N-acetyl-seryl-aspartyl-lysyl-proline is a valuable endogenous antifibrotic peptide for kidney fibrosis in diabetes: An update and translational aspects.

N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) is an endogenous peptide that has been confirmed to show excellent organ-protective effects. Even though originally discovered as a modulator of hemotopoietic stem cells, during the recent two decades, AcSDKP has been recognized as valuable antifibrotic peptide. The antifibrotic mechanism of AcSDKP is not yet clear; we have established that AcSDKP could target endothelial-mesenchymal transition program through the induction of the endothelial fibroblast growth factor receptor signaling pathway. Also, recent reports suggested the clinical significance of AcSDKP. The aim of this review was to update recent advances of the mechanistic action of AcSDKP and discuss translational research aspects.

Introduction

With the recent type 2 diabetes pandemic, the number of people with diabetic complications could increase in the future. Diabetic kidney disease, such as diabetic nephropathy or other kidney insults, is the major cause of end‐stage renal disease and renal replacement therapy. Kidney fibrosis, which is characterized by matrix deposition and glomerulosclerosis, is the final pathway of progressive kidney diseases. Fundamentally, organ fibrosis is an important tissue repair process; progressive kidney fibrosis might be due to disruption of the normal wound healing mechanisms1, 2. The primary factors in the fibrosis pathway; for example, cellular induction of fibrosis or production of excess extracellular matrix (ECM), are difficult to identify. All cell types, either resident or non‐resident, in the kidney, such as resident fibroblasts, tubular epithelial cells, pericytes, endothelial cells, vascular smooth muscle cells, mesangial cells, podocytes and migrated bone marrow derived cells or inflammatory cells, are involved in this process3, 4. All these cell types could play vital roles in kidney fibrosis. Additionally, the conversion programs of cells from an epithelial or endothelial phenotype to a mesenchymal phenotype through epithelial–mesenchymal transition (EMT) and endothelial–mesenchymal transition (EndMT) could promote the generation of matrix‐producing mesenchymal‐like cells4, 5. However, there has been intensive discussion about the contribution or even the presence of EMT or EndMT. Most likely, the complete form of EMT or EndMT would make a minor contribution, if any; the presence of partial EMT or EndMT, the transition phase of epithelial/endothelial cells into mesenchymal cells, would likely have a major impact, because mesenchymal marker expression is common in either epithelial or endothelial cells in fibrotic kidney biopsies6, 7, 8. Therefore, targeting these mesenchymal programs is essential when considering therapeutic options for kidney fibrosis in diabetes. Diabetes is associated with endothelial damage induced by various insults, such as high glucose, hyperinsulinemia, oxidative stress and transforming growth factor (TGF)‐β. The present author and co‐authors have investigated endothelial damage and the EndMT program among the many potential mesenchymal programs in diabetic kidneys.

N‐acetyl‐seryl‐aspartyl‐lysyl‐proline (AcSDKP) is an endogenous peptide that has antifibrotic effects. The suppressive effects of AcSDKP on EndMT have been intensively analyzed. In this review, AcSDKP as a therapeutic option for diabetic kidney disease is focused on, especially the recent advances in endothelial protection induced by AcSDKP.

AcSDKP synthesis

AcSDKP is a tetrapeptide isolated from fetal calf bone marrow9, and studies have shown its strong antifibrotic properties in various organs of multiple disease models. Although the molecular pathway of endogenous AcSDKP synthesis has not been fully elucidated, thymosin β4 (Tβ4), a globular‐actin‐sequestering peptide, is the strongest candidate for the AcSDKP precursor (Figure 1)10, 11. Small interfering ribonucleic acid (RNA) knockdown of Tβ4 in HeLa cells resulted in significant suppression of AcSDKP levels11. Elegantly, Lenfant et al. 10 showed that [3H] AcSDKP was produced when radiolabeled [3H] Tβ4 was incubated with either bone marrow cells or bone marrow lysate. AcSDKP is the N‐terminal sequence of Tβ4 (Figure 1), and a subsequent study showed that prolyl oligopeptidase (POP; in some studies described as prolyl endopeptidase) is responsible for Tβ4‐mediated AcSDKP production (Figure 1)12. Interestingly, POP has shown to be suppressed in patients with relapsing–emitting multiple sclerosis13.

