The activation of mixed lineage kinase-like (MLKL) by receptor-interacting protein kinase-3 (RIPK3) results in plasma membrane (PM) disruption and a form of regulated necrosis, called necroptosis. ...Here, we show that, during necroptosis, MLKL-dependent calcium (Ca2+) influx and phosphatidylserine (PS) exposure on the outer leaflet of the plasma membrane preceded loss of PM integrity. Activation of MLKL results in the generation of broken, PM “bubbles” with exposed PS that are released from the surface of the otherwise intact cell. The ESCRT-III machinery is required for formation of these bubbles and acts to sustain survival of the cell when MLKL activation is limited or reversed. Under conditions of necroptotic cell death, ESCRT-III controls the duration of plasma membrane integrity. As a consequence of the action of ESCRT-III, cells undergoing necroptosis can express chemokines and other regulatory molecules and promote antigenic cross-priming of CD8+ T cells.
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•Phosphatidylserine is exposed prior to loss of cell integrity during necroptosis•ESCRT-III facilitates the shedding of MLKL-damaged plasma membrane from intact cells•ESCRT-III supports cell survival and functions downstream of active MLKL•ESCRT-III allows necroptotic cells to signal surrounding cells
Cells undergoing necroptosis are not always headed towards death; ESCRT-III helps preserve the plasma membrane in these cells, contributing to survival.
Klotho is a membrane-bound protein predominantly expressed in the kidney, where it acts as a permissive co-receptor for Fibroblast Growth Factor 23. In its shed form, Klotho exerts anti-fibrotic ...effects in several tissues. Klotho-deficient mice spontaneously develop fibrosis and Klotho deficiency exacerbates the disease progression in fibrotic animal models. Furthermore, Klotho overexpression or supplementation protects against fibrosis in various models of renal and cardiac fibrotic disease. These effects are mediated at least partially by the direct inhibitory effects of soluble Klotho on TGFβ1 signaling, Wnt signaling, and FGF2 signaling. Soluble Klotho, as present in the circulation, appears to be the primary mediator of anti-fibrotic effects. Similarly, through inhibition of the TGFβ1, Wnt, FGF2, and IGF1 signaling pathways, Klotho also inhibits tumorigenesis. The Klotho promoter gene is generally hypermethylated in cancer, and overexpression or supplementation of Klotho has been found to inhibit tumor growth in various animal models. This review focuses on the protective effects of soluble Klotho in inhibiting renal fibrosis and fibrosis in distant organs secondary to renal Klotho deficiency. We also discuss the structure-function relationships of Klotho domains and biological effects in the context of potential targeted treatment strategies.
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The endogenous repair process can result in recovery after acute kidney injury (AKI) with adaptive proliferation of tubular epithelial cells, but repair can also lead to fibrosis and progressive ...kidney disease. There is currently limited knowledge about transcriptional regulators regulating these repair programs. Herein we establish the enhancer and super-enhancer landscape after AKI by ChIP-seq in uninjured and repairing kidneys on day two after ischemia reperfusion injury (IRI). We identify key transcription factors including HNF4A, GR, STAT3 and STAT5, which show specific binding at enhancer and super-enhancer sites, revealing enhancer dynamics and transcriptional changes during kidney repair. Loss of bromodomain-containing protein 4 function before IRI leads to impaired recovery after AKI and increased mortality. Our comprehensive analysis of epigenetic changes after kidney injury in vivo has the potential to identify targets for therapeutic intervention. Importantly, our data also call attention to potential caveats involved in use of BET inhibitors in patients at risk for AKI.
Matrix Gla protein (MGP) is a potent inhibitor of vascular calcification (VC) and requires carboxylation by vitamin K to exert calcification inhibition. Chronic kidney disease (CKD) patients undergo ...early vascular aging often involving extensive VC. The present cross-sectional study investigated the association between circulating dp-ucMGP levels, MGP expression in vascular tissue and MGP polymorphisms. In 141 CKD stage 5 patients, CAC score was significantly increased in the highest tertile of dp-ucMGP (p = 0.002), and a high medial VC score was associated with elevated dp-ucMGP levels. MGP vascular expression was associated with increased circulating dp-ucMGP and CAC scores. MGP SNP analysis revealed that patients homozygous for the C allele of the rs1800801 variant had a higher CAC score (median 15 range 0-1312) compared to patients carrying a T allele (median 0 range 0-966 AU). These results indicate that plasma levels of dp-ucMGP are an independent predictor of increased VC in CKD5 patients and correlate with both higher CAC scores and degree of medial calcification. Additionally, high vascular expression of MGP was associated with higher CAC scores and plasma dp-ucMGP levels. Taken together, our results support that MGP is involved in the pathogenesis of VC.
