ By contrast, biglycan deficiency ameliorated the progression of LN.¹⁶ In renal IRI in mice, soluble biglycan enhanced plasma and renal levels of TNF-α, CXCL1, CCL2, and CCL5; caused infiltration of neutrophils, macrophages, and T cells; and worsened renal function. Concomitant ablation of TLR2 and TLR4 diminished these effects.⁴¹ Apart from direct evidence, there are several indirect indications that biglycan-TLR2/4 signaling in the kidney is associated with aggravation of renal injury in various kidney disease conditions.^{14,17,40,52,56}

To activate TLR4, biglycan is known to engage two adaptor molecules, MyD88 and TRIF, whereas interaction with TLR2 requires involvement of MyD88.⁴⁸ Transient overexpression of soluble biglycan in MyD88- and TRIF-deficient mice provides direct proof of biglycan/MyD88-dependent production of CXCL1, CCL2, which enhances recruitment of neutrophils and macrophages into the kidney.^27,41,48 At the same time, biglycan induces CCL5 through TRIF and attracts both T cells and macrophages into the kidney.⁴⁸

The importance of biglycan-dependent stimulation of the inflammasome in the kidney is confirmed in models of sterile renal inflammation and fibrosis (unilateral ureteral obstruction [UUO]) and in LN.^16,45 In both models, biglycan-deficient animals display lower levels of active caspase-1 and mature IL-1β in the affected kidneys in correlation with less severe renal damage.^16,45 In LN mice, overexpressing soluble biglycan, renal abundance of active caspase-1 and mature IL-1β was increased and tissue damage was aggravated.¹⁶

Despite the inherent complexity of in vivo systems, there is some evidence for the crosstalk between biglycan- and ROS-signaling in the kidney. NOX2-deficient mice have an inflammatory phenotype potentiated in the kidney by renal IRI.³⁵ However, biglycan deficiency improved the outcome of renal IRI by rescuing kidney function, reducing renal Hsp70, CCL2, and IL-1β production, and limiting macrophage recruitment.³⁵ Recent in vivo data unambiguously showed the relevance of crosstalk between biglycan- and SphK1-driven ECM- and sphingolipid-signaling for the outcome of renal inflammation. Transient overexpression of soluble biglycan in mice lacking Sphk1 caused enhanced production of renal CCL2 and CCL5 and recruitment of macrophages into the kidney.³⁶

Thus, there is growing evidence that biglycan is a potent trigger of sterile inflammation in the kidney. Even though the inflammatory reaction following renal injury is primarily a protective response,⁵⁷ biglycan-induced hyperinflammation is clearly detrimental and in the end aggravates kidney injury. Thus, inhibition of biglycan signaling might be a promising therapeutic strategy to mitigate renal inflammation and fibrogenesis.

Proinflammatory Signaling of Decorin: Implications for Renal Inflammatory Diseases

Decorin is well established in nephrology as a TGF-β-binding partner and inhibitor of its activity with beneficial effects on renal fibrogenesis.⁵⁸ Therefore, in this context, identification of soluble decorin as ECM-derived DAMP²⁸ is quite surprising. In addition, decorin is highly homologous to biglycan and contains, similarly to TLR2/4 and CD14, LRRs within its protein structure.¹⁸ Thus, it is not surprising that decorin and maybe other SLRPs will act as DAMPs.

It is widely recognized that decorin is a high-affinity ligand of TLR2 and TLR4 that triggers the production of proinflammatory TNF-α and IL-12p70 in a NF-κB- and MAPK-dependent manner.²⁸ Interestingly, different effects regarding the regulation of Il10 mRNA and IL-10 protein expression are displayed by decorin as opposed to biglycan. On one hand, decorin induces the anti-inflammatory Il10 gene expression via TLR2/4. On the other hand, it inhibits active TGF-β1. For details regarding decorin interactions with the TGF-β, please refer to more topical reviews.^14,19,26 TGF-β1 inhibition leads to a reduction in processing of primary miR-21 to precursor miR-21 and thus its transformation into mature miR-21.^28,59 By decreasing miR-21, which acts as a posttranscriptional inhibitor of programmed cell death 4 (PDCD4), the levels of PDCD4 increase. Enhanced levels of PDCD4, a posttranscriptional inhibitor of IL-10 production, result in lower abundance of the anti-inflammatory cytokine IL-10 in macrophages.²⁸ Thus, this dual signaling of decorin involving TLR2/4 activation and TGF-β1 inhibition strongly promotes a proinflammatory response.²⁸