Figure 1

Generation of N‐acetyl‐seryl‐aspartyl‐lysyl‐proline (AcSDKP) from precursor polypeptide thymosin β4 (Tβ4). Tβ4 involves the amino acid sequence AcSDKP in the N‐terminal. Maprin‐α digested Tβ4 peptide into shorter sequence which can be cleaved by prolyl oligopeptidase (POP). In the second step, POP cleaves the AcSDKP sequence containing a shorter fragment of Tβ4 (1–13~24), and AcSDKP is excised out. Generated AcSDKP is degraded by angiotensin‐converting enzyme (ACE).

The Tβ4 peptide also shows antifibrotic and tissue‐protective effects. Tβ4 is a 43 amino acid peptide (4.9 kDa) that sequesters globular‐actin and regulates polymerization of F‐actin14, 15. Tβ4 is ubiquitously expressed in various organs14, 15. Tβ4 administration through either the intracardiac or the intraperitoneal route significantly rescued cardiac function and promoted neovascularization in a mouse model of experimental myocardial infarction16. Tβ4 also induced epicardial progenitor cell mobilization17. These organ‐protective effects of Tβ4 are primarily mediated by the synthesis of AcSDKP18, 19. In addition to Tβ4, Tβ15, which has the AcSDKP peptide sequence, might also contribute to the genesis of AcSDKP20, 21. The Tβ15 concentration is believed to be low compared with that of Tβ4 (~1,000‐fold lower); Tβ15 levels could be induced in some disease states, such as cancer20, 21. Interestingly, although the levels of AcSDKP were mostly suppressed, they were not completely abolished in mice with Tβ4 deficiency in the kidney and heart. These results might indicate that peptides homologous to Tβ4 (such as Tβ15) could compensate for the absence of Tβ422; however, caution is required in interpreting these data, because the AcSDKP level in the organs was determined with enzyme immunoassays for plasma/serum or urine, and such assays often show false positive values.

In the generation process of AcSDKP from Tβ4, recent evidence suggested the requirement of additional enzymatic cleavage processing. Kumar et al. 23 focused on the biology of POP. POP can cleave the peptide that is shorter than the sequence of 30 amino acids. However, Tβ4 is indeed 43 amino acids; therefore, theoretically, an additional enzymatic cleavage process should be required. Kumar et al.23 clearly showed that meprin‐α is a potential responsible enzyme to cleave Tβ4 and subsequently enable POP to cleave Tβ4 fragments to synthesize AcSDKP. Indeed, meprin‐α knockout mice showed significantly lower levels of AcSDKP. In regard to this, exogenous‐derived Tβ4‐increased AcSDKP levels in vivo were diminished by actinonin, the potential meprin‐α inhibitor23. These results suggested two‐step generation of AcSDKP from Tβ4 through meprin‐α and then POP (Figure 1).

AcSDKP degradation and the role of angiotensin‐converting enzyme

Angiotensin‐converting enzyme (ACE) was confirmed to hydrolyze AcSDKP (Figures 1,2). AcSDKP is maintained at a low level in the plasma, and the AcSDKP concentration increased fivefold after captopril administration24. ACE has N‐terminus (ACEN) and C‐terminus (ACEC) zinc‐binding catalytic domains that are responsible for the cleavage of target substrates25, 26. The amino acid homology between the ACEN and ACEC domain is approximately 60%; approximately 89% homology can be observed in the catalytic regions26. The ACE gene in higher organisms is believed to be the result of the ancient gene duplication27, and the resultant ACE, which has two catalytic sites, is found in many organs and cells (somatic ACE). In contrast, an ACE composed of only the ACEC without the ACEN is only found in the testis, and this testis ACE is vital for fertility28, 29, 30. The testis ACE is likely the primitive type of ACE26. This duplication of the ACE gene occurred in early evolution, and separate promoter regions regulate the different enzyme expression levels (Figure 2)27, 31, 32. These similar, but distinct, catalytic domains were described in detail in a previous report33.

Figure 2

The role of the distinct two catalytic sites of angiotensin‐converting enzyme (ACE) in tissue fibrosis. The C‐terminal catalytic site of ACE shows a higher affinity to angiotensin I. In contrast, the N‐terminal catalytic site of ACE shows a high affinity to N‐acetyl‐seryl‐aspartyl‐lysyl‐proline (AcSDKP), and AcSDKP could be only degraded by the N‐terminal catalytic site.

