A 69-year-old Japanese man developed abdominal pain, purpura, proteinuria, and hematuria while receiving treatment for pulmonary tuberculosis. A skin biopsy revealed IgA-positive leukocytoclastic vasculitis, and a renal biopsy showed IgA-positive mesangial proliferative glomerulonephritis with crescent formation. Based on these findings, we diagnosed IgA vasculitis with nephritis (IgAVN) and initiated treatment. The patient’s abdominal symptoms improved following factor XIII supplementation and corticosteroids. Corticosteroids were administered, and after 5 months, the proteinuria was in complete remission. Although IgAVN often follows a prior infection, it is rarely complicated by tuberculosis. In this case, staining for galactose-deficient IgA1, which is specifically positive in IgA nephropathy and IgAVN, was positive. Nephritis-associated plasmin receptor staining was also positive, suggesting some involvement of infectious glomerulonephritis. Therefore, the patient was considered to have IgAVN associated with pulmonary tuberculosis. In adult-onset cases, IgAVN is often severe. This patient was presented with adult-onset nephrosis and International Study of Kidney Disease in Children grade IIIb IgAVN, suggesting a poor prognosis. Therefore, we immediately initiated treatment with corticosteroids, factor XIII supplementation, a renin-aldosterone-system inhibitor, and a sodium–glucose cotransporter 2 inhibitor. The patient recovered uneventfully with no worsening of tuberculosis.

The progress in the research field of diabetic kidney disease (DKD) has been disturbed by the lack of reliable animal models. Angiotensin II (Ang II) type 1 receptor (AT1R)-associated protein (ATRAP) promotes internalization of AT1R and selectively inhibits pathological AT1R signaling. In this study, we investigated whether overactivation of the renin-angiotensin system (RAS) through a combination of ATRAP deletion with Ang II stimulation developed a progressive DKD model in C57BL/6 mice, which are resistant to the development of kidney injury. Eight-week-old male systemic ATRAP-knockout mice on the C57BL/6 strain (KO) and their littermate wild-type mice (Ctrl) were divided into five groups: 1) Ctrl, 2) Ctrl-streptozotocin (STZ), 3) KO-STZ, 4) Ctrl-STZ-Ang II, and 5) KO-STZ-Ang II. Ang II was administered for 6 weeks from 4 weeks after STZ administration. At 10 weeks after STZ administration, mice were euthanized to evaluate kidney injuries. Neither ATRAP deletion alone nor Ang II stimulation alone developed a progressive DKD model in STZ-induced diabetic C57BL/6 mice. However, a combination of ATRAP deletion with Ang II stimulation accelerated the development of DKD as manifested by overt albuminuria, glomerular hypertrophy, podocyte loss, mesangial expansion, kidney interstitial fibrosis and functional insufficiency, concomitant with increased angiotensinogen and AT1R expression in the kidneys. In STZ-induced diabetic C57BL/6 mice that are resistant to the development of kidney injury, the combination of ATRAP deletion and Ang II stimulation accelerates the development of DKD, which may be associated with intrarenal RAS overactivation.

Although some dietary supplements have been reported to cause renal dysfunction, there have been few reports of supplement-induced Fanconi syndrome. We present the case of a 56-year-old woman with Fanconi syndrome that developed after she consumed a red yeast rice supplement. She was referred to our hospital because of renal dysfunction, and was found to have electrolyte abnormalities, including hypophosphatemia and hypouricemia, renal diabetes, and hyperchloremic metabolic acidosis, and was, therefore, diagnosed with Fanconi syndrome. Renal biopsy revealed proximal tubular injury characterized by severely degenerated tubular epithelial cells as well as mild hypocellular fibrosis. We speculated that the red yeast rice supplement, which the patient had been consuming for approximately 1 year, might be a cause of her syndrome, because reports of renal dysfunction associated with the consumption of red yeast rice supplements have emerged in Japan since 2024. After the supplement was discontinued and oral prednisolone treatment was initiated, the patient’s renal function improved and her electrolyte abnormalities were ameliorated. Furthermore, even after tapering off and discontinuing the prednisolone over approximately 12 weeks, her renal function remained. Because Fanconi syndrome may be caused by various exogenous substances, the taking of a thorough medical history is crucial, including with respect to the use not only of prescription medications, but also other substances, including supplements.

