Abstract
Tubulointerstitial inflammation plays an important role in the progression of diabetic nephropathy (DN), and tubular epithelial cells (TECs) are crucial promoters of the inflammatory cascade. Exchange protein activated by cAMP (Epac) has been shown to suppress the angiotensin II (Ang-II)-induced release of inflammatory cytokines in tubular cells. However, the role of Epac in TEC-mediated tubulointerstitial inflammation in DN remains unknown. We found that administering the Epac agonist 8-pCPT-2′-O-Me-cAMP (8-O-cAMP) to db/db mice inhibited tubulointerstitial inflammation characterized by macrophage infiltration and increased inflammatory cytokine release and consequently alleviated tubulointerstitial fibrosis in the kidney. Furthermore, 8-O-cAMP administration restored CCAAT/enhancer binding protein β (C/EBP-β) expression and further upregulated the expression of Suppressor of cytokine signaling 3 (SOCS3), while inhibiting p-STAT3, MCP-1, IL-6, and TNF-α expression in the kidney cortex in db/db mice. And in vitro study showed that macrophage migration and MCP-1 expression induced by high glucose (HG, 30 mM) were notably reduced by 8-O-cAMP in human renal proximal tubule epithelial (HK-2) cells. In addition, 8-O-cAMP treatment restored C/EBP-β expression in HK-2 cells and promoted C/EBP-β translocation to the nucleus, where it transcriptionally upregulated SOCS3 expression, subsequently inhibiting STAT3 phosphorylation. Under HG conditions, siRNA-mediated knockdown of C/EBP-β or SOCS3 in HK-2 cells partially blocked the inhibitory effect of Epac activation on the release of MCP-1. In contrast, SOCS3 overexpression inhibited HG-induced activation of STAT3 and MCP-1 expression in HK-2 cells. These findings indicate that Epac activation via 8-O-cAMP ameliorates tubulointerstitial inflammation in DN through the C/EBP-β/SOCS3/STAT3 pathway.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
References
Zhang L, Long J, Jiang W, Shi Y, He X, Zhou Z, et al. Trends in chronic kidney disease in China. N Engl J Med. 2016;375:905–6.
Lim AK, Tesch GH. Inflammation in diabetic nephropathy. Mediators Inflamm. 2012;2012:146154.
Klessens CQF, Zandbergen M, Wolterbeek R, Bruijn JA, Rabelink TJ, Bajema IM, et al. Macrophages in diabetic nephropathy in patients with type 2 diabetes. Nephrol Dial Transpl. 2017;32:1322–9.
Niu S, Bian Z, Tremblay A. Broad infiltration of macrophages leads to a proinflammatory state in streptozotocin-induced hyperglycemic mice. J Immunol. 2016;197:3293–301.
Tesch GH. Macrophages and diabetic nephropathy. Semin Nephrol. 2010;30:290–301.
Gilbert RE. Proximal tubulopathy: prime mover and key therapeutic target in diabetic kidney disease. Diabetes. 2017;66:791–800.
Lanaspa MA, Ishimoto T, Cicerchi C, Tamura Y, Roncal-Jimenez CA, Chen W, et al. Endogenous fructose production and fructokinase activation mediate renal injury in diabetic nephropathy. J Am Soc Nephrol. 2014;25:2526–38.
Ortiz-Munoz G, Lopez-Parra V, Lopez-Franco O, Fernandez-Vizarra P, Mallavia B, Flores C, et al. Suppressors of cytokine signaling abrogate diabetic nephropathy. J Am Soc Nephrol. 2010;21:763–72.
Yang W, Luo Y, Yang S, Zeng M, Zhang S, Liu J, et al. Ectopic lipid accumulation: potential role in tubular injury and inflammation in diabetic kidney disease. Clin Sci (Lond). 2018;132:2407–22.
Liu W, Chen X, Wang Y, Chen Y, Chen S, Gong W, et al. Micheliolide ameliorates diabetic kidney disease by inhibiting Mtdh-mediated renal inflammation in type 2 diabetic db/db mice. Pharmacol Res. 2019;150:104506.
de Rooij J, Zwartkruis FJ, Verheijen MH, Cool RH, Nijman SM, Wittinghofer A, et al. Epac is a Rap1 guanine-nucleotide-exchange factor directly activated by cyclic AMP. Nature. 1998;396:474–7.
de Rooij J, Rehmann H, van Triest M, Cool RH, Wittinghofer A, Bos JL. Mechanism of regulation of the Epac family of cAMP-dependent RapGEFs. J Biol Chem. 2000;275:20829–36.