Only the ACEN is contributing AcSDKP hydrolysis (Figure 2). In fact, the testis (where the germinal‐type ACE without the ACEN is expressed) is known to have higher levels of AcSDKP than other organs34, 35. A seminal report from Li et al.36 showed that ACEN‐knockout mice, which showed high levels of AcSDKP, had significantly less bleomycin‐induced lung fibrosis, as shown by lung histology and hydroxyproline levels, than ACEC knockout mice, suggesting the vital role of the ACEN in AcSDKP degradation. Furthermore, they investigated whether S‐17092, a POP inhibitor, could decrease the lung‐protective effects in ACEN knockout mice, and as expected, S‐17092 abolished lung protection in these mice. AcSDKP suppressed bleomycin‐induced lung fibrosis in wild‐type mice. Overall, that study showed that physiological elevation of AcSDKP through inhibition of the ACEN leads to endogenous antifibrosis programs in the lungs36.

Kidney fibrosis

After kidney insults, profibrotic cytokines accumulate in the microenvironment and subsequently stimulate target cell activation, producing ECM, which is fundamental for renal fibrogenesis. The majority of matrix‐producing cells responsible for the production of interstitial matrix components (including fibronectin and type I and type III collagens) are fibroblasts37. Activated fibroblasts or myofibroblasts, which are characterized by increased expression of alpha smooth muscle actin (αSMA), are believed to be a significant source of ECM‐producing renal cells. However, almost all cell types in the kidney (either resident or non‐resident kidney cells) play a vital role in ECM production38. As described earlier, these cell types develop a profibrotic phenotype, and TGF‐β, a profibrotic cytokine that plays a central role in fibrogenesis, has an essential role in this process. Thus, inhibition of either TGF‐β or the TGF‐β‐stimulated Smad transcription factor signaling pathway shows antifibrotic effects39. Activated fibroblasts in the kidney express αSMA and are often called myofibroblasts, which show unique contractile properties37. However, αSMA expression was observed not only in fibroblasts, but also in tubular cells6 and endothelial cells8, suggesting that these cells could contribute to kidney fibrosis through EMT or EndMT program. Although the presence of these programs is controversial, at least in diabetic models, these processes are most likely important in kidney fibrosis. Interestingly, most studies did not detect EMT or EndMT in unilateral ureteral obstruction or ischemic reperfusion injury models. Unilateral ureteral obstruction is often utilized in kidney fibrosis studies, but the relationship of this model to human kidney disease is unclear. Differences are observed in the progression time and the background physiological condition. In diabetic models, EMT or EndMT has been detected (>150 papers on EMT and 20 papers on EndMT). Therefore, these programs should still be credible to be investigated in kidney fibrosis in diabetes.

Antifibrotic effects of AcSDKP

AcSDKP has been confirmed to show antifibrotic organ‐protective effects in diverse preclinical models18, 40, 41, 42, 43, 44, 45. In the first study of db/db mice42, the present author and co‐authors showed the preclinical utility of AcSDKP in protection against diabetic kidney disease, with a focus on kidney fibrosis.

Although various studies have shown the apparent antifibrotic effects in vivo and the direct effects of AcSDKP on culture fibroblasts in vitro, the effects of AcSDKP on fibroblast activation/myofibroblast differentiation programs are still unclear. Peng et al.46 reported that AcSDKP inhibited TGF‐β1‐induced differentiation of human cardiac fibroblasts into myofibroblasts, as shown by increased expression of αSMA and the embryonic isoform of smooth muscle myosin. Xu et al.47 reported that AcSDKP inhibits myofibroblast accumulation in silicotic nodules in the lung and the TGF‐β1‐induced myofibroblast differentiation of pulmonary fibroblasts. In a study of the AcSDKP‐inhibited myofibroblast differentiation program, the present author and co‐authors focused on the anti‐EndMT effects of AcSDKP associated with TGF‐β‐Smad signal transduction33, 48, 49, 50, 51.

