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BACKGROUND: Sepsis is characterized by a dysregulated immune response and metabolic alterations, including decreased high-density lipoprotein cholesterol (HDL-C) levels. HDL exhibits beneficial properties, such as lipopolysaccharides (LPS) scavenging, exerting anti-inflammatory effects and providing endothelial protection. We investigated the effects of CER-001, an engineered HDL-mimetic, in a swine model of LPS-induced acute kidney injury (AKI) and a Phase 2a clinical trial, aiming to better understand its molecular basis in systemic inflammation and renal function. METHODS: We carried out a translational approach to study the effects of HDL administration on sepsis. Sterile systemic inflammation was induced in pigs by LPS infusion. Animals were randomized into LPS (n = 6), CER20 (single dose of CER-001 20 mg/kg; n = 6), and CER20 × 2 (two doses of CER-001 20 mg/kg; n = 6) groups. Survival rate, endothelial dysfunction biomarkers, pro-inflammatory mediators, LPS, and apolipoprotein A-I (ApoA-I) levels were assessed. Renal and liver histology and biochemistry were analyzed. Subsequently, we performed an open-label, randomized, dose-ranging (Phase 2a) study included 20 patients with sepsis due to intra-abdominal infection or urosepsis, randomized into Group A (conventional treatment, n = 5), Group B (CER-001 5 mg/kg BID, n = 5), Group C (CER-001 10 mg/kg BID, n = 5), and Group D (CER-001 20 mg/kg BID, n = 5). Primary outcomes were safety and efficacy in preventing AKI onset and severity; secondary outcomes include changes in inflammatory and endothelial dysfunction markers. RESULTS: CER-001 increased median survival, reduced inflammatory mediators, complement activation, and endothelial dysfunction in endotoxemic pigs. It enhanced LPS elimination through the bile and preserved liver and renal parenchyma. In the clinical study, CER-001 was well-tolerated with no serious adverse events related to study treatment. Rapid ApoA-I normalization was associated with enhanced LPS removal and immunomodulation with improvement of clinical outcomes, independently of the type and gravity of the sepsis. CER-001-treated patients had reduced risk for the onset and progression to severe AKI (stage 2 or 3) and, in a subset of critically ill patients, a reduced need for organ support and shorter ICU length of stay. CONCLUSIONS: CER-001 shows promise as a therapeutic strategy for sepsis management, improving outcomes and mitigating inflammation and organ damage. TRIAL REGISTRATION: The study was approved by the Agenzia Italiana del Farmaco (AIFA) and by the Local Ethic Committee (N° EUDRACT 2020-004202-60, Protocol CER-001- SEP_AKI_01) and was added to the EU Clinical Trials Register on January 13, 2021.
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Lesión Renal Aguda , Sepsis , Humanos , Animales , Porcinos , Lipoproteínas HDL , Apolipoproteína A-I/uso terapéutico , Apolipoproteína A-I/química , Apolipoproteína A-I/farmacología , Lipopolisacáridos , Investigación Biomédica Traslacional , Inflamación , Sepsis/tratamiento farmacológico , Lesión Renal Aguda/tratamiento farmacológico , Mediadores de InflamaciónRESUMEN
The long non-coding RNAs (lncRNA) play an important role in several biological processes, including some renal diseases. Nevertheless, little is known about lncRNA that are expressed in the healthy kidneys and involved in renal cell homeostasis and development, and even less is known about lncRNA involved in the maintenance of human adult renal stem/progenitor cells (ARPCs) that have been shown to be very important for renal homeostasis and repair processes. Through a whole-genome transcriptome screening, we found that the HOTAIR lncRNA is highly expressed in renal progenitors and potentially involved in cell cycle and senescence biological processes. By CRISPR/Cas9 genome editing, we generated HOTAIR knockout ARPC lines and established a key role of this lncRNA in ARPC self-renewal properties by sustaining their proliferative capacity and limiting the apoptotic process. Intriguingly, the HOTAIR knockout led to the ARPC senescence and to a significant decrease in the CD133 stem cell marker expression which is an inverse marker of ARPC senescence and can regulate renal tubular repair after the damage. Furthermore, we found that ARPCs expressed high levels of the α-Klotho anti-aging protein and especially 2.6-fold higher levels compared to that secreted by renal proximal tubular cells (RPTECs). Finally, we showed that HOTAIR exerts its function through the epigenetic silencing of the cell cycle inhibitor p15 inducing the trimethylation of the histone H3K27. Altogether, these results shed new light on the mechanisms of regulation of these important renal cells and may support the future development of precision therapies for kidney diseases.
