Knockout of the KH-Type Splicing Regulatory Protein Drives Glomerulonephritis in MRL-Faslpr Mice

KH-type splicing regulatory protein (KSRP) is an RNA-binding protein that promotes mRNA decay and thereby negatively regulates cytokine expression at the post-transcriptional level. Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by dysregulated cytokine expression causing multiple organ manifestations; MRL-Faslpr mice are an established mouse model to study lupus disease pathogenesis. To investigate the impact of KSRP on lupus disease progression, we generated KSRP-deficient MRL-Faslpr mice (MRL-Faslpr/KSRP−/− mice). In line with the predicted role of KSRP as a negative regulator of cytokine expression, lupus nephritis was augmented in MRL-Faslpr/KSRP−/− mice. Increased infiltration of immune cells, especially of IFN-γ producing T cells and macrophages, driven by enhanced expression of T cell-attracting chemokines and adhesion molecules, seems to be responsible for worsened kidney morphology. Reduced expression of the anti-inflammatory interleukin-1 receptor antagonist may be another reason for severe inflammation. The increase of FoxP3+ T cells detected in the kidney seems unable to dampen the massive kidney inflammation. Interestingly, lymphadenopathy was reduced in MRL-Faslpr/KSRP−/− mice. Altogether, KSRP appears to have a complex role in immune regulation; however, it is clearly able to ameliorate lupus nephritis.

Chronic activation of AMP-activated protein kinase leads to early onset polycystic kidney phenotype

AMP-activated protein kinase (AMPK) plays a key role in the cellular response to low energy stress and has emerged as an attractive therapeutic target for tackling metabolic diseases. Whilst significant progress has been made regarding the physiological role of AMPK, its function in the kidney remains only partially understood. We use a mouse model expressing a constitutively active mutant of AMPK to investigate the effect of AMPK activation on kidney function in vivo. Kidney morphology and changes in gene and protein expression were monitored and serum and urine markers were measured to assess kidney function in vivo. Global AMPK activation resulted in an early onset polycystic kidney phenotype, featuring collecting duct cysts and compromised renal function in adult mice. Mechanistically, the cystic kidneys had increased cAMP levels and ERK activation, increased hexokinase I expression, glycogen accumulation and altered expression of proteins associated with autophagy. Kidney tubule-specific activation of AMPK also resulted in a polycystic phenotype, demonstrating that renal tubular AMPK activation caused the cystogenesis. Importantly, human ADPKD kidney sections revealed similar protein localisation patterns to that observed in the murine cystic kidneys. Our findings show that early onset chronic AMPK activation leads to a polycystic kidney phenotype, suggesting dysregulated AMPK signalling is a contributing factor in cystogenesis.

Early type 1 diabetes aggravates renal ischemia/reperfusion-induced acute kidney injury

The present study aimed to investigate the interaction between early diabetes and renal IR-induced AKI and to clarify the mechanisms involved. C57BL/6J mice were assigned to the following groups: (1) sham-operated; (2) renal IR; (3) streptozotocin (STZ—55 mg/kg/day) and sham operation; and (4) STZ and renal IR. On the 12th day after treatments, the animals were subjected to bilateral IR for 30 min followed by reperfusion for 48 h, at which time the animals were euthanized. Renal function was assessed by plasma creatinine and urea levels, as well urinary protein contents. Kidney morphology and gene and protein expression were also evaluated. Compared to the sham group, renal IR increased plasma creatinine, urea and albuminuria levels and decreased Nphs1 mRNA expression and nephrin and WT1 protein staining. Tubular injury was observed with increased Havcr1 and Mki67 mRNA expression accompanied by reduced megalin staining. Renal IR also resulted in increased SQSTM1 protein expression and increased proinflammatory and profibrotic factors mRNA expression. Although STZ treatment resulted in hyperglycemia, it did not induce significant changes in renal function. On the other hand, STZ treatment aggravated renal IR-induced AKI by exacerbating renal dysfunction, glomerular and tubular injury, inflammation, and profibrotic responses. Thus, early diabetes constitutes a relevant risk factor for renal IR-induced AKI.