More than 15 years ago, the present author and co‐authors found that AcSDKP inhibited TGF‐β‐induced Smad2 phosphorylation, and the anti‐TGF‐β/Smad pathway is the key to understanding the antifibrotic effects of AcSDKP in rat cardiac fibroblasts and human mesangial cells52, 53. Indeed, AcSDKP is the first endogenous circulatory polypeptide shown to inhibit TGF‐β‐induced Smad2 phosphorylation (only a Smad2 phosphorylation antibody was available at this time). Smads are TGF‐β superfamily‐specific transcription factors and are essential players in signal transduction39, 54, 55, 56. Smad transcription factors are categorized as follows: (i) receptor‐regulated (R)‐Smads (Smad2 and 3); (ii) common (co)‐Smads (Smad4); and (iii) inhibitory (I)‐Smads (Smad6 and 7). After TGF‐β binding to the type II receptor, this receptor physically interacts with and subsequently induces serine residue phosphorylation on the type I receptor (TGF‐βRI)57. The phosphorylated type I receptor interacts with and phosphorylates R‐Smads, and phosphorylated R‐Smads subsequently interact with co‐Smads and translocate into the nucleus with the help of importin‐β58. The R‐co‐Smad heterodimer binds to the Smad‐binding elements of the target promoter deoxyribonucleic acid regions, whereas I‐Smads localize to the nucleus53 and translocate to the cytoplasm from the nucleus after stimulation with TGF‐β to inhibit R‐Smad phosphorylation by type I receptors.

The precise molecular mechanisms by which AcSDKP suppresses TGF‐β‐induced R‐Smad phosphorylation are still unclear; I‐Smads could have a role in this process. The present author and co‐authors first reported that the I‐Smad Smad7 was translocated after AcSDKP incubation in human mesangial cells,53 and several groups have shown that the Smad7 levels were indeed elevated by AcSDKP administration in vivo 43. However, alternative molecular mechanisms by which AcSDKP‐inhibited R‐Smad phosphorylation could be linked to fibroblast growth factor (FGF) receptor (FGFR)49 and FGFR‐associated induction of the microRNA (miRNA) let‐7‐mediated suppression of TGF‐β159.

FGFR1

Fibroblast growth factor signaling, mediated by FGFRs, has an important role in maintaining endothelial homeostasis and cardiovascular integrity60, 61, 62, 63. The FGFR family, a subfamily of receptor tyrosine kinases, consists of four members (FGFR1, FGFR2, FGFR3 and FGFR4)64. Chen et al.59 reported that FGF/FGFR1 signaling suppressed TGF‐β‐induced EndMT through inducing the miRNA let‐7. The present author and co‐authors showed that a fibrotic strain of diabetic mice displayed suppression of FGFR1 on endothelial cells, and that AcSDKP restored the levels of endothelial FGFR1 in these mice49. FGFR1 is a fundamental molecule that inhibits the TGF‐β/Smad signaling pathway in endothelial cells65. Additionally, a key adaptor of the FGFR signaling pathway, FGF receptor substrate 2, has been shown to suppress EndMT induction, and endothelial‐specific FGF receptor substrate 2 knockout mice showed increased levels of EndMT‐ and EndMT‐derived smooth muscle cells59. A deficiency in mesodermal FGF receptor substrate 2 resulted in misalignment and hypoplasia of the cardiac outflow tract66. FGFR1 also plays an essential role in cardiomyocyte development and cardiac chamber identity in the developing ventricle67, 68.

Effect of AcSDKP on the FGFR1‐mitogen‐activated protein kinase kinase kinase kinase 4 pathway

Mitogen‐activated protein kinase kinase kinase kinase 4 (MAP4K4), a member of the Sterile 20 family of kinases, has various biological roles69. In endothelial cells, MAP4K4 has been shown to suppress integrin β1 activity by inducing moesin70. Integrin β1 stimulates EMT and organ fibrosis through TGF‐β/Smad signaling activation71, 72, 73. The present author and co‐authors showed that the interaction between dipeptidyl peptidase (DPP)‐4 and integrin β1 induces EndMT, and that this interaction disrupts endothelial homeostasis74. Misshapen (msn) kinases (the mammalian orthologs of MAP4K4) have been shown to suppress TGF‐β/Smad signaling75.