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ARN Largo no Codificante , Adulto , Humanos , Senescencia Celular/genética , Histonas/metabolismo , Riñón/metabolismo , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Células Madre/metabolismo , Proteínas KlothoRESUMEN
Acute kidney injury (AKI) is a common consequence of sepsis with a mortality rate of up to 40%. The pathogenesis of septic AKI is complex and involves several mechanisms leading to exacerbated inflammatory response associated with renal injury. A large body of evidence suggests that inflammation is tightly linked to AKI through bidirectional interaction between renal and immune cells. Preclinical data from our and other laboratories have identified in complement system activation a crucial mediator of AKI. Partial recovery following AKI could lead to long-term consequences that predispose to chronic dysfunction and may also accelerate the progression of preexisting chronic kidney disease. Recent findings have revealed striking morphological and functional changes in renal parenchymal cells induced by mitochondrial dysfunction, cell cycle arrest via the activation of signaling pathways involved in aging process, microvascular rarefaction, and early fibrosis. Although major advances have been made in our understanding of the pathophysiology of AKI, there are no available preventive and therapeutic strategies in this field. The identification of ideal clinical biomarkers for AKI enables prompt and effective therapeutic strategy that could prevent the progression of renal injury and promote repair process. Therefore, the use of novel biomarkers associated with clinical and functional criteria could provide early interventions and better outcome. Several new drugs for AKI are currently being investigated; however, the complexity of this disease might explain the failure of pharmacological intervention targeting just one of the many systems involved. The hypothesis that blood purification could improve the outcome of septic AKI has attracted much attention. New relevant findings on the role of polymethylmethacrylate-based continuous veno-venous hemofiltration in septic AKI have been reported. Herein, we provide a comprehensive literature review on advances in the pathophysiology of septic AKI and potential therapeutic approaches in this field.
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End-stage renal disease (ESRD) is characterized by deep disorders in both innate and adaptive immune systems that imply unbalance deactivation and immunosuppression. The central, widely recognized factors responsible for this immune dysregulation are uremia, uremic toxin retention, hemodialysis membrane biocompatibility, and related cardiovascular complications. Recently, several studies strengthened the concept that dialysis membranes are not considered as a simple diffusive/adsorptive device but as a platform to personalize a dialysis approach to improve the quality of life of ESRD patients. Therefore, understanding of the molecules associated with altered immune response is crucial and could lead to therapeutically intervention or adaptation of the dialysis procedure itself for the management of immunological dysfunction of ESRD patients. The polymethyl methacrylate (PMMA)-based membrane is characterized by a symmetrical structure with large-sized pores, providing a better hydrophobic and cationic adsorption capacity compared to the other synthetic membranes. Together with hydrophobic interactions, the high adsorption rate of cytokines (i.e., IL-6) can also be enhanced by the size of nano-pores placed on the membrane surface. PMMA membranes exhibit adsorptive properties for a large amount of uremic toxins including p-cresol and indoxyl sulfate, as well as ß2-microglobulin characterized by higher molecular weight, maintaining the diffusive clearance of small molecules like urea with a great biocompatibility. Besides exerting a strong anti-inflammatory effects in line with the improvement of immune responses in patients undergoing dialysis, PMMA also plays a role in modulating adaptive immune response, i.e., can clear blood from soluble CD40, a natural antagonist of the CD40/CD40L signaling that acts inhibiting immunoglobulin production by B cells. This review provides an overview of the main concepts and current understanding of immune dysfunction in hemodialysis and summarizes the recent findings regarding PMMA-based dialysis as potential strategy to restore immune balance in ESRD patients.
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In the past years, indoxyl sulfate has been strongly implicated in kidney disease progression and contributed to cardiovascular morbidity. Moreover, as a result of its elevated albumin affinity rate, indoxyl sulfate is not adequately cleared by extracorporeal therapies. Within this scenario, although LC-MS/MS represents the conventional approach for IS quantification, it requires dedicated equipment and expert skills and does not allow real-time analysis. In this pilot study, we implemented a fast and simple technology designed to determine serum indoxyl sulfate levels that can be integrated into clinical practice. Indoxyl sulfate was detected at the time of enrollment by Tandem MS from 25 HD patients and 20 healthy volunteers. Next, we used a derivatization reaction to transform the serum indoxyl sulfate into Indigo blue. Thanks to the spectral shift to blue, its quantity was measured by the colorimetric assay at a wavelength of 420-450 nm. The spectrophotometric analysis was able to discriminate the levels of IS between healthy subjects and HD patients corresponding to the LC-MS/MS. In addition, we found a strong linear relationship between indoxyl sulfate levels and Indigo levels between the two methods (Tandem MS and spectrophotometry). This innovative method in the assessment of gut-derived indoxyl sulfate could represent a valid tool for clinicians to monitor CKD progression and dialysis efficacy.