Introduction To date, there is limited evidence on the effects of bronchodilators on respiratory dynamics in chronic obstructive pulmonary disease (COPD). Dynamic chest radiography (DCR) is a novel radiographic modality that provides real-time, objective and quantifiable kinetic data, including changes in the lung area (Rs), tracheal diameter, diaphragmatic kinetics and pulmonary ventilation during respiration, at a lower radiation dose than that used by fluoroscopic or CT imaging. However, the therapeutic effect of dual bronchodilators on respiratory kinetics, such as chest wall dynamics and respiratory muscle function, has not yet been prospectively evaluated using DCR.Aim This study aims to evaluate the effects of bronchodilator therapy on respiratory kinetics in patients with COPD using DCR.Methods and analysis This is an open-label, prospective, single-centre, non-controlled, comparative study. A total of 35 patients with COPD, aged 40–85 years, with a forced expiratory volume in the first second of 30–80%, will be enrolled. After a 2–4 weeks washout period, patients will receive tiotropium/olodaterol therapy for 6 weeks. Treatment effects will be evaluated based on DCR findings, pulmonary function test results and patient-related outcomes obtained before and after treatment. The primary endpoint is the change in Rs after therapy. The secondary endpoints include differences in other DCR parameters (diaphragmatic kinetics, tracheal diameter change and maximum pixel value change rate), pulmonary function test results and patient-related outcomes between pre-therapy and post-therapy values. All adverse events will be reported.Ethics and dissemination Ethical approval for this study was obtained from the Ethics Committee of Chiba University Hospital. The results of this trial will be published in a peer-reviewed journal.Trial registration number jRCTs032210543.

Objective: Angiotensin II type 1 receptor (AT1R)-associated protein (ATRAP) was originally identified as a specific binding protein of AT1R. We have shown that ATRAP promotes constitutive internalization of AT1R so as to inhibit the pathological activation of its downstream signaling. Also, we reported that genetic knockdown of ATRAP exacerbates kidney fibrosis in mice along with functional mitochondrial abnormalities and subsequent increases in ROS production. These effects of ATRAP on kidney fibrosis were suggested to be AT1R-independent actions, but the molecular mechanism is unclear. The present study was designed to explore a novel interacting protein of ATRAP that was involved in the mechanism by which ATRAP protected against kidney fibrosis, independent of the interaction with AT1R. Design and method: We established Human Embryonic Kidney 293 cells which were able to induce the expression of Flag-tagged ATRAP (HEK293_Flag-ATRAP cells) in a doxycycline-dependent manner. After immunoprecipitation with anti-Flag antibodies, the Flag-ATRAP complex was analyzed with a mass spectrometer. Among identified proteins, we focused on transferrin receptor1(TfR1). To confirm the molecular interaction, co-immunoprecipitation was performed using anti-Flag or anti-TfR1 antibodies. Additionally, to validate functional interactions, we analyzed intracellular iron concentrations using fluorescent probe of ferric iron. Furthermore, we verified the TfR1 expression in two ways. First, immunofluorescence staining of TfR1 in the whole cell or on the cell surface only. Second, Western blot with whole cell lysate treated with ferristatin II which degraded TfR1 on the cell surface. Results: On the mass spectrometric analysis, 377 proteins were identified as the ATRAP binding protein candidates. Enrichment analysis indicated that proteins related to endocytosis and vesicle trafficking were enriched. TfR1 is also identified. We confirmed the molecular interaction between ATRAP and TfR1 by co-immunoprecipitation. Enhanced ATRAP expression decreased the cellular iron level as well as the expression of TfR1 on the cell surface, despite no evident change in whole cell. Furthermore, we showed that ATRAP suppressed the effect of ferristatin II. Conclusion: We propose a molecular and functional link between ATRAP and TfR1. ATRAP would regulate TfR1 availability via downregulation of cell surface TfR1 via promotion of its internalization. TfR1 promotes intracellular localization of the ferric iron-bound transferrin. Iron is a key factor in the process of kidney fibrosis via production of ROS in relation to deterioration of mitochondrial function. Taken together, this novel ATRAP-TfR1 axis might be the mechanism relevant to the ROS/mitochondrial dysfunction-mediated process of kidney fibrosis.