We found that AcSDKP restored the TGF‐β2‐suppressed levels of FGFR1 on endothelial cells, and that the FGFR1 levels were associated with MAP4K4 phosphorylation (Figure 3). AcSDKP was detected in close proximity to FGFR1. TGF‐β2 suppressed FGFR1, and AcSDKP restored the FGFR1 levels in association with the AcSDKP–FGFR1 interaction (Figure 3); however, mutant peptides, such as AcDSPK, AcSDKA and AcADKP, did not alter the levels of FGFR150. FGFR1 physically interacted with phosphorylated MAP4K4, and neutralizing FGFR1 abolished the AcSDKP‐induced MAP4K4 phosphorylation (Figure 3). Additionally, knockdown of MAP4K4 in endothelial cells abolished the anti‐EndMT and anti‐TGF‐β1/Smad effects of AcSDKP. Furthermore, the hearts of diabetic mice showed suppressed levels of both FGFR1 and MAP4K4 associated with Smad3 phosphorylation on endothelial cells; AcSDKP intervention normalized all of these parameters50. All these data showed that the FGFR1–MAP4K4 pathway is involved in the anti‐EndMT effects of AcSDKP (Figure 3).

Figure 3

N‐acetyl‐seryl‐aspartyl‐lysyl‐proline (AcSDKP)–fibroblast growth factor receptor 1 (FGFR1)–mitogen‐activated protein kinase kinase kinase kinase 4 (MAP4K4) axis‐induced suppression of integrin–transforming growth factor‐β receptors (TGFβRs) interaction and endothelial mesenchymal transition (EndMT). AcSDKP shows close proximity and would stabilize FGFR1. Alternatively, AcSDKP suppresses interferon‐γ (IFN‐γ), which inhibits FGFR1 levels in endothelial cells. FGFR1 physically interacts with MAP4K4 by which integrin β1 signaling is suppressed. Integrin β1 is essential for the heterodimer formations of TGFβRs; integrin β1 suppression directly results in the suppression of TGF‐β1‐smad signaling and EndMT.

β‐Klotho

Klotho proteins are type I single‐pass transmembrane proteins that share homology with family 1 β‐glycosidases, and are composed of α‐klotho (KLA)76, β‐klotho (KLB)77 and γ‐klotho78. Klotho proteins consist of short intracellular and relatively large extracellular domains with two internal repeats, termed KL1 and KL279. Klotho proteins act as coreceptors of FGFRs, facilitating the binding of the FGF19 subfamily on FGFRs (e.g., KLA for FGF23 and KLB for FGF19/FGF21)80, 81, and have emerged as essential metabolism‐regulating factors through their interactions during the past decades82, 83, 84, 85, 86, 87, 88.

KLA was shown to have anti‐aging effects76, and the functions of KLA in endothelial cells and the cardiovascular system have been reported89, 90, 91, 92. However, the biological significance of KLB is still unclear. KLB is expressed in human umbilical vein endothelial cells93 and human brain microvascular endothelial cells94, where KLB contributes to the formation of the blood–brain barrier.

We have reported that FGFR1 deficiency induced activation of TGF‐β/Smad3 signaling50; however, KLB knockdown was not associated with an obvious alteration of Smad3 phosphorylation51. As KLB knockdown did not directly alter the FGFR1 levels, FGFR1 could be essential for the regulation of Smad3 phosphorylation. In contrast, the mitogen‐activated protein kinase signaling pathways involving mitogen‐activated protein kinase kinase (MEK)1/2 and extracellular signal‐regulated kinases 1/2 were activated51. Consistent with this finding, a MEK inhibitor decreased KLB deficiency‐induced EndMT. Liu et al.95 reported that KLB overexpression inhibited EMT, the epithelial program sharing similar molecular mechanisms to EndMT, through the suppression of mitogen‐activated protein kinase. Unexpectedly, the KLB‐deficient activated mitogen‐activated protein kinase pathway without Smad3 phosphorylation is sufficient for the induction of EndMT. Either FGF19 or FGF21, a ligand of the FGFR1–KLB complex, synergistically diminished EndMT and MEK–extracellular signal‐regulated kinase pathway activation in AcSDKP‐treated cultured endothelial cells (Figure 4)51. Inhibition of FGFR1 and KLB protein levels was found in streptozotocin‐induced diabetic mouse hearts; AcSDKP restored these levels, suggesting the important in vivo effects of AcSDKP on FGFR1 and KLB complex formation51.These reports showed the essential role of the FGFR1‐KLB complex in the suppression of EndMT by AcSDKP, but the qualitative difference between Smad‐dependent EndMT and Smad‐independent, mitogen‐activated protein kinase‐dependent EndMT should be further investigated.