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Insuficiencia Renal Crónica , Tóxinas Urémicas , Humanos , Indicán , Diálisis Renal , Cromatografía Liquida , Proyectos Piloto , Espectrometría de Masas en Tándem/métodos , Insuficiencia Renal Crónica/terapiaRESUMEN
Extracellular vesicles (EV) are emerging mediators in several diseases. However, their role in the pathophysiology of antibody-mediated allograft rejection (AMR) has been poorly investigated. Here, we investigated the role of EV isolated from AMR patients in inducing tubular senescence and endothelial to mesenchymal transition (EndMT) and analyzed their miRNA expression profile. By multiplex bead flow cytometry, we characterized the immunophenotype of plasma AMR-derived EV and found a prevalent platelet and endothelial cell origin. In vitro, AMR-derived EV induced tubular senescence by upregulating SA-ß Gal and CDKN1A mRNA. Furthermore, AMR-derived EV induced EndMT. The occurrence of tubular senescence and EndMT was confirmed by analysis of renal biopsies from the same AMR patients. Moreover, AMR-derived EV induced C3 gene upregulation and CFH downregulation in tubular epithelial cells, with C4d deposition on endothelial cells. Interestingly, RNase-mediated digestion of EV cargo completely abrogated tubular senescence and EndMT. By microarray analysis, miR-604, miR-515-3p, miR-let-7d-5p, and miR-590-3p were significantly upregulated in EV from AMR group compared with transplant controls, whereas miR-24-3p and miR-29a-3p were downregulated. Therefore, EV-associated miRNA could act as active player in AMR pathogenesis, unraveling potential mechanisms of accelerated graft senescence, complement activation and early fibrosis that might lead to new therapeutic intervention.
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Vesículas Extracelulares , MicroARNs , Células Endoteliales/metabolismo , Células Epiteliales/metabolismo , Vesículas Extracelulares/metabolismo , Humanos , MicroARNs/genética , ARN Mensajero/metabolismoRESUMEN
BACKGROUND: Data from large cardiac surgery registries have been depicting a downward trend of mortality and morbidities in the last 20 years. However, despite decades of medical evolution, cardiac surgery and cardiopulmonary bypass still provoke a systemic inflammatory response, which occasionally leads to worsened outcome. This article seeks to outline the mechanism of the phenomenon. METHODS: A thorough review of the literature has been performed. Criteria for considering studies for this non-systematic review were as follows: observational and interventional studies investigating the systemic inflammatory response to cardiac surgery, experimental studies describing relevant molecular mechanisms, and essential review studies pertinent to the topic. RESULTS: The intrinsic variability of the inflammatory response to cardiac surgery, together with its heterogenous perception among clinicians, as well as the arduousness to early discriminate high-responder patients from those who will not develop a clinically relevant reaction, concurred to hitherto unconclusive randomized controlled trials. Furthermore, peremptory knowledge about the pathophysiology of maladaptive inflammation following heart surgery is still lacking. CONCLUSIONS: Systemic inflammation following cardiac surgery is a frequent entity that occasionally becomes clinically relevant. Specific genomic differences, age, and other preoperative factors influence the magnitude of the response, which elements display extreme redundancy and pleiotropism that the target of a single pathway cannot represent a silver bullet.
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Procedimientos Quirúrgicos Cardíacos , Puente Cardiopulmonar , Procedimientos Quirúrgicos Cardíacos/efectos adversos , Puente Cardiopulmonar/efectos adversos , Humanos , Inflamación/etiologíaRESUMEN
High-density lipoproteins (HDLs) are a class of blood particles, principally involved in mediating reverse cholesterol transport from peripheral tissue to liver. Omics approaches have identified crucial mediators in the HDL proteomic and lipidomic profile, which are involved in distinct pleiotropic functions. Besides their role as cholesterol transporter, HDLs display anti-inflammatory, anti-apoptotic, anti-thrombotic, and anti-infection properties. Experimental and clinical studies have unveiled significant changes in both HDL serum amount and composition that lead to dysregulated host immune response and endothelial dysfunction in the course of sepsis. Most SARS-Coronavirus-2-infected patients admitted to the intensive care unit showed common features of sepsis disease, such as the overwhelmed systemic inflammatory response and the alterations in serum lipid profile. Despite relevant advances, episodes of mild to moderate acute kidney injury (AKI), occurring during systemic inflammatory diseases, are associated with long-term complications, and high risk of mortality. The multi-faceted relationship of kidney dysfunction with dyslipidemia and inflammation encourages to deepen the clarification of the mechanisms connecting these elements. This review analyzes the multifaced roles of HDL in inflammatory diseases, the renal involvement in lipid metabolism, and the novel potential HDL-based therapies.