Objective: One of the obstacles toward understanding the pathophysiology of diabetic nephropathy (DN) has been the lack of reliable animal models that faithfully replicate features of human DN. Angiotensin II (Ang II) type 1 receptor (AT1R)-associated protein (ATRAP) promotes internalization of AT1R from the cell surface into the cytoplasm, resulting in the suppression of AT1R signaling pathway. We have recently reported that systemic ATRAP deficiency exaggerates streptozotocin (STZ)-induced DN via an activation of renal renin-angiotensin system (RAS) (Haruhara K, et al. Kidney Int 2022). However, STZ-induced diabetic ATRAP knockout mice still exhibited a modest increase in albuminuria with limited pathological changes just in glomerulus. To establish more robust DN models, we examined the effect of Ang II stimulation on the development of DN in STZ-treated ATRAP knockout mice. Design and method: Male eight weeks old C57BL/6 mice (Ctrl) and systemic ATRAP-knockout mice (KO) were divided into three groups: 1) Ctrl-STZ, 2) Ctrl-STZ-Ang II, and 3) KO-STZ-Ang II. Hyperglycemia was induced by intraperitoneal injection of 55 mg/kg STZ for consecutive 5 days. From 4 weeks after STZ administration, Ang II (1000 ng/kg/min) was continuously administered by osmotic mini-pumps for 6 weeks. During the experimental period, body weights and levels of blood glucose were measured every 2 weeks. At 6 and 10 weeks after STZ administration, 24-hour urine samples were collected in metabolic cages, and urinary albumin excretion (UAE) was evaluated by ELISA. At the end of experimental period, mice were euthanized to evaluate renal pathological changes. Results: During the experimental period, body weight gain and levels of blood glucose were not significantly different between the three groups. Nonetheless, although levels of UAE at 6 weeks after STZ administration were still modest and comparable between the three groups, levels of UAE only in the KO-STZ-Ang II group were remarkably increased compared to those in the Ctrl-STZ and Ctrl-STZ-Ang II groups at 10 weeks after STZ administration (679.1 ± 583.7 vs 69.9 ± 10.0 vs 61.3 ± 8.2 μg/day, respectively). Furthermore, in the KO-STZ-Ang II group, exacerbation of mesangial expansion and interstitial fibrosis was observed. Conclusions: In STZ-induced diabetic mice, combination of systemic ATRAP deficiency with Ang II stimulation accelerated the development of DN, suggesting a potential to become a promising mouse model replicating key features of human DN.

The renin-angiotensin system plays a crucial role in the regulation of blood pressure. Activation of the angiotensin II (Ang II)-Ang II type 1 receptor (AT1R) signaling pathway contributes to the pathogenesis of hypertension and subsequent organ damage. AT1R-associated protein (ATRAP) has been identified as an endogenous inhibitory protein of the AT1R pathological activation. We have shown that mouse Atrap (Atrap) represses various Ang II-AT1R-mediated pathologies, including hypertension in mice. The expression of human ATRAP (ATRAP)/Atrap can be altered in various pathological states in humans and mice, such as Ang II stimulation and serum starvation. However, the regulatory mechanisms of ATRAP/Atrap are not yet fully elucidated. miRNAs are 21 to 23 nucleotides of small RNAs that post-transcriptionally repress gene expression. Single miRNA can act on hundreds of target mRNAs, and numerous miRNAs have been identified as the Ang II-AT1R signaling-associated disease phenotype modulator, but nothing is known about the regulation of ATRAP/Atrap. In the present study, we identified miR-125a-5p/miR-125b-5p as the evolutionarily conserved miRNAs that potentially act on ATRAP/Atrap mRNA. Further analysis revealed that miR-125a-5p/miR-125b-5p can directly repress both ATRAP and Atrap. In addition, the inhibition of miR-125a-5p/miR-125b-5p resulted in the suppression of the Ang II-AT1R signaling in mouse distal convoluted tubule cells. Taken together, miR-125a-5p/miR-125b-5p activates Ang II-AT1R signaling by the suppression of ATRAP/Atrap. Our results provide new insights into the potential approaches for achieving the organ-protective effects by the repression of the miR-125 family associated with the enhancement of ATRAP/Atrap expression.