Renal and Extra Renal Manifestations in Adult Zebrafish Model of Cystinosis

Cystinosis is a rare, incurable, autosomal recessive disease caused by mutations in the CTNS gene. This gene encodes the lysosomal cystine transporter cystinosin, leading to lysosomal cystine accumulation in all cells of the body, with kidneys being the first affected organs. The current treatment with cysteamine decreases cystine accumulation, but does not reverse the proximal tubular dysfunction, glomerular injury or loss of renal function. In our previous study, we have developed a zebrafish model of cystinosis through a nonsense mutation in the CTNS gene and have shown that zebrafish larvae recapitulate the kidney phenotype described in humans. In the current study, we characterized the adult cystinosis zebrafish model and evaluated the long-term effects of the disease on kidney and extra renal organs through biochemical, histological, fertility and locomotor activity studies. We found that the adult cystinosis zebrafish presents cystine accumulation in various organs, altered kidney morphology, impaired skin pigmentation, decreased fertility, altered locomotor activity and ocular anomalies. Overall, our data indicate that the adult cystinosis zebrafish model reproduces several human phenotypes of cystinosis and may be useful for studying pathophysiology and long-term effects of novel therapies.

Evaluation of the Therapeutic Effects of Protocatechuic Aldehyde in Diabetic Nephropathy

Diabetic nephropathy (DN) is one of the most severe chronic kidney diseases in diabetes and is the main cause of end-stage renal disease (ESRD). Protocatechuic aldehyde (PCA) is a natural product with a variety of effects on pulmonary fibrosis. In this study, we examined the effects of PCA in C57BL/KS db/db male mice. Kidney morphology, renal function indicators, and Western blot, immunohistochemistry, and hematoxylin and eosin (H&E) staining data were analyzed. The results revealed that treatment with PCA could reduce diabetic-induced renal dysfunction, as indicated by the urine albumin-to-creatinine ratio (db/m: 120.1 ± 46.1μg/mg, db/db: 453.8 ± 78.7 µg/mg, db/db + 30 mg/kg PCA: 196.6 ± 52.9 µg/mg, db/db + 60 mg/kg PCA: 163.3 ± 24.6 μg/mg, p < 0.001). However, PCA did not decrease body weight, fasting plasma glucose, or food and water intake in db/db mice. H&E staining data revealed that PCA reduced glomerular size in db/db mice (db/m: 3506.3 ± 789.3 μm2, db/db: 6538.5 ± 1818.6 μm2, db/db + 30 mg/kg PCA: 4916.9 ± 1149.6 μm2, db/db + 60 mg/kg PCA: 4160.4 ± 1186.5 μm2p < 0.001). Western blot and immunohistochemistry staining indicated that PCA restored the normal levels of diabetes-induced fibrosis markers, such as transforming growth factor-beta (TGF-β) and type IV collagen. Similar results were observed for epithelial–mesenchymal transition-related markers, including fibronectin, E-cadherin, and α-smooth muscle actin (α-SMA). PCA also decreased oxidative stress and inflammation in the kidney of db/db mice. This research provides a foundation for using PCA as an alternative therapy for DN in the future.

Challenging Disease Ontology by Instances of Atypical PKHD1 and PKD1 Genetics

BackgroundAutosomal polycystic kidney disease is distinguished into dominant (ADPKD) and recessive (ARPKD) inheritance usually caused by either monoallelic (PKD1/PKD2) or biallelic (PKHD1) germline variation. Clinical presentations are genotype-dependent ranging from fetal demise to mild chronic kidney disease (CKD) in adults. Additionally, exemptions from dominant and recessive inheritance have been reported in both disorders resulting in respective phenocopies. Here, we comparatively report three young adults with microcystic-hyperechogenic kidney morphology based on unexpected genetic alterations beyond typical inheritance.MethodsNext-generation sequencing (NGS)-based gene panel analysis and multiplex ligation-dependent probe amplification (MLPA) of PKD-associated genes, familial segregation analysis, and reverse phenotyping.ResultsThree unrelated individuals presented in late adolescence for differential diagnosis of incidental microcystic-hyperechogenic kidneys with preserved kidney and liver function. Upon genetic analysis, we identified a homozygous hypomorphic PKHD1 missense variant causing pseudodominant inheritance in a family, a large monoallelic PKDH1-deletion with atypical transmission, and biallelic PKD1 missense hypomorphs with recessive inheritance.ConclusionBy this report, we illustrate clinical presentations associated with atypical PKD-gene alterations beyond traditional modes of inheritance. Large monoallelic PKHD1-alterations as well as biallelic hypomorphs of both PKD1 and PKHD1 may lead to mild CKD in the absence of prominent macrocyst formation and functional liver impairment. The long-term renal prognosis throughout life, however, remains undetermined. Increased detection of atypical inheritance challenges our current thinking of disease ontology not only in PKD but also in Mendelian disorders in general.