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COVID-19/patología , Lipoproteínas HDL/metabolismo , Sepsis/patología , Lesión Renal Aguda/etiología , COVID-19/complicaciones , COVID-19/metabolismo , COVID-19/virología , Colesterol/metabolismo , Proteínas del Sistema Complemento/metabolismo , Humanos , Metabolismo de los Lípidos , Lipoproteínas HDL/química , SARS-CoV-2/aislamiento & purificación , SARS-CoV-2/fisiología , Sepsis/complicaciones , Sepsis/metabolismo , Internalización del VirusRESUMEN
Diabetic nephropathy (DN) is the most frequent cause of end-stage renal disease. Tubulointerstitial accumulation of lysine 63 (K63)-ubiquitinated (Ub) proteins is involved in the progression of DN fibrosis and correlates with urinary miR-27b-3p downregulation. We explored the renoprotective effect of an inhibitor of K63-Ub (NSC697923), alone or in combination with the ACE-inhibitor ramipril, in vitro and in vivo. Proximal tubular epithelial cells and diabetic DBA/2J mice were treated with NSC697923 and/or ramipril. K63-Ub protein accumulation along with α-SMA, collagen I and III, FSP-1, vimentin, p16INK4A expression, SA-α Gal staining, Sirius Red, and PAS staining were measured. Finally, we measured the urinary albumin to creatinine ratio (uACR), and urinary miR-27b-3p expression in mice. NSC697923, both alone and in association with ramipril, in vitro and in vivo inhibited hyperglycemia-induced epithelial to mesenchymal transition by significantly reducing K63-Ub proteins, α-SMA, collagen I, vimentin, FSP-1 expression, and collagen III along with tubulointerstitial and glomerular fibrosis. Treated mice also showed recovery of urinary miR-27b-3p and restored expression of p16INK4A. Moreover, NSC697923 in combination with ramipril demonstrated a trend in the reduction of uACR. In conclusion, we suggest that selective inhibition of K63-Ub, when combined with the conventional treatment with ACE inhibitors, might represent a novel treatment strategy to prevent the progression of fibrosis and proteinuria in diabetic nephropathy and we propose miR-27b-3p as a biomarker of treatment efficacy.
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Diabetes Mellitus Experimental/complicaciones , Nefropatías Diabéticas/prevención & control , Fibrosis/prevención & control , Lisina/química , Nitrofuranos/farmacología , Ramipril/farmacología , Sulfonas/farmacología , Ubiquitinación , Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Animales , Nefropatías Diabéticas/etiología , Nefropatías Diabéticas/metabolismo , Nefropatías Diabéticas/patología , Quimioterapia Combinada , Femenino , Fibrosis/etiología , Fibrosis/metabolismo , Fibrosis/patología , Ratones , Ratones Endogámicos DBARESUMEN
Endothelial dysfunction is a hallmark of LPS-induced acute kidney injury (AKI). Endothelial cells (ECs) acquired a fibroblast-like phenotype and contributed to myofibroblast generation through the endothelial-to-mesenchymal transition (EndMT) process. Of note, human adult renal stem/progenitor cells (ARPCs) enhance the tubular regenerative mechanism during AKI but little is known about their effects on ECs. Following LPS exposure, ECs proliferated, decreased EC markers CD31 and vascular endothelial cadherin, and up-regulated myofibroblast markers, collagen I, and vimentin. The coculture with ARPCs normalized the EC proliferation rate and abrogated the LPS-induced EndMT. The gene expression analysis showed that most of the genes modulated in LPS-stimulated ARPCs belong to cell activation and defense response pathways. We showed that the ARPC-specific antifibrotic effect is exerted by the secretion of CXCL6, SAA4, and BPIFA2 produced after the anaphylatoxin stimulation. Next, we investigated the molecular signaling that underlies the ARPC protective mechanism and found that renal progenitors diverge from differentiated tubular cells and ECs in myeloid differentiation primary response 88-independent pathway activation. Finally, in a swine model of LPS-induced AKI, we observed that activated ARPCs secreted CXCL6, SAA4, and BPIFA2 as a defense response. These data open new perspectives on the treatment of both sepsis- and endotoxemia-induced AKI, suggesting an underestimated role of ARPCs in preventing endothelial dysfunction and novel strategies to protect the endothelial compartment and promote kidney repair.-Sallustio, F., Stasi, A., Curci, C., Divella, C., Picerno, A., Franzin, R., De Palma, G., Rutigliano, M., Lucarelli, G., Battaglia, M., Staffieri, F., Crovace, A., Pertosa, G. B., Castellano, G., Gallone, A., Gesualdo, L. Renal progenitor cells revert LPS-induced endothelial-to-mesenchymal transition by secreting CXCL6, SAA4, and BPIFA2 antiseptic peptides.