Considering the prevalence of obesity and global aging, the consumption of a high-protein diet (HPD) may be advantageous. However, an HPD aggravates kidney dysfunction in patients with chronic kidney disease (CKD). Moreover, the effects of an HPD on kidney function in healthy individuals are controversial. In this study, we employed a remnant kidney mouse model as a CKD model and aimed to evaluate the effects of an HPD on kidney injury under conditions of non-CKD and CKD. Mice were divided into four groups: a sham surgery (sham) + normal diet (ND) group, a sham + HPD group, a 5/6 nephrectomy (Nx) + ND group and a 5/6 Nx + HPD group. Blood pressure, kidney function and kidney tissue injury were compared after 12 weeks of diet loading among the four groups. The 5/6 Nx groups displayed blood pressure elevation, kidney function decline, glomerular injury and tubular injury compared with the sham groups. Furthermore, an HPD exacerbated glomerular injury only in the 5/6 Nx group; however, an HPD did not cause kidney injury in the sham group. Clinical application of these results suggests that patients with CKD should follow a protein-restricted diet to prevent the exacerbation of kidney injury, while healthy individuals can maintain an HPD without worrying about the adverse effects.

Objective: We previously reported that genetic knockdown of angiotensin II type 1 receptor (AT1R)-associated protein (ATRAP) exacerbated the aging-associated kidney tubulointerstitial fibrosis in mice (Uneda K, et al. J Am Heart Assoc 2017). However, little is known about whether enhancement of ATRAP expression could affect any pathological stimuli-induced kidney fibrosis and inflammation. Recently we proposed that aristolochic acid nephropathy (AAN) might be a useful model of kidney aging along with tubulointerstitial fibrosis (Urate S, et al. Int J Mol Sci 2021). The present study was designed to investigate the functional role of ATRAP in kidney fibrosis and inflammation, using ATRAP transgenic mice subjected to AAN. Design and method: We generated ATRAP transgenic (Tg19) mice under the control of chicken β;-actin promoter. (Wakui H, et al. Am J Physiol Renal Physiol 2010). The level of kidney ATRAP protein expression was about 4-fold higher in the Tg19 mice compared with wild-type littermate control (LC) mice. The Tg19 mice and LC mice were administered either vehicle or aristolochic acid (AA) (3 mg/kg) for 4 weeks, followed by a 4-week remodeling period. Kidney fibrosis was examined by histopathology and macrophage infiltration was determined by immunohistochemistry. Expression levels of genes associated with inflammation, fibrosis, and senescence were determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immuno blot analyses. Results: AA administration provoked kidney fibrosis and inflammation accompanied by decreased kidney function. There were no significant differences in AA-induced kidney fibrosis estimated by qRT-PCR and histological analysis between the Tg19 and LC mice. Creatinine clearance was similarly decreased in the Tg19 and LC mice with AAN. However, AA-induced macrophage infiltration was significantly suppressed in the kidney of Tg19 mice compared with that of the LC mice. In addition, inflammation-, aging-, and oxidative stress-related genes (tumor necrosis factor-α, interleukin-1β;, cyclin-dependent kinase inhibitor 2A, wnt family member 9A, nicotinamide adenine dinucleotide phosphate oxidase 2) in response to AA administration were significantly suppressed in the kidney of the Tg19 mice compared with that of the LC mice. Conclusion: These results indicate that the enhancement of ATRAP expression suppresses kidney inflammation despite no evident effects on kidney fibrosis in a mouse model of AAN. ATRAP may be a therapeutic target for inflammation associated with CKD.