Dual Actions of A2A and A3 Adenosine Receptor Ligand Prevents Obstruction-Induced Kidney Fibrosis in Mice

Kidney fibrosis is the final outcome of chronic kidney disease (CKD). Adenosine plays a significant role in protection against cellular damage by activating four subtypes of adenosine receptors (ARs), A1AR, A2AAR, A2BAR, and A3AR. A2AAR agonists protect against inflammation, and A3AR antagonists effectively inhibit the formation of fibrosis. Here, we showed for the first time that LJ-4459, a newly synthesized dual-acting ligand that is an A2AAR agonist and an A3AR antagonist, prevents the progression of tubulointerstitial fibrosis. Unilateral ureteral obstruction (UUO) surgery was performed on 6-week-old male C57BL/6 mice. LJ-4459 (1 and 10 mg/kg) was orally administered for 7 days, started at 1 day before UUO surgery. Pretreatment with LJ-4459 improved kidney morphology and prevented the progression of tubular injury as shown by decreases in urinary kidney injury molecular-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL) excretion. Obstruction-induced tubulointerstitial fibrosis was attenuated by LJ-4459, as shown by a decrease in fibrotic protein expression in the kidney. LJ-4459 also inhibited inflammation and oxidative stress in the obstructed kidney, with reduced macrophage infiltration, reduced levels of pro-inflammatory cytokines, as well as reduced levels of reactive oxygen species (ROS). These data demonstrate that LJ-4459 has potential as a therapeutic agent against the progression of tubulointerstitial fibrosis.

MO036DAPAGLIFLOZIN RESCUES THE RENAL PHENOTYPE OF GLYCOGEN STORAGE DISEASE TYPE IB

Background and Aims Glycogenosis I type b (GsdI-b) is a rare metabolic disease and immune disorder characterized by hepato-renal glycogen accumulation caused by a deficiency in the Glucose-6-phosphate transporter (G6PT). G6PT transports glucose-6-phosphate (G6P) from cytoplasm to endoplasmic reticulum (ER) where a G6Pase catalyses the hydrolysis of G6P in glucose and phosphate. G6PT deficiency lead to impaired glucose homeostasis, myeloid disfunction and long-term risk of hepatocellular adenomas. No causal therapy is so far available for GSDI-b patients besides a dietary approach to control glycemia and the use of Granulocyte Colony-Stimulating Factor (GCSF) to improve neutropenia. Over time, these supports increase the chronicity of GSDI-b with some complications. A mouse model recapitulating the GDSI-b has been recently generated by inducing G6PT suppression after tamoxifen injection. Here, we characterized the renal phenotype of TM-G6PT-/- mice model focusing on the molecular mechanisms that lead to renal dysfunction. Finally, we evaluated the efficiency of Dapagliflozin, a selective inhibitor of SGLT2, on kidney functions in terms of therapeutic effect. Method Machine learning approach to computer based evaluation of renal morphology was used to analyze the renal sections from TM-G6PT-/- treated with or without dapagliflozin. Results: G6PT is expressed in all renal zones and a severe downregulation of G6PT mRNA expression in whole kidney of TM-G6PT-/- mice can be observed. TM-G6PT-/- mice show tubular vacuolization and overall cellular dysfunction of PT due to a high glycogen accumulation. TM-G6PT-/- mice manifest glycosuria, phosphaturia and polyuria associated with a down regulation of main transporters of PT cells. The urine concentrating defect is due to a primarily role of G6PT in CNT/CD cells confirmed by a downregulation of AQP2, main water channel along CD segments. This mouse model recapitulates the human GSD-Ib renal phenotype characterized by a disfunction of PT but also CNT/CD cells. In order to evaluate whether targeting the glucose metabolism would improve the renal phenotype of these mice we limited glucose flux across the apical membrane of PT cells, applying the SGLT2-inhibitor dapagliflozin to reduce new glycogen formation. After one month of treatment, Dapagliflozin prevents glycogen accumulation in TM-G6PT-/- mice and ameliorates the main dysregulated markers of PT function. This finding was paralleled by an improvement of the histological features of kidney morphology in dapagliflozin treated TM-G6PT-/- mice. Conclusion Our data provide evidence that treatment with dapagliflozin ameliorates intracellular glycogen storage and improves the renal functions in TM-G6PT-/- mice.