Klotho was discovered as an antiaging gene, and α-Klotho (Klotho) is expressed in multiple tissues with a broad set of biologic functions. Membrane-bound Klotho binds fibroblast growth factor 23 ...(FGF23), but a soluble form of Klotho is also produced by alternative splicing or cleavage of the extracellular domain of the membrane-bound protein. The relative organ-specific contributions to the levels and effects of circulating Klotho remain unknown. We explored these issues by generating a novel mouse strain with Klotho deleted throughout the nephron (Six2-KL(-/-)). Klotho shedding from Six2-KL(-/-) kidney explants was undetectable and the serum Klotho level was reduced by approximately 80% in Six2-KL(-/-) mice compared with wild-type littermates. Six2-KL(-/-) mice exhibited severe growth retardation, kyphosis, and premature death, closely resembling the phenotype of systemic Klotho knockout mice. Notable biochemical changes included hyperphosphatemia, hypercalcemia, hyperaldosteronism, and elevated levels of 1,25-dihydroxyvitamin D and Fgf23, consistent with disrupted renal Fgf23 signaling. Kidney histology demonstrated interstitial fibrosis and nephrocalcinosis in addition to absent dimorphic tubules. A direct comparative analysis between Six2-KL(-/-) and systemic Klotho knockout mice supports extensive, yet indistinguishable, extrarenal organ manifestations. Thus, our data reveal the kidney as the principal contributor of circulating Klotho and Klotho-induced antiaging traits.
Diabetic kidney disease (DKD) is the most common cause of severe renal disease, and few treatment options are available today that prevent the progressive loss of renal function. DKD is characterized ...by altered glomerular filtration and proteinuria. A common observation in DKD is the presence of renal steatosis, but the mechanism(s) underlying this observation and to what extent they contribute to disease progression are unknown. Vascular endothelial growth factor B (VEGF-B) controls muscle lipid accumulation through regulation of endothelial fatty acid transport. Here, we demonstrate in experimental mouse models of DKD that renal VEGF-B expression correlates with the severity of disease. Inhibiting VEGF-B signaling in DKD mouse models reduces renal lipotoxicity, re-sensitizes podocytes to insulin signaling, inhibits the development of DKD-associated pathologies, and prevents renal dysfunction. Further, we show that elevated VEGF-B levels are found in patients with DKD, suggesting that VEGF-B antagonism represents a novel approach to treat DKD.
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•Targeting VEGF-B signaling to reduce renal lipotoxicity prevents DKD•The beneficial effect is due to re-sensitizing podocytes to insulin signaling•Renal VEGF-B levels are increased in both experimental models and subjects with DKD•Reducing VEGF-B signaling may be a therapeutic strategy for the treatment of DKD
Human studies have highlighted that hyperglycemia may not be the underlying cause of diabetic kidney disease (DKD). Falkevall et al. highlight a role for VEGF-B in renal lipotoxicity in mouse models and patients, offering a potential novel therapeutic approach to DKD.
Sclerostin, an osteocyte-derived inhibitor of bone formation, is linked to mineral bone disorder. In order to validate its potential as a predictor of vascular calcification, we explored associations ...of circulating sclerostin with measures of calcification in 89 epigastric artery biopsies from patients with end-stage renal disease. Significantly higher sclerostin levels were found in the serum of patients with epigastric and coronary artery calcification (calcification score 100 or more). In Spearman’s rank correlations, sclerostin levels significantly associated with age, intact parathyroid hormone, bone-specific alkaline phosphatase, and percent calcification. Multivariable regression showed that age, male gender, and sclerostin each significantly associated with the presence of medial vascular calcification. Receiver operating characteristic curve analysis showed that sclerostin (AUC 0.68) predicted vascular calcification. Vascular sclerostin mRNA and protein expressions were low or absent, and did not differ between calcified and non-calcified vessels, suggesting that the vasculature is not a major contributor to circulating levels. Thus, high serum sclerostin levels associate with the extent of vascular calcification as evaluated both by coronary artery CT and scoring of epigastric artery calcification. Among circulating biomarkers of mineral bone disorder, only sclerostin predicted vascular calcification.