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Lesión Renal Aguda/metabolismo , Lesión Renal Aguda/patología , Células Madre Adultas/patología , Quimiocina CXCL6/metabolismo , Células Endoteliales/patología , Proteínas y Péptidos Salivales/metabolismo , Proteína Amiloide A Sérica/metabolismo , Lesión Renal Aguda/genética , Células Madre Adultas/efectos de los fármacos , Células Madre Adultas/metabolismo , Animales , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Células Cultivadas , Técnicas de Cocultivo , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Expresión Génica/efectos de los fármacos , Humanos , Riñón/efectos de los fármacos , Riñón/metabolismo , Riñón/patología , Lipopolisacáridos/toxicidad , Factor 88 de Diferenciación Mieloide/metabolismo , Miofibroblastos/efectos de los fármacos , Miofibroblastos/metabolismo , Miofibroblastos/patología , Regeneración/fisiología , Transducción de Señal/efectos de los fármacos , Sus scrofaRESUMEN
Adult Renal Stem/Progenitor Cells (ARPCs) have been recently identified in the human kidney and several studies show their active role in kidney repair processes during acute or chronic injury. However, little is known about their immunomodulatory properties and their capacity to regulate specific T cell subpopulations. We co-cultured ARPCs activated by triggering Toll-Like Receptor 2 (TLR2) with human peripheral blood mononuclear cells for 5 days and 15 days and studied their immunomodulatory capacity on T cell subpopulations. We found that activated-ARPCs were able to decrease T cell proliferation but did not affect CD8+ and CD4+ T cells. Instead, Tregs and CD3+ CD4- CD8- double-negative (DN) T cells decreased after 5 days and increased after 15 days of co-culture. In addition, we found that PAI1, MCP1, GM-CSF, and CXCL1 were significantly expressed by TLR2-activated ARPCs alone and were up-regulated in T cells co-cultured with activated ARPCs. The exogenous cocktail of cytokines was able to reproduce the immunomodulatory effects of the co-culture with activated ARPCs. These data showed that ARPCs can regulate immune response by inducing Tregs and DN T cells cell modulation, which are involved in the balance between immune tolerance and autoimmunity.
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Células Madre Adultas/metabolismo , Riñón/citología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Biomarcadores , Proliferación Celular , Quimiocinas/metabolismo , Humanos , Inmunomodulación , Inmunofenotipificación , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Receptor Toll-Like 2/metabolismoRESUMEN
During sepsis, the increased synthesis of circulating lipopolysaccharide (LPS)-binding protein (LBP) activates LPS/TLR4 signaling in renal resident cells, leading to acute kidney injury (AKI). Pericytes are the major source of myofibroblasts during chronic kidney disease (CKD), but their involvement in AKI is poorly understood. Here, we investigate the occurrence of pericyte-to-myofibroblast trans-differentiation (PMT) in sepsis-induced AKI. In a swine model of sepsis-induced AKI, PMT was detected within 9 h from LPS injection, as evaluated by the reduction of physiologic PDGFRß expression and the dysfunctional α-SMA increase in peritubular pericytes. The therapeutic intervention by citrate-based coupled plasma filtration adsorption (CPFA) significantly reduced LBP, TGF-ß, and endothelin-1 (ET-1) serum levels, and furthermore preserved PDGFRß and decreased α-SMA expression in renal biopsies. In vitro, both LPS and septic sera led to PMT with a significant increase in Collagen I synthesis and α-SMA reorganization in contractile fibers by both SMAD2/3-dependent and -independent TGF-ß signaling. Interestingly, the removal of LBP from septic plasma inhibited PMT. Finally, LPS-stimulated pericytes secreted LBP and TGF-ß and underwent PMT also upon TGF-ß receptor-blocking, indicating the crucial pro-fibrotic role of TLR4 signaling. Our data demonstrate that the selective removal of LBP may represent a therapeutic option to prevent PMT and the development of acute renal fibrosis in sepsis-induced AKI.