Use of Anti-Diabetic Agents in Non-Diabetic Kidney Disease: From Bench to Bedside

New drugs were recently developed to treat hyperglycemia in patients with type 2 diabetes mellitus (T2D). However, metformin remains the first-line anti-diabetic agent because of its cost-effectiveness. It has pleiotropic action that produces cardiovascular benefits, and it can be useful in diabetic nephropathy, although metformin-associated lactic acidosis is a hindrance to its use in patients with kidney failure. New anti-diabetic agents, including glucagon-like peptide-1 receptor (GLP-1R) agonists, dipeptidyl peptidase-4 (DPP-4) inhibitors, and sodium-glucose transporter-2 (SGLT-2) inhibitors, also produce cardiovascular or renal benefits in T2D patients. Their glucose-independent beneficial actions can lead to cardiorenal protection via hemodynamic stabilization and inflammatory modulation. Systemic hypertension is relieved by natriuresis and improved vascular dysfunction. Enhanced tubuloglomerular feedback can be restored by SGLT-2 inhibition, reducing glomerular hypertension. Patients with non-diabetic kidney disease might also benefit from those drugs because hypertension, proteinuria, oxidative stress, and inflammation are common factors in the progression of kidney disease, irrespective of the presence of diabetes. In various animal models of non-diabetic kidney disease, metformin, GLP-1R agonists, DPP-4 inhibitors, and SGLT-2 inhibitors were favorable to kidney morphology and function. They strikingly attenuated biomarkers of oxidative stress and inflammatory responses in diseased kidneys. However, whether those animal results translate to patients with non-diabetic kidney disease has yet to be evaluated. Considering the paucity of new agents to treat kidney disease and the minimal adverse effects of metformin, GLP-1R agonists, DPP-4 inhibitors, and SGLT-2 inhibitors, these anti-diabetic agents could be used in patients with non-diabetic kidney disease. This paper provides a rationale for clinical trials that apply metformin, GLP-1R agonists, DPP-4 inhibitors, and SGLT-2 inhibitors to non-diabetic kidney disease.

A novel STING antagonist H151 ameliorates cisplatin-induced acute kidney injury and mitochondrial dysfunction

Stimulator of interferon genes (STING) is an important adaptor in the cytosolic DNA sensing pathways. Recent study found that the deletion of STING ameliorated cisplatin-induced acute kidney injury (AKI), suggesting that STING could serve as a potential target for AKI therapy. Up to now, a series of small-molecule STING inhibitors/antagonists have been identified. However, none of research was performed to explore the role of human STING inhibitors in AKI. Here we investigated the effect of a newly generated covalent antagonist H151 targeting both human and murine STING, in cisplatin-induced AKI. We found that H151 treatment significantly ameliorated cisplatin-induced kidney injury as shown by the improvement of renal function, kidney morphology and renal inflammation. Besides, tubular cell apoptosis and the increased renal tubular injury marker NGAL induced by cisplatin were also effectively attenuated in H151-treated mice. Moreover, the mitochondrial injury caused by cisplatin was also reversed as evidenced by the improved mitochondrial morphology, restored mitochondrial DNA (mtDNA) content, and reversed mitochondrial genes expression. Finally, we observed enhanced mtDNA levels in the plasma of patients receiving platinum-based chemotherapy compared to the healthy controls, which could potentially activate STING signaling. Taken together, these findings suggested that H151 could be a potential therapeutic agent for treating AKI possibly through inhibiting STING-mediated inflammation and mitochondrial injury.