Background and aim Dysregulation of angiotensin II type 1 receptor-associated protein (ATRAP) expression in cardiovascular, kidney, and adipose tissues is involved in the pathology of hypertension, cardiac hypertrophy, atherosclerosis, kidney injury, and metabolic disorders. Furthermore, ATRAP is highly expressed in bone marrow-derived immune cells; however, the functional role of immune cell ATRAP in obesity-related pathology remains unclear. Thus, we sought to identify the pathophysiological significance of immune cell ATRAP in the development of visceral obesity and obesity-related metabolic disorders using a mouse model of diet-induced obesity. Methods Initially, we examined the effect of high-fat diet (HFD)-induced obesity on the expression of immune cell ATRAP in wild-type mice. Subsequently, we conducted bone marrow transplantation to generate two types of chimeric mice: bone marrow wild-type chimeric (BM-WT) and bone marrow ATRAP knockout chimeric (BM-KO) mice. These chimeric mice were provided an HFD to induce visceral obesity, and then the effects of immune cell ATRAP deficiency on physiological parameters and adipose tissue in the chimeric mice were investigated. Results In wild-type mice, body weight increase by HFD was associated with increased expression of immune cell ATRAP. In the bone marrow transplantation experiments, BM-KO mice exhibited amelioration of HFD-induced weight gain and visceral fat expansion with small adipocytes compared BM-WT mice. In addition, BM-KO mice on the HFD showed significant improvements in white adipose tissue metabolism, inflammation, glucose tolerance, and insulin resistance, compared with BM-WT mice on the HFD. Detailed analysis of white adipose tissue revealed significant suppression of HFD-induced activation of transforming growth factor-beta signaling, a key contributor to visceral obesity, via amelioration of CD206+ macrophage accumulation in the adipose tissue of BM-KO mice. This finding suggests a relevant mechanism for the anti-obesity phenotype in BM-KO mice on the HFD. Finally, transcriptome analysis of monocytes indicated the possibility of genetic changes, such as the enhancement of interferon-γ response at the monocyte level, affecting macrophage differentiation in BM-KO mice. Conclusion Collectively, our results indicate that ATRAP in bone marrow-derived immune cells plays a role in the pathogenesis of visceral obesity. The regulation of ATRAP expression in immune cells may be a key factor against visceral adipose obesity with metabolic disorders.

Abstract Aims Angiotensin receptor-neprilysin inhibitor (ARNI) is an established treatment for heart failure. However, whether ARNI has renoprotective effects beyond renin-angiotensin system inhibitors alone in cardiorenal syndrome (CRS) has not been fully elucidated. Here, we examined the effects of ARNI on the heart and kidneys of CRS model mice with overt albuminuria and identified the mechanisms underlying ARNI-induced kidney protection. Methods and results C57BL6 mice were subjected to chronic angiotensin II infusion, nephrectomy, and salt loading (ANS); they developed CRS phenotypes and were divided into the vehicle treatment (ANS-vehicle), sacubitril/valsartan treatment (ANS-ARNI), and two different doses of valsartan treatment (ANS-VAL M, ANS-VAL H) groups. Four weeks after treatment, the hearts and kidneys of each group were evaluated. The ANS-vehicle group showed cardiac fibrosis, cardiac dysfunction, overt albuminuria, and kidney fibrosis. The ANS-ARNI group showed a reduction in cardiac fibrosis and cardiac dysfunction compared with the valsartan treatment groups. However, regarding the renoprotective effects characterized by albuminuria and fibrosis, ARNI was less effective than valsartan. Kidney transcriptomic analysis showed that the ANS-ARNI group exhibited a significant enhancement in the phosphoinositide 3-kinase (PI3K)-AKT signalling pathway compared with the ANS-VAL M group. Adding PI3K inhibitor treatment to ARNI ameliorated kidney injury to levels comparable with those of ANS-VAL M while preserving the superior cardioprotective effect of ARNI. Conclusion PI3K pathway activation has been identified as a key mechanism affecting remnant kidney injury under ARNI treatment in CRS pathology, and blockading the PI3K pathway with simultaneous ARNI treatment is a potential therapeutic strategy for treating CRS with overt albuminuria.