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Lesión Renal Aguda/etiología , Lesión Renal Aguda/metabolismo , Proteínas de Fase Aguda/metabolismo , Proteínas Portadoras/metabolismo , Transdiferenciación Celular , Glicoproteínas de Membrana/metabolismo , Miofibroblastos/metabolismo , Pericitos/metabolismo , Receptor Toll-Like 4/metabolismo , Lesión Renal Aguda/patología , Animales , Biopsia , Transdiferenciación Celular/genética , Células Cultivadas , Modelos Animales de Enfermedad , Endotoxinas/efectos adversos , Fibrosis , Inmunohistoquímica , Modelos Biológicos , Miofibroblastos/citología , PorcinosRESUMEN
Sepsis remains a serious cause of morbidity and mortality in critically ill patients, with limited therapeutic options available. Of the several disorders connected with sepsis, acute kidney injury (AKI) is one of the major complications. The pathophysiology of sepsis-induced AKI is characterized by severe inflammation in renal parenchyma with endothelial dysfunction, intra-glomerular thrombosis and tubular injury. Endothelial dysfunction is regulated by several mechanisms implicated in cellular de-differentiation, such as endothelial-to-mesenchymal transition (EndMT). Gram-negative bacteria and their cell wall component lipopolysaccharides (LPSs) are frequently involved in the pathogenesis of AKI. The host recognition of LPS requires a specific receptor, which belongs to the Toll-like receptor (TLR) family of proteins, called TLR4, and two carrier proteins, namely the LPS-binding protein (LBP) and cluster of differentiation 14 (CD14). In particular, LBP is released as a consequence of Gram-negative infection and maximizes the activation of TLR4 signalling. Recent findings regarding the emerging role of LBP in mediating sepsis-induced AKI, and the possible beneficial effects resulting from the removal of this endogenous adaptor protein, will be discussed in this review.
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Lesión Renal Aguda/etiología , Lesión Renal Aguda/metabolismo , Proteínas de Fase Aguda/metabolismo , Proteínas Portadoras/metabolismo , Lipopolisacáridos/efectos adversos , Glicoproteínas de Membrana/metabolismo , Sepsis/complicaciones , Lesión Renal Aguda/patología , Animales , Humanos , Transducción de SeñalRESUMEN
BACKGROUND: Increasing evidence demonstrates a phenotypic plasticity of endothelial cells (ECs). Endothelial-to-mesenchymal transition (EndMT) contributes to the development of tissue fibrosis. However, the pathogenic factors and signalling pathways regulating this process in ischaemia/reperfusion (I/R) injury are still poorly understood. METHODS: We investigated the possible role of complement in the induction of this endothelial dysfunction in a swine model of renal I/R injury by using recombinant C1 inhibitor in vivo. RESULTS: Here, we showed that I/R injury reduced the density of renal peritubular capillaries and induced tissue fibrosis with generation of CD31(+)/α-SMA(+) and CD31(+)/FPS-1(+) cells indicating EndMT. When we inhibited complement, the process of EndMT became rare, with preserved density of peritubular capillaries and significant reduction in renal fibrosis. When we activated ECs by anaphylatoxins in vitro, C3a and C5a led to altered endothelial phenotype with increased expression of fibroblast markers and decrease expression of specific endothelial markers. The activation of Akt pathway was pivotal for the C3a and C5a-induced EndMT in vitro. In accordance, inhibition of complement in vivo led to the abrogation of Akt signalling, with hampered EndMT and tissue fibrosis. CONCLUSIONS: Our data demonstrate a critical role for complement in the acute induction of EndMT via the Akt pathway. Therapeutic inhibition of these systems may be essential to prevent vascular damage and tissue fibrosis in transplanted kidney.
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Anafilatoxinas/metabolismo , Células Endoteliales/metabolismo , Enfermedades Renales/patología , Riñón/patología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Daño por Reperfusión/complicaciones , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Células Endoteliales/patología , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Fibrosis/complicaciones , Fibrosis/metabolismo , Fibrosis/patología , Humanos , Riñón/metabolismo , Enfermedades Renales/etiología , Enfermedades Renales/metabolismo , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Transducción de Señal , PorcinosRESUMEN
INTRODUCTION: The pathophysiology of endotoxemia-induced acute kidney injury (AKI) is characterized by an intense activation of the host immune system and renal resident cells by lipopolysaccharide (LPS) and derived proinflammatory products. However, the occurrence of renal fibrosis in this setting has been poorly investigated. The aim of the present study was to investigate the possible association between endothelial dysfunction and acute development of tissue fibrosis in a swine model of LPS-induced AKI. Moreover, we studied the possible effects of coupled plasma filtration adsorption (CPFA) in this setting. METHODS: After 9 hours from LPS infusion and 6 hours of CPFA treatment, histologic and biochemical changes were analyzed in pigs. Apoptosis and endothelial dysfunction were assessed on renal biopsies. The levels of LPS-binding protein (LBP) were quantified with enzyme-linked immunosorbent assay (ELISA). Endothelial cells (ECs) were stimulated in vitro with LPS and cultured in the presence of swine sera and were analyzed with FACS and real-time RT-PCR. RESULTS: In a swine model of LPS-induced AKI, we observed that acute tubulointerstitial fibrosis occurred within 9 hours from LPS injection. Acute fibrosis was associated with dysfunctional alpha-smooth muscle actin (α-SMA)+ ECs characterized by active proliferation (Ki-67+) without apoptosis (caspase-3-). LPS led to EC dysfunction in vitro with significant vimentin and N-cadherin expression and increased collagen I mRNA synthesis. Therapeutic intervention by citrate-based CPFA significantly prevented acute fibrosis in endotoxemic animals, by preserving the EC phenotype in both peritubular capillaries and renal arteries. We found that the removal of LBP from plasma was crucial to eliminate the effects of LPS on EC dysfunction, by blocking LPS-induced collagen I production. CONCLUSIONS: Our data indicate that EC dysfunction might be pivotal in the acute development of tubulointerstitial fibrosis in LPS-induced AKI. Selective removal of the LPS adaptor protein LBP might represent a future therapeutic option to prevent EC dysfunction and tissue fibrosis in endotoxemia-induced AKI.
