Osteocrin (OSTN), a bone-derived humoral factor, was reported to act on heart and bone by potentiating the natriuretic peptide (NP) system. Ostn gene polymorphisms have been associated with renal function decline, but its pathophysiological role in the kidney remains unclear.The role of endogenous OSTN was investigated using systemic Ostn-knockout (KO) mice. As a model for OSTN administration, liver-specific Ostn-overexpressing mice crossed with KO (KO-Tg) were generated. These mice were subjected to unilateral ischemia-reperfusion injury (IRI) and renal lesions after 21 days of insult were evaluated. A comprehensive analysis of the Wnt/β-catenin pathway was performed using a polymerase chain reaction (PCR) array. Reporter plasmid-transfected proximal tubular cells (NRK52E) were used to investigate the mechanism by which OSTN affects the pathway.After injury, KO mice showed marginal worsening of renal fibrosis compared with wild-type mice, with comparable renal atrophy. KO-Tg mice showed significantly ameliorated renal atrophy, fibrosis and tubular injury, together with reduced expressions of fibrosis- and inflammation-related genes. The PCR array showed that the activation of the Wnt/β-catenin pathway was attenuated in KO-Tg mice. The downstream targets Mmp7, Myc and Axin2 showed similar results. MMP7 and Wnt2 were induced in corticomedullary proximal tubules after injury, but not in KO-Tg. In NRK52E, OSTN significantly potentiated the inhibitory effects of NP on transforming growth factor β1-induced activation of the Wnt/β-catenin pathway, which was reproduced by a cyclic guanosine monophosphate analog.Ectopic Ostn overexpression ameliorated subsequent renal injury following ischemia-reperfusion. OSTN could represent possible renoprotection in acute to chronic kidney disease transition, thus serving as a potential therapeutic strategy.

AXIN2

Renal ischemia

10.1093/ndt/gfab286

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Novel Drug Screening Strategy to Explore Exosome-Targeted Intervention of Intraglomerular Cross-Talk for Diabetic Kidney Disease

Journal of the American Society of Nephrology (2023)

Daisuke Fujimoto Shuro Umemoto Takashige Kuwabara Ryosuke Date Teruhiko Mizumoto

Fujimoto, Daisuke; Umemoto, Shuro; Kuwabara, Takashige; Date, Ryosuke; Mizumoto, Teruhiko; Kakizoe, Yutaka; Izumi, Yuichiro; Mukoyama, Masashi Author Information

Exosome

10.1681/asn.20233411s1834c

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POS-018 A NOVEL ATP-RETAINING COMPOUND, KUS121, ENHANCES ER-ASSOCIATED DEGRADATION AND EXERTS RENOPROTECTIVE EFFECTS AGAINST ACUTE KIDNEY INJURY

Kidney International Reports (2022)

Yusuke Hata Takashige Kuwabara Daisuke Fujimoto Ryosuke Date Shuro Umemoto

Pathophysiology

10.1016/j.ekir.2022.01.026

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Calprotectin Plays an Important Role in the Pathogenesis of Vascular Calcification in CKD

Journal of the American Society of Nephrology (2024)

Jingxuan Zhang Takashige Kuwabara Ryosuke Date Daisuke Fujimoto Tomoko Kanki

Pathogenesis

10.1681/asn.2024tfsexsq3

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Role of Exosome-Mediated Intraglomerular Cross-Talk Between Mesangial Cells and Macrophages in the Progression of Diabetic Kidney Disease

Journal of the American Society of Nephrology (2022)

Daisuke Fujimoto Shuro Umemoto Takashige Kuwabara Ryosuke Date Yusuke Hata

Fujimoto, Daisuke; Umemoto, Shuro; Kuwabara, Takashige; Date, Ryosuke; Hata, Yusuke; Mizumoto, Teruhiko; Izumi, Yuichiro; Mukoyama, Masashi Author Information

Exosome

Mesangial cell

Kidney Glomerulus

10.1681/asn.20223311s1107a

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A novel VCP modulator KUS121 exerts renoprotective effects in ischemia-reperfusion injury with retaining ATP and restoring ERAD-processing capacity

AJP Renal Physiology (2022)

Yusuke Hata Ryosuke Date Daisuke Fujimoto Hanako Ohashi Ikeda Shuro Umemoto

Acute kidney injury (AKI) is a life-threatening condition and often progresses to chronic kidney disease or the development of other organ dysfunction even after recovery. Despite the increased recognition and high prevalence of AKI worldwide, there has been no established treatment so far. The aim of this study was to investigate the renoprotective effect of Kyoto University substance 121 (KUS121), a novel valosin-containing protein modulator, on AKI. In in vitro experiments, we evaluated cell viability and ATP levels of proximal tubular cells with or without KUS121 under endoplasmic reticulum (ER) stress conditions. In in vivo experiments, the effects of KUS121 were examined in mice with AKI caused by ischemia-reperfusion injury. ER-associated degradation (ERAD)-processing capacity was evaluated by quantification of the ERAD substrate CD3delta-YFP. KUS121 protected proximal tubular cells from cell death under ER stress. The apoptotic response was mitigated as indicated by the suppression of C/EBP homologous protein expression and caspase-3 cleavage, with maintained intracellular ATP levels by KUS121 administration. KUS121 treatment suppressed the elevation of serum creatinine and neutrophil gelatinase-associated lipocalin levels and attenuated renal tubular damage after ischemia-reperfusion. The expression of inflammatory cytokines in the kidney was also suppressed in the KUS121-treated group. Valosin-containing protein expression levels were not altered by KUS121 both in vitro and in vivo. KUS121 treatment restored ERAD-processing capacity associated with potentiation of its upstream pathway, phosphorylated inositol-requiring enzyme-1α, and spliced X box-binding protein-1. In conclusion, these findings indicate that KUS121 can protect renal tubular cells from ER stress-induced injury, suggesting that KUS121 could be a novel and promising therapeutic compound for ischemia-associated AKI.NEW & NOTEWORTHY Novel findings of this study are as follows: 1) Kyoto University substance 121 (KUS121), a novel valosin-containing protein (VCP) modulator, can reduce ATP consumption of VCP; 2) KUS121 reduced endoplasmic reticulum (ER) stress and improved cell viability in proximal tubular cells; 3) KUS121 exerted renoprotective effects against ischemia-reperfusion injury; and 4) KUS121 may prevent ischemic acute kidney injury with ATP retention and restoring ER-associated degradation capacity.