Figure 4

Fibroblast growth factor receptor 1 (FGFR1)–β klotho complex plays roles in endothelial homeostasis. (a) In normal endothelial cells, N‐acetyl‐seryl‐aspartyl‐lysyl‐proline (AcSDKP)‐induced FGFR1 levels and FGFR1–KLB complex. Such FGFR1–KLB complex is essential for the endothelial homeostasis by inhibiting both smad3 and mitogen‐activated protein kinase kinase (MEK)–extracellular signal‐regulated kinases (ERK) signaling pathway. (b) In the endothelial cells with KLB deficiency, MEK–ERK pathway dependent EndMT could be induced. KLB deficiency is not associated with alteration in FGFR1 levels in endothelial cells. AcSDKP cannot inhibit MEK–ERK pathway‐dependent EndMT in KLB‐deficient endothelial cells. (c) In the endothelial cells with FGFR1 deficiency, KLB levels are also significantly diminished, and both smad3 and the MEK–ERK pathway are activated. FGFR1‐deficient endothelial cells also show AcSDKP‐resistant endothelial mesenchymal transition (EndMT).

MiRNA cross‐talk

As described earlier, AcSDKP‐induced FGFR1 is associated with the anti‐EndMT miRNA let‐7 that targets TGF‐βRI; miRNA let‐7 induction through the AcSDKP–FGFR1 axis played further roles in endothelial protection through inducing miRNA‐298, 96, 97 and vice versa (Figure 5).

Figure 5

Micro ribonucleic acid (miRNA) cross‐talk between miR 29 and miR let‐7 in the anti‐endothelial mesenchymal transition action of N‐acetyl‐seryl‐aspartyl‐lysyl‐proline (AcSDKP). AcSDKP increases the levels of fibroblast growth factor receptor 1 (FGFR1) and, subsequently, miR let‐7s are induced. MiR let‐7 suppressed transforming growth factor β receptor (TGFβR1) and downstream Smad3 signaling, the major miR 29 inhibitory pathway. Therefore, the cells treated with AcSDKP also show induction of miR 29. miR 29 suppressed profibrotic and inflammatory cytokines and proteins, such as interferon‐γ, dipeptidyl peptidase‐4(DPP‐4) and so on. Interferon‐γ is known to suppress FGFR1. Interestingly, DPP‐4 inhibitor‐mediated suppression of DPP‐4 subsequently induced miR 29, and such DPP‐4 inhibitor‐activated miR 29 induced FGFR1‐dependent miR let‐7, as vice versa.

[Correction added on 1 April 2020, after first online publication: The figure has been amended to match the original.]

In a follow‐up study, the present authors and co‐authors found that AcSDKP‐induced let‐7 was associated with miRNA 29, and miRNA 29 expression was abolished by an antagomir for miRNA let‐7, suggesting antifibrotic cross‐talk between miRNA let‐7 and miRNA 2948. This conclusion is reasonable, because activation of TGF‐β signaling significantly suppresses miRNA 29, and the induction of miRNA let‐7 by miRNA 29 in endothelial cells is a reasonable sequence (Figure 5). Interestingly, miRNA 29 induction also suppressed EndMT, and was associated with the induction of miRNA let‐748. In this complex process, the present author and co‐authors identified the precise molecular mechanisms underlying the miRNA cross‐talk. MiRNA 29 targets several profibrotic molecules, such as integrin β1 and DPP‐448. Integrin β1 and DPP‐4 form a complex on the cell surface of endothelial cells, and induce TGF‐β receptor heterodimer formation and subsequent activation of the Smad signaling pathway74. TGF‐β‐induced EndMT was also inhibited by the DPP‐4 inhibitor linagliptin, which further restored miRNA 29. The induction of miRNA 29 is likely important for the antifibrotic effects of AcSDKP. MiRNA 29 also targets interferon‐γ, a cytokine related to inflammation and organ fibrosis. Interferon‐γ can inhibit endothelial FGFR1, the key mediator of miRNA let‐7 induction and the suppression of EndMT by AcSDKP59. The present author and co‐authors showed that the AcSDKP–FGFR1 axis was critical for maintaining endothelial mitochondrial biogenesis through induction of miRNA let‐7b‐5p98. These data showed that the AcSDKP–FGFR1 axis on endothelial cells is essential for endothelial homeostasis through the cross‐talk between miRNA let‐7 and miRNA 29 (Figure 5).