Lezoualc’h F, Fazal L, Laudette M, Conte C. Cyclic AMP sensor EPAC proteins and their role in cardiovascular function and disease. Circ Res. 2016;118:881–97.
Robichaux WG, Cheng X. Intracellular cAMP Sensor EPAC: physiology, pathophysiology, and therapeutics development. Physiol Rev. 2018;98:919–1053.
Qin Y, Stokman G, Yan K, Ramaiahgari S, Verbeek F, de Graauw M, et al. cAMP signalling protects proximal tubular epithelial cells from cisplatin-induced apoptosis via activation of Epac. Br J Pharmacol. 2012;165:1137–50.
Parnell E, Palmer TM, Yarwood SJ. The future of EPAC-targeted therapies: agonism versus antagonism. Trends Pharmacol Sci. 2015;36:203–14.
Stokman G, Qin Y, Booij TH, Ramaiahgari S, Lacombe M, Dolman ME, et al. Epac-Rap signaling reduces oxidative stress in the tubular epithelium. J Am Soc Nephrol. 2014;25:1474–85.
Stokman G, Qin Y, Genieser HG, Schwede F, de Heer E, Bos JL, et al. Epac-Rap signaling reduces cellular stress and ischemia-induced kidney failure. J Am Soc Nehprol. 2011;22:859–72.
Ding H, Bai F, Cao H, Xu J, Fang L, Wu J, et al. PDE/cAMP/Epac/C/EBP-beta signaling cascade regulates mitochondria biogenesis of tubular epithelial cells in renal fibrosis. Antioxid Redox Signal. 2018;29:637–52.
Fang F, Liu GC, Kim C, Yassa R, Zhou J, Scholey JW. Adiponectin attenuates angiotensin II-induced oxidative stress in renal tubular cells through AMPK and cAMP-Epac signal transduction pathways. Am J Physiol Ren Physiol. 2013;304:F1366–74.
Xie P, Joladarashi D, Dudeja P, Sun L, Kanwar YS. Modulation of angiotensin II-induced inflammatory cytokines by the Epac1-Rap1A-NHE3 pathway: implications in renal tubular pathobiology. Am J Physiol Ren Physiol. 2014;306:F1260–74.
Yoshimura A, Naka T, Kubo M. SOCS proteins, cytokine signalling and immune regulation. Nat Rev Immunol. 2007;7:454–65.
Jin H, Fujita T, Jin M, Kurotani R, Hidaka Y, Cai W, et al. Epac activation inhibits IL-6-induced cardiac myocyte dysfunction. J Physiol Sci. 2018;68:77–87.
Sands WA, Woolson HD, Yarwood SJ, Palmer TM. Exchange protein directly activated by cyclic AMP-1-regulated recruitment of CCAAT/enhancer-binding proteins to the suppressor of cytokine signaling-3 promoter. Methods Mol Biol. 2012;809:201–14.
Hellstrom M, Harvey AR. Cyclic AMP and the regeneration of retinal ganglion cell axons. Int J Biochem Cell Biol. 2014;56:66–73.
Zhan M, Usman IM, Sun L, Kanwar YS. Disruption of renal tubular mitochondrial quality control by Myo-inositol oxygenase in diabetic kidney disease. J Am Soc Nephrol. 2015;26:1304–21.
Xiao L, Xu X, Zhang F, Wang M, Xu Y, Tang D, et al. The mitochondria-targeted antioxidant MitoQ ameliorated tubular injury mediated by mitophagy in diabetic kidney disease via Nrf2/PINK1. Redox Biol. 2017;11:297–311.
Crowe AR, Yue W. Semi-quantitative determination of protein expression using immunohistochemistry staining and analysis: an integrated protocol. Bio-protoc. 2019;9:e3465.
Xie P, Sun L, Nayak B, Haruna Y, Liu FY, Kashihara N, et al. C/EBP-beta modulates transcription of tubulointerstitial nephritis antigen in obstructive uropathy. J Am Soc Nephrol. 2009;20:807–19.