Kidney fibrosis is a common pathway that leads to chronic kidney disease. Angiotensin II type-1 receptor (AT1R)-associated protein (ATRAP) was originally identified as an AT1R-binding protein. Previously, we reported that systemic knockout of ATRAP exacerbates kidney fibrosis in aged mice. Although these effects of ATRAP appeared to be AT1R-independent actions, the molecular mechanism remains poorly understood. To elucidate the molecular mechanism of ATRAP independent of AT1R, we explored novel ATRAP-interacting proteins. Mass spectrometric analysis of the immunoprecipitants of a Flag-tagged ATRAP complex revealed 376 candidate proteins that potentially interact with ATRAP. Gene ontology analysis revealed that proteins related to vesicle trafficking, membrane transport, and many membrane proteins, including transferrin receptor 1 (TfR1), were enriched. Because TfR1 promotes cellular iron uptake and iron is a key factor involved in kidney fibrosis, we focused on TfR1 and confirmed that it interacts with ATRAP. In addition, our findings revealed that enhanced ATRAP expression decreased cell-surface TfR1 expression without altering the overall cellular TfR1 expression levels. Furthermore, enhanced ATRAP expression attenuated cellular iron levels. Together, our results highlight the role of ATRAP as a suppressor of TfR1 that functions by facilitating TfR1 internalization, which affects iron metabolism and oxidative stress signaling.

Abstract Numerous animal models of chronic kidney disease (CKD) have been developed. However, mice are relatively resistant to kidney injury. We aimed to evaluate the effects of high-protein diet (HPD) loading and 5/6 nephrectomy (Nx) in a susceptible strain of mice (129/Sv) over a long-term period. 129/Sv mice were divided into three groups: sham surgery (sham) + normal diet (ND) group, sham + HPD group, and 5/6 Nx + HPD group. Blood pressure, kidney function, and kidney tissue injury were compared longitudinally for 12 weeks among the three groups. The 5/6 Nx + HPD group displayed blood pressure elevation, kidney function decline, severe albuminuria, glomerular injury, and tubular injury compared with the sham + ND and sham + HPD groups. However, there was no significant difference in kidney injuries between the sham + ND and sham + HPD groups. Furthermore, the 5/6 Nx + ND group was added in comparison with the 5/6 Nx + HPD group. The glomerular injury was significantly exacerbated in the 5/6 Nx + HPD group than in the 5/6 Nx + ND group. These results indicate that HPD loading alone has little effect on kidney injury, while it exacerbates glomerular injury in the remnant kidney model.

Urate, Shingo; Wakui, Hiromichi; Tanaka, Shohei; Abe, Eriko; Tsukamoto, Shunichiro; Taguchi, Shinya; Suzuki, Toru; Azushima, Kengo; Tamura, Kouichi Author Information

Tumor necrosis factor (TNF)-α is a potent mediator of inflammation and is involved in the pathophysiology of chronic kidney disease (CKD). However, the effects of TNF-α inhibition on the progression of kidney fibrosis have not been fully elucidated. We examined the effects of TNF-α inhibition by etanercept (ETN) on kidney inflammation and fibrosis in mice with aristolochic acid (AA) nephropathy as a model of kidney fibrosis. C57BL/6 J mice were administered AA for 4 weeks, followed by a 4-week remodeling period. The mice exhibited kidney fibrosis, functional decline, and albuminuria concomitant with increases in renal mRNA expression of inflammation- and fibrosis-related genes. The 8-week ETN treatment partially but significantly attenuated kidney fibrosis and ameliorated albuminuria without affecting kidney function. These findings were accompanied by significant suppression of interleukin (IL)-1β, IL-6, and collagen types I and III mRNA expression. Moreover, ETN tended to reduce the AA-induced increase in interstitial TUNEL-positive cells with a significant reduction in Bax mRNA expression. Renal phosphorylated p38 MAPK was significantly upregulated by AA but was normalized by ETN. These findings indicate a substantial role for the TNF-α pathway in the pathogenesis of kidney fibrosis and suggest that TNF-α inhibition could become an adjunct therapeutic strategy for CKD with fibrosis.