Perspective: Translational research of AcSDKP

As described earlier, AcSDKP has the potential to combat fibroproliferative diseases, such as kidney complications in diabetes. Although AcSDKP has not yet been translated into the clinic, recent data have shown that AcSDKP could be a useful clinical biomarker in some disease conditions.

Sodium intake is a known detrimental factor in the clinical outcome of renal diseases99. Kwakernaak et al.100 examined the potential relevance of AcSDKP in renal protection. The researchers enrolled 46 non‐diabetic chronic kidney disease patients with overt proteinuria who showed mild‐to‐moderate renal insufficiency. In a cross‐over design/double‐blind analysis for a 6‐week study period with a regular or a low‐sodium diet and either lisinopril or lisinopril plus valsartan, sodium restriction was confirmed to increase the plasma level of AcSDKP during either single or dual renin–angiotensin system blockade100. Interestingly, a recent report utilizing mice with knockout of the N‐terminal sequence of ACE, which resulted in AcSDKP degradation, found that they showed increased levels of AcSDKP associated with enhanced urine excretion of sodium without alterations in renal angiotensin II levels101. The molecular regulation and physiological relevance of the interaction of AcSDKP levels and sodium intake/excretion have not yet been elucidated, and further research is required.

Finally, the present author recently reported that the urine levels of AcSDKP could be a potential biomarker of alteration of renal function in normoalbuminuric diabetes patients with an estimated glomerular filtration rate (eGFR) ≥30 mL/min/1.73 m2 102. When compared with that observed two decades ago, the diabetes population with renal deficiency without urine albuminuria has increased103, 104. The explanation for these changes is unknown, but evidence‐based treatment of diabetes patients (such as appropriate blood glucose control and renin–angiotensin system blockade) could result in the reduction of urine albumin with a decrease in eGFR103, 104. Urine albumin‐negative diabetes patients rarely show a progressive decline in renal function, few, but some, can progress to end‐stage renal disease104. Most importantly, there is no established biomarker for this urine albumin‐negative diabetes population. The present author and co‐authors have already shown that CD‐1 mice, a fibrotic strain with diabetes, displayed a diabetes‐associated decline in urine AcSDKP levels48.

Based on this background, the present author and co‐authors hypothesized that the urine AcSDKP‐to‐urine creatine ratio can predict kidney injury and future renal function. A total of 21 diabetes patients with normoalbuminuria and an eGFR ≥ 30 mL/min/1.73 m2 were divided into two groups based on the median values: low or high urinary AcSDKP groups (uAcSDKP/Crlow or uAcSDKP/Crhigh) in the first morning urine and follow up at ~4 years to monitor eGFR. In the uAcSDKP/Crhigh group, the alteration in eGFR (ΔeGFRop [ΔeGFR observational periods]) showed significant stability compared with that of the uAcSDKP/Crlow group over time (P = 0.003, χ2 = 8.58). Levels of the urine kidney injury molecule‐1 (uKim‐1), which is known to increase in the injured kidney, were also measured, and in the low uKim‐1 group, ΔeGFRop showed stable kidney function over time compared with that of the high uKim‐1 group (P = 0.004, χ2 = 8.38). There was no significant interaction between the levels of AcSDKP and Kim‐1. Patients who had both uAcSDKP/Crhigh and uKim‐1low were much more likely to show stable ΔeGFRop (P < 0.001, χ2 = 30.4) than other patients. The plasma AcSDKP level (P = 0.015, χ2 = 5.94) can also weakly, but significantly, predict ΔeGFRop. As kidney tubulointerstitial fibrosis is a major determinant of kidney dysfunction, the antifibrotic peptide, AcSDKP, could be a functional biomarker for kidney injury and a future decline in renal function102.

Conclusion

A therapeutic strategy with appropriate molecular biomarkers is essential to prevent kidney disease progression in diabetes. Fibrosis is a pathological process that destroys normal parenchyma; therefore, an antifibrotic strategy based on the mechanisms underlying kidney fibrogenesis could help to inhibit diabetic kidney disease progression. From this point of view, further elucidation of the potential of AcSDKP should be carried out. AcSDKP could be valuable, as both a therapeutic treatment and molecular biomarker in kidney disease, especially for diabetes patients.

Disclosure

KK is an advisory board member of Boehringer Ingelheim and collaborates with a study on a similar topic contained in this manuscript.