Shu S, Zhu J, Liu Z, Tang C, Cai J, Dong Z. Endoplasmic reticulum stress is activated in post-ischemic kidneys to promote chronic kidney disease. EBioMedicine. 2018;37:269–80.
You H, Gao T, Cooper TK, Brian Reeves W, Awad AS. Macrophages directly mediate diabetic renal injury. Am J Physiol Ren Physiol. 2013;305:F1719–27.
Awad AS, You H, Gao T, Cooper TK, Nedospasov SA, Vacher J, et al. Macrophage-derived tumor necrosis factor-α mediates diabetic renal injury. Kidney Int. 2015;88:722–33.
Scurt FG, Menne J, Brandt S, Bernhardt A, Mertens PR, Haller H, et al. Systemic inflammation precedes microalbuminuria in diabetes. Kidney Int Rep. 2019;4:1373–86.
Yarwood SJ, Borland G, Sands WA, Palmer TM. Identification of CCAAT/enhancer-binding proteins as exchange protein activated by cAMP-activated transcription factors that mediate the induction of the SOCS-3 gene. J Biol Chem. 2008;283:6843–53.
Tuazon Kels MJ, Ng E, Al Rumaih Z. TNF deficiency dysregulates inflammatory cytokine production, leading to lung pathology and death during respiratory poxvirus infection. Proc Natl Acad Sci USA. 2020;117:15935–46.
Chow FY, Nikolic-Paterson DJ, Ozols E, Atkins RC, Rollin BJ, Tesch GH. Monocyte chemoattractant protein-1 promotes the development of diabetic renal injury in streptozotocin-treated mice. Kidney Int. 2006;69:73–80.
Zhang MH, Feng L, Zhu MM, Gu JF, Jiang J, Cheng XD, et al. The anti-inflammation effect of Moutan Cortex on advanced glycation end products-induced rat mesangial cells dysfunction and High-glucose-fat diet and streptozotocin-induced diabetic nephropathy rats. J Ethnopharmacol. 2014;151:591–600.
Usui HK, Shikata K, Sasaki M, Okada S, Matsuda M, Shikata Y, et al. Macrophage scavenger receptor-a-deficient mice are resistant against diabetic nephropathy through amelioration of microinflammation. Diabetes. 2007;56:363–72.
Boring L, Gosling J, Chensue SW, Kunkel SL, Farese RV Jr., Broxmeyer HE, et al. Impaired monocyte migration and reduced type 1 (Th1) cytokine responses in C-C chemokine receptor 2 knockout mice. J Clin Invest. 1997;100:2552–61.
Eardley KS, Zehnder D, Quinkler M, Lepenies J, Bates RL, Savage CO, et al. The relationship between albuminuria, MCP-1/CCL2, and interstitial macrophages in chronic kidney disease. Kidney Int. 2006;69:1189–97.
Viau A, Bienaime F, Lukas K, Todkar AP, Knoll M, Yakulov TA, et al. Cilia-localized LKB1 regulates chemokine signaling, macrophage recruitment, and tissue homeostasis in the kidney. EMBO J. 2018;37:e98615.
Yang Z, Guo Z, Dong J, Sheng S, Wang Y, Yu L, et al. miR-374a regulates inflammatory response in diabetic nephropathy by targeting MCP-1 expression. Front Pharmacol. 2018;9:900.
Klueh U, Czajkowski C, Ludzinska I, Qiao Y, Frailey J, Kreutzer DL. Impact of CCL2 and CCR2 chemokine/receptor deficiencies on macrophage recruitment and continuous glucose monitoring in vivo. Biosens Bioelectron. 2016;86:262–9.
Feigerlová E, Battaglia-Hsu SF. IL-6 signaling in diabetic nephropathy: from pathophysiology to therapeutic perspectives. Cytokine Growth Factor Rev. 2017;37:57–65.
Mezzano S, Aros C, Droguett A, Burgos ME, Ardiles L, Flores C, et al. NF-kappaB activation and overexpression of regulated genes in human diabetic nephropathy. Nephrol Dial Transpl. 2004;19:2505–12.
Lu TC, Wang ZH, Feng X, Chuang PY, Fang W, Shen Y, et al. Knockdown of Stat3 activity in vivo prevents diabetic glomerulopathy. Kidney Int. 2009;76:63–71.