The in vivo verification of decorin-driven dual proinflammatory signaling is provided in sepsis and tumor xenografts.²⁸ Even though data regarding proinflammatory effects of decorin in the kidney are sparse, direct evidence in favor of the mechanism described above²⁸ is provided by transient overexpression of soluble decorin in UUO.¹⁷ Indeed, enhanced levels of soluble wild-type decorin increased the infiltration of macrophages in the interstitium of both contralateral and obstructed kidneys.¹⁷ As UUO causes increased expression and accumulation of collagen type I,¹⁵ part of the soluble decorin might be captured by collagen and sequestered into the ECM.⁶⁰ This hypothesis has been proven in UUO by transiently overexpressing a soluble decorin mutant, which does not bind to collagen type I.¹⁷ As expected, this mutant resulted in higher macrophage numbers recruited into contralateral and obstructed kidneys as compared with wild-type decorin.¹⁷ Thus, it appears that similar to biglycan, only non-sequestered decorin is capable of interacting with TLRs to promote proinflammatory signaling. Identification of decorin as DAMP might explain why initial findings regarding the antifibrotic effects of decorin have not been transferred into clinical practice.

Signaling of Other SLRPs and Their Involvement in Renal Inflammation

Our knowledge regarding signaling pathways evoked of other SLRPs is limited to a few studies referring to inflammation. Lumican potentiates the TLR4/CD14- and NF-κB-mediated inflammatory response in LPS-induced sepsis.⁴⁷ Accordingly, in a hapten 2-4-5, trinitrobenzene sulfonic acid-induced colitis model lumican-deficient mice display impaired synthesis of TNF-α, IL-1β, and IL-6.⁶¹ Mechanistically, it was suggested that lumican binds LPS to present it to CD14.⁴⁷ Only recently, lumican has been associated with renal disease.⁶² LPS treatment of lumican-overexpressing mice led to enhanced production of TNF-α, IL-6, IL-4, and IL-10 in renal tissue compared with wild-type animals, which was associated with an acute decline of renal function.⁶² However, it is still necessary to understand whether the soluble or sequestered or both forms of lumican are responsible for these effects. Asporin (class I SLRP) and fibromodulin (class II SLRP)¹⁸ appear to display anti-inflammatory effects as described in the chapter below.^63,64

Anti-Inflammatory Effects of SLRPs

Recent data show that SLRPs, besides their conventional role as proinflammatory DAMPs, are also involved in self-limitation of inflammation.^28,35,36,51 It is well known that biglycan by selectively triggering TLR2/4 and their adaptor molecules tightly regulates inflammation.^{35,36,45,51,52} A prototypical example is the biglycan-dependent control of IL-1β synthesis and maturation in macrophages. On one side, biglycan stimulates receptor-signaling collaboration between TLR2/4, the P2X₇ receptor, and the NLRP3 inflammasome to produce and activate proinflammatory IL-1β (Fig. 1).⁴⁵ On the other side, biglycan promotes an anti-inflammatory mechanism controlling IL-1β overproduction by TLR4/TRIF-dependent synthesis and NOX2 (Fig. 2).³⁵ In addition, biglycan triggers activation of the NOX2 complex through TLR4/MyD88-dependent and Rac1- and Erk-mediated translocation of p47^phox (Fig. 2). The inhibitory effects of NOX2 on biglycan-stimulated IL-1β synthesis, and probably on the maturation as well, were shown in wild-type versus NOX2-deficient macrophages. The presence of NOX2 in macrophages markedly reduces biglycan-evoked expression of caspase-1, Nlrp3, and Il1β mRNAs as well as lowers mature IL-1β levels.³⁵ Thus, it is tempting to speculate that these anti-inflammatory mechanisms are involved under physiological conditions to fine-tune biglycan-dependent IL-1β synthesis and maturation. In inflammatory renal diseases, biglycan causes Hsp70 production, which binds NOX2, thereby blocking the inhibitory effects on IL-1β production and promoting inflammation.³⁵

The active NOX2 complex is also involved in inhibition of biglycan-triggered Ccl2 expression in macrophages and in the kidney challenged by IRI (Fig. 2). Interestingly, this mechanism is not employed in the regulation of the biglycan-triggered expression of Ccl5 (own unpublished data). This suggests a very specific regulation of inflammation. Thus, identification of physiological inhibitors of single proinflammatory cytokines and chemokines would be a milestone for translational research in proteoglycan biology.

As pleiotropic anti-inflammatory effects of Epo have been described in chronic inflammatory or infectious diseases,⁶⁵ biglycan-mediated Epo production in liver and kidneys might represent an additional biglycan-dependent anti-inflammatory mechanism.⁵¹

Data regarding the anti-inflammatory effects of other SLRPs are limited. An interesting anti-inflammatory mechanism was described for fibromodulin. By direct binding, fibromodulin sequesters TGF-β in its inactive form in the ECM.⁶³ Following fibromodulin degradation by matrix metalloproteinase (MMP)-8, the sequestered form of TGF-β is released. Subsequently, TGF-β potentiates the M2-macrophage polarization⁶³ (Fig. 2) associated with the resolution of inflammation.^66,67 The capability of soluble decorin to trigger the transcription of the anti-inflammatory Il10 gene in macrophages following binding to TLR2 and TLR4 indicates that decorin under certain conditions is capable of downregulating inflammation (Fig. 2).²⁸ The relevance of these findings for renal inflammation has not yet been determined.