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Lesión Renal Aguda/patología , Células Endoteliales/fisiología , Riñón/patología , Lesión Renal Aguda/inducido químicamente , Proteínas de Fase Aguda , Animales , Proteínas Portadoras , Caspasa 3/metabolismo , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática/métodos , Femenino , Fibrosis , Riñón/irrigación sanguínea , Riñón/fisiopatología , Glicoproteínas de Membrana , PorcinosRESUMEN
Despite recent technical advances in dialysis care over the past decades, the mortality rate of critically ill patients with acute kidney injury (AKI) requiring dialysis and of chronic kidney disease (CKD) remains unacceptably high. Several preclinical studies have increased our knowledge of the principal mechanisms involved in the pathophysiology of AKI and CKD. Additionally, the development of efficient and specific compensatory sorbent systems in renal replacement therapy to remove unwanted compounds has created the possibility to treat renal diseases and their underlying pathological triggers. Recently, several biomedical blood purification materials have been developed to improve the removal of waste and inflammatory compounds, improve the quality of treatment, and reduce the duration of treatment. This chapter is focused on the principal mechanisms involved in AKI and CKD and the current state of the art for blood purification strategies to identify the most feasible solution to reduce immunological dysfunction and waste compound clearance. In this regard, the current literature underlines the high efficacy of polymethyl methacrylate membrane hemofilters to overcome the shortcomings in the efficiency of current methodologies in removing the excess of metabolic waste and inflammatory mediators from blood. The purpose of this chapter is therefore to enhance physicians' knowledge about PMMA.
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Lesión Renal Aguda , Insuficiencia Renal Crónica , Humanos , Polimetil Metacrilato , Diálisis Renal , Insuficiencia Renal Crónica/terapia , Terapia de Reemplazo Renal , Lesión Renal Aguda/terapiaRESUMEN
During the past decades, the gut microbiome emerged as a key player in kidney disease. Dysbiosis-related uremic toxins together with pro-inflammatory mediators are the main factors in a deteriorating kidney function. The toxicity of uremic compounds has been well-documented in a plethora of pathophysiological mechanisms in kidney disease, such as cardiovascular injury (CVI), metabolic dysfunction, and inflammation. Accumulating data on the detrimental effect of uremic solutes in kidney disease supported the development of many strategies to restore eubiosis. Fecal microbiota transplantation (FMT) spread as an encouraging treatment for different dysbiosis-associated disorders. In this scenario, flourishing studies indicate that fecal transplantation could represent a novel treatment to reduce the uremic toxins accumulation. Here, we present the state-of-the-art concerning the application of FMT on kidney disease to restore eubiosis and reverse the retention of uremic toxins.