Said E, Zaitone SA, Eldosoky M, Elsherbiny NM. Nifuroxazide, a STAT3 inhibitor, mitigates inflammatory burden and protects against diabetes-induced nephropathy in rats. Chem Biol Interact. 2018;281:111–20.
Huang JS, Chuang LY, Guh JY, Huang YJ, Hsu MS. Antioxidants attenuate high glucose-induced hypertrophic growth in renal tubular epithelial cells. Am J Physiol Ren Physiol. 2007;293:F1072–82.
Wang X, Shaw S, Amiri F, Eaton DC, Marrero MB. Inhibition of the Jak/STAT signaling pathway prevents the high glucose-induced increase in tgf-beta and fibronectin synthesis in mesangial cells. Diabetes. 2002;51:3505–9.
Tan JC, Rabkin R. Suppressors of cytokine signaling in health and disease. Pediatr Nephrol. 2005;20:567–75.
Sands WA, Woolson HD, Milne GR, Rutherford C, Palmer TM. Exchange protein activated by cyclic AMP (Epac)-mediated induction of suppressor of cytokine signaling 3 (SOCS-3) in vascular endothelial cells. Mol Cell Biol. 2006;26:6333–46.
Marrero MB, Banes-Berceli AK, Stern DM, Eaton DC. Role of the JAK/STAT signaling pathway in diabetic nephropathy. Am J Physiol Ren Physiol. 2006;290:F762–8.
Hwang M, Go Y, Park JH, Shin SK, Song SE, Oh BC, et al. Epac2a-null mice exhibit obesity-prone nature more susceptible to leptin resistance. Int J Obes (Lond). 2017;41:279–88.
Chepurny OG, Kelley GG, Dzhura I, Leech CA, Roe MW, Dzhura E, et al. PKA-dependent potentiation of glucose-stimulated insulin secretion by Epac activator 8-pCPT-2′-O-Me-cAMP-AM in human islets of Langerhans. Am J Physiol Endocrinol Metab. 2010;298:E622–33.
Kai AK, Lam AK, Chen Y, Tai AC, Zhang X, Lai AK, et al. Exchange protein activated by cAMP 1 (Epac1)-deficient mice develop β-cell dysfunction and metabolic syndrome. FASEB J. 2013;27:4122–35.
Kelley GG, Chepurny OG, Schwede F, Genieser HG, Leech CA, Roe MW, et al. Glucose-dependent potentiation of mouse islet insulin secretion by Epac activator 8-pCPT-2′-O-Me-cAMP-AM. Islets. 2009;1:260–5.
Métrich M, Morel E, Berthouze M, Pereira L, Charron P, Gomez AM, et al. Functional characterization of the cAMP-binding proteins Epac in cardiac myocytes. Pharmacol Rep. 2009;61:146–53.
Herfindal L, Nygaard G, Kopperud R, Krakstad C, Døskeland SO, Selheim F. Off-target effect of the Epac agonist 8-pCPT-2′-O-Me-cAMP on P2Y12 receptors in blood platelets. Biochem Biophys Res Commun. 2013;437:603–8.
Acknowledgements
We thank all who contributed to the present study for their time and effort. This work was supported by the National Natural Sciences Foundation of China (No. 81570658).
Author information
Authors and Affiliations
Contributions
WXY, LX: Conceptualization; WXY, LX: Data curation; WXY, YL, LX: Formal analysis; LX: Funding acquisition; WXY, SMZ, YFL, HFW, JLL, XHL, MRZ, YZH: Investigation; WXY, LX: Methodology; LX: Project administration; WXY, LX: Resources; WXY, LX: Software; LX: Supervision; WXY, LX: Validation; WXY, LX: Visualization; WXY: Writing—original draft; YL, FYL, LS, LX: Writing—review and editing.
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing interests.
Rights and permissions
About this article
Cite this article
Yang, Wx., Liu, Y., Zhang, Sm. et al. Epac activation ameliorates tubulointerstitial inflammation in diabetic nephropathy. Acta Pharmacol Sin 43, 659–671 (2022). https://doi.org/10.1038/s41401-021-00689-2
Received:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/s41401-021-00689-2
Keywords
This article is cited by
-
EphrinB2 alleviates tubulointerstitial fibrosis in diabetic kidney disease
Journal of Translational Medicine (2025)
-
Identification of transcription factors related to diabetic tubulointerstitial injury
Journal of Translational Medicine (2023)