Endoplasmic-reticulum-associated protein degradation

Viability assay

10.1152/ajprenal.00392.2021

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PS-B08-2: EXOSOME-MEDIATED INTERCELLULAR CROSSTALK BETWEEN MESANGIAL CELLS AND MACROPHAGES PARTICIPATES IN THE PROGRESSION OF DIABETIC KIDNEY DISEASE.

Journal of Hypertension (2023)

Daisuke Fujimoto Shuro Umemoto Takashige Kuwabara Ryosuke Date Yusuke Hata

Background: Extracellular vesicles (EVs) are important mediators of intercellular communication and can play a key role in the regulation of pathophysiological processes. Particularly, exosome-mediated intercellular crosstalk has been addressed in several disorders such as cancer and lifestyle-related diseases including hypertensive and diabetic vascular diseases. In diabetic kidney disease (DKD), it has been reported that macrophages infiltrate the mesangial region and may play an important role through local inflammation in glomeruli. Design and method: In this study, we focused on exosome as a factor that acts in a paracrine manner in glomeruli and examined the effects of mesangial cell-derived exosomes cultured under high-glucose conditions on macrophages. In order to identify new therapeutic agents, we screened a validated compound library that can efficiently inhibit this mechanism and also studied their effects on DKD. Results: Exosomes released from mesangial cells induced inflammation in macrophages, as indicated by the NFκB transcriptional activity and TNFα and IL-1β mRNA expression. In addition, the effect was significantly enhanced in exosomes from mesangial cells cultured under high-glucose conditions compared to low-glucose conditions. We also observed that fluorescent-labeled (DiO) exosomes were endocytosed by macrophages in vitro and in vivo. Next, we conducted drug screening using a validated compound library to find compounds that could specifically and effectively inhibit the inflammation in macrophages induced by exosomes. The screening was divided into four steps, and we succeeded in narrowing down the list to 30 candidate compounds from a total of 1,364 compounds. Finally, an HSP90 inhibitor, alvespimycin, was identified as a compound with a strong inhibitory effect on both exosome uptake and the NFκB transcriptional activity. Treatment of a diabetic rat model with alvespimycin significantly reduced proteinuria, and showed a trend toward suppression of mesangial expansion. Conclusions: We found that mesangial cell-derived exosomes are important for inducing the local inflammation by intercellular crosstalk between mesangial cells and macrophages in DKD. Furthermore, alvespimycin, one of the HSP90 inhibitors obtained by drug screening, can effectively ameliorate the disease progression, suggesting that this mechanism could become a novel therapeutic target for DKD.

Exosome

Crosstalk

Mesangial cell

10.1097/01.hjh.0000916612.45016.0d

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Identification and verification of novel therapeutic agents for diabetic kidney disease based on exosome-targeted high-throughput chemical screening

Research Square (Research Square) (2024)

Daisuke Fujimoto Shuro Umemoto Teruhiko Mizumoto Tomoko Kanki Yusuke Hata

Abstract Exosomes are important mediators of intercellular communication and play key roles in the regulation of pathophysiological processes. In diabetic kidney disease (DKD), it has been reported that macrophages recruited in the mesangial region may play pathogenic roles through inducing local inflammation in glomeruli. We focused on exosome-mediated crosstalk between mesangial cells (MC) and macrophages as a novel therapeutic target for DKD. Exosomes released from MC induced inflammation in macrophages and the effect was enhanced under high-glucose conditions. For discovering novel therapeutic agents which can inhibit such exosome-mediated mechanisms, drug repositioning is considered as aneffective tool. We established a unique screening strategy and screened agents to aim at maximizing their specificity and potency to inhibit exosomal mechanisms, along with minimizing their toxicity. We succeeded in identifying alvespimycin, an HSP90 inhibitor. Treatment of diabetic rats with alvespimycin significantly suppressed mesangial expansion, inflammatory gene activation including macrophage markers, and proteinuria. The inhibitory effect on exosome uptake was specific to alvespimycin compared with other known HSP90 inhibitors. MC-derived exosomes are crucial for inflammation by intercellular crosstalk between MC and macrophages in DKD, and alvespimycin effectively ameliorated the progression of DKD by suppressing exosome-mediated actions, suggesting that exosome-targeted agents can be a novel therapeutic strategy.

Identification

Exosome

High-Throughput Screening

10.21203/rs.3.rs-4010567/v1

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