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Microbioma Gastrointestinal , Insuficiencia Renal Crónica , Humanos , Tóxinas Urémicas , Trasplante de Microbiota Fecal , Insuficiencia Renal Crónica/metabolismo , DisbiosisRESUMEN
Kidney transplantation is the first-choice treatment for end-stage renal disease (ESRD). Kidney transplant recipients (KTRs) are at higher risk of experiencing a life-threatening event requiring intensive care unit (ICU) admission, mainly in the late post-transplant period (more than 6 months after transplantation). Urosepsis and bloodstream infections account for almost half of ICU admissions in this population; in addition, potential side effects related to immunosuppressive treatment should be accounted for cytotoxic and ischemic changes induced by calcineurin inhibitor (CNI), sirolimus/CNI-induced thrombotic microangiopathy and posterior reversible encephalopathy syndrome. Throughout the ICU stay, Acute Kidney Injury (AKI) incidence is common and ranges from 10% to 80%, and up to 40% will require renal replacement therapy. In-hospital mortality can reach 30% and correlates with acute illness severity and admission diagnosis. Graft survival is subordinated to baseline estimated glomerular filtration rate (eGFR), clinical presentation, disease severity and potential drug nephrotoxicity. The present review aims to define the impact of AKI events on short- and long-term outcomes in KTRs, focusing on the epidemiologic data regarding AKI incidence in this subpopulation; the pathophysiological mechanisms underlying AKI development and potential AKI biomarkers in kidney transplantation, graft and patients' outcomes; the current diagnostic work up and management of AKI; and the modulation of immunosuppression in ICU-admitted KTRs.
RESUMEN
Cisplatin is one of the most effective chemotherapeutic agents strongly associated with nephrotoxicity. Tubular adult renal progenitor cells (tARPC) can regenerate functional tubules and participate in the repair processes after cisplatin exposition. This study investigated the molecular mechanisms underlying the protective effect of tARPC on renal epithelium during cisplatin nephrotoxicity. By performing a whole-genome transcriptomic analysis, we found that tARPC, in presence of cisplatin, can strongly influence the gene expression of renal proximal tubular cell [RPTEC] by inducing overexpression of CYP1B1, a member of the cytochrome P450 superfamily capable of metabolizing cisplatin and of hypoxia/cancer-related lncRNAs as MIR210HG and LINC00511. Particularly, tARPC exerted renoprotection and regeneration effects via extracellular vesicles (EV) enriched with CYP1B1 and miR-27b-3p, a well-known CYP1B1 regulatory miRNA. The expression of CYP1B1 by tARPC was confirmed by analyzing biopsies of cisplatin-treated renal carcinoma patients that showed the colocalization of CYP1B1 with the tARPC marker CD133. CYP1B1 was also overexpressed in urinary EV purified from oncologic patients that presented nephrotoxicity episodes after cisplatin treatment. Interestingly CYP1B1 expression significantly correlated with creatinine and eGFR levels. Taken together, our results show that tARPC are able to counteract cisplatin-induced nephrotoxicity via CYP1B1 release through EV. These findings provide a promising therapeutic strategy for nephrotoxicity risk assessment that could be related to abundance of renal progenitors.
Asunto(s)
Cisplatino , Riñón , MicroARNs , Células Madre , Adulto , Humanos , Cisplatino/efectos adversos , Cisplatino/metabolismo , Cisplatino/farmacología , Citocromo P-450 CYP1B1/genética , Citocromo P-450 CYP1B1/metabolismo , Regulación hacia Abajo/genética , Vesículas Extracelulares/genética , Vesículas Extracelulares/metabolismo , Riñón/efectos de los fármacos , Riñón/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Células Madre/metabolismoRESUMEN
BACKGROUND: Immunoglobulin A nephropathy (IgAN) is the most frequent primary glomerulonephritis and the role of IL-6 in pathogenesis is becoming increasingly important. A recent whole genome DNA methylation screening in IgAN patients identified a hypermethylated region comprising the non-coding RNA Vault RNA 2-1 (VTRNA2-1) that could explain the high IL-6 levels. METHODS: The pathway leading to IL-6 secretion controlled by VTRNA2-1, PKR, and CREB was analyzed in peripheral blood mononuclear cells (PBMCs) isolated from healthy subjects (HS), IgAN patients, transplanted patients with or without IgAN. The role of double and single-strand RNA in controlling the pathway was investigated. RESULTS: VTRNA2-1 was downregulated in IgAN compared to HS and in transplanted IgAN patients (TP-IgAN) compared to non-IgAN transplanted (TP). The loss of the VTRNA2-1 natural restrain in IgAN patients caused PKR hyperphosphorylation, and consequently the activation of CREB by PKR, which, in turn, led to high IL-6 production, both in IgAN and in TP-IgAN patients. IL-6 levels could be decreased by the PKR inhibitor imoxin. In addition, PKR is normally activated by bacterial and viral RNA, and we found that both the RNA poly(I:C), and the COVID-19 RNA-vaccine stimulation significantly increased the IL-6 levels in PBMCs from HS but had an opposite effect in those from IgAN patients. CONCLUSION: The discovery of the upregulated VTRNA2-1/PKR/CREB/IL-6 pathway in IgAN patients may provide a novel approach to treating the disease and may be useful for the development of precision nephrology and personalized therapy by checking the VTRNA2-1 methylation level in IgAN patients.