Lumican, by presenting LPS to TLR4,⁴⁷ is able to potentiate LPS-induced IL-4 and IL-10 anti-inflammatory cytokine expression (Fig. 2).⁶² However, this was not associated with preventing the decline of renal function.⁶²

Antifibrotic Effects of Decorin in the Kidney

There are no doubts that the final outcome of pro- and anti-inflammatory effects of biglycan in inflammatory kidney diseases promotes tissue injury and loss of renal function.^{16,35,41,45,51,55} Taking into account the dual proinflammatory signaling of decorin in macrophages,²⁸ described above, even stronger deteriorating effects on the outcome of renal diseases would be expected for decorin versus biglycan. However, in various organs and diseases resulting in fibrogenesis, decorin clearly exerts antifibrotic effects. For details, please refer to reviews focused on organ fibrosis.^{14,17,19,26,29,68,69}

Beneficial effects of decorin in the kidney are based on neutralizing effects on two major profibrotic factors, TGF-β1^70,71 and connective tissue growth factor (CTGF)/CCN2.^72–74 Several direct and indirect mechanisms of decorin-dependent TGF-β inactivation^70,75–78 caused by physical interaction, inhibition of TGF-β signaling, and regulation of TGF-β modulators (e.g., fibrillin-1) have been described at length in earlier reviews.^14,19,26

Besides inhibition of TGF-β1 and CTGF, decorin-mediated protection of tubular epithelial cells from apoptosis appears of particular importance in renal fibrogenesis. This is clearly indicated by the fact that decorin-deficient mice challenged by UUO and streptozotocin-induced diabetes display markedly aggravated renal fibrosis, caused by TGF-β-independent apoptotic loss of tubular epithelial cells.^15,74,76 Mechanistically, the decorin protein core binds to and phosphorylates the insulin-like growth factor (IGF)-I receptor with subsequent activation of Akt/protein kinase B (Akt/PKB) and induction of p21^WAF1.^76,79 Thus, it appears that the crosstalk of decorin signaling in macrophages and in renal resident cells does not promote chronification of inflammation and protects against fibrosis.

Recent data identified decorin as an important regulator of autophagy in epithelial and carcinoma cells.^80–86 Details regarding autophagic signaling pathways evoked by decorin are described in recent reviews.^87–90 It is conceivable that autophagy could be one of the decorin-mediated antifibrotic mechanisms. Thus, experimental data are needed to proof this hypothesis in the kidney.

Over the years, much has been argued on whether SLRPs promote fibrosis or protect against it. Their intricacy is due to the fact that on one hand they are ECM components and therefore in their sequestered form a part of a fibrotic scar. On the other hand, in their soluble form, they turn into signaling molecules. Identification of numerous signaling receptors for decorin and biglycan provided a better understanding of their complex role in renal diseases. In particular, recognition of biglycan and decorin as ECM-derived DAMPs emphasized their biological relevance in inflammatory renal diseases. SLRPs are capable of simultaneously interacting with and orchestrating various receptor signaling.²⁹ By “choosing” different adaptor molecules, SLRPs modify signaling pathways and downstream effects of the same receptor.^35,48 To further add to the complexity of SLRP signaling, recent data have shown that the outcome of SLRP signaling via a single innate immune receptor might differ in immune as opposed to renal resident cells.^27,51 Therefore, it is conceivable that the influence of SLRPs on renal inflammation and disease outcome will depend to a large extent on the specific biological context: structural versus soluble form of SLRPs, tissue and cell types involved, type and stage of disease.

In addition, it became widely accepted that SLRPs besides acting as “proinflammatory DAMPs” are deeply involved in the regulation of the inflammatory response. In fact, newer findings clearly showed that under certain conditions soluble biglycan also exerts anti-inflammatory effects. Biglycan is capable of suppressing inflammation by enrolling TLRs and crosstalking with other transduction pathways such as ROS- or sphingolipid-signaling.^35,36,52

Even though many mechanisms have been already revealed, we have only just started to understand the complex signaling cascades, in which SLRPs are involved during renal inflammation and fibrosis. Therefore, it is of utmost importance to further determine which factors impact on the pro- and anti-inflammatory balance within SLRPs signaling pathways. In addition, there is a need for addressing the signaling capability of other members of the SLRP family, besides biglycan and decorin.

Taken together, targeting SLRP signaling in a selective and specific manner depending on the SLRP member, the involved receptor and adaptor molecules, the type of pro- or anti-inflammatory outcome, tissue/cell types, and stage of disease might be a promising therapeutic approach for inflammatory and fibrotic renal diseases.