Osteocytes within the mineralized bone matrix control bone remodeling by regulating osteoblast and osteoclast activity. Osteocytes express the aging suppressor Klotho, but the functional role of this ...protein in skeletal homeostasis is unknown. Here we identify Klotho expression in osteocytes as a potent regulator of bone formation and bone mass. Targeted deletion of Klotho from osteocytes led to a striking increase in bone formation and bone volume coupled with enhanced osteoblast activity, in sharp contrast to what is observed in Klotho hypomorphic (kl/kl) mice. Conversely, overexpression of Klotho in cultured osteoblastic cells inhibited mineralization and osteogenic activity during osteocyte differentiation. Further, the induction of chronic kidney disease with high-turnover renal osteodystrophy led to downregulation of Klotho in bone cells. This appeared to offset the skeletal impact of osteocyte-targeted Klotho deletion. Thus, our findings establish a key role of osteocyte-expressed Klotho in regulating bone metabolism and indicate a new mechanism by which osteocytes control bone formation.
Klotho acts as a co-receptor for and dictates tissue specificity of circulating FGF23. FGF23 inhibits PTH secretion, and reduced Klotho abundance is considered a pathogenic factor in renal secondary ...hyperparathyroidism. To dissect the role of parathyroid gland resident Klotho in health and disease, we generated mice with a parathyroid-specific Klotho deletion (PTH-KL(-/-)). PTH-KL(-/-) mice had a normal gross phenotype and survival; normal serum PTH and calcium; unaltered expression of the PTH gene in parathyroid tissue; and preserved PTH response and sensitivity to acute changes in serum calcium. Their PTH response to intravenous FGF23 delivery or renal failure did not differ compared to their wild-type littermates despite disrupted FGF23-induced activation of the MAPK/ERK pathway. Importantly, calcineurin-NFAT signaling, defined by increased MCIP1 level and nuclear localization of NFATC2, was constitutively activated in PTH-KL(-/-) mice. Treatment with the calcineurin-inhibitor cyclosporine A abolished FGF23-mediated PTH suppression in PTH-KL(-/-) mice whereas wild-type mice remained responsive. Similar results were observed in thyro-parathyroid explants ex vivo. Collectively, we present genetic and functional evidence for a novel, Klotho-independent, calcineurin-mediated FGF23 signaling pathway in parathyroid glands that mediates suppression of PTH. The presence of Klotho-independent FGF23 effects in a Klotho-expressing target organ represents a paradigm shift in the conceptualization of FGF23 endocrine action.
Renal Klotho controls mineral metabolism by directly modulating tubular reabsorption of phosphate and calcium and by acting as a co-receptor for the phosphaturic and vitamin D-regulating hormone ...fibroblast growth factor-23 (FGF23). Klotho null mice have a markedly abnormal phenotype. We sought to determine effects of renal-specific and partial deletion of Klotho to facilitate investigation of its roles in health and disease. We generated a mouse model with partial deletion of Klotho in distal tubular segments (Ksp-KL(-/-)). In contrast to Klotho null mice, Ksp-KL(-/-) mice were fertile, had a normal gross phenotype, and did not have vascular or tubular calcification on renal histology. However, Ksp-KL(-/-) mice were hyperphosphatemic with elevated FGF23 levels and abundant expression of the sodium-phosphate cotransporter Npt2a at the brush border membrane. Serum calcium and 1,25-dihydroxyvitamin D(3) levels were normal but parathyroid hormone levels were decreased. TRPV5 protein was reduced with a parallel mild increase in urinary calcium excretion. Renal expression of vitamin D regulatory enzymes and vitamin D receptor was higher in Ksp-KL(-/-) mice than controls, suggesting increased turnover of vitamin D metabolites and a functional increase in vitamin D signaling. There was a threshold effect of residual renal Klotho expression on FGF23: deletion of >70% of Klotho resulted in FGF23 levels 30-250 times higher than in wild-type mice. A subgroup of Ksp-KL(-/-) mice with normal phosphate levels had elevated FGF23, suggesting a Klotho-derived renal-bone feedback loop. Taken together, renal FGF23-Klotho signaling, which is disrupted in CKD, is essential for homeostatic control of mineral metabolism.
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