Loss of NLRP6 increases the severity of kidney fibrosis.

While NLRP3 contributes to kidney fibrosis, the function of most NOD-like receptors (NLRs) in chronic kidney disease (CKD) remains unexplored. To identify further NLR members involved in the pathogenesis of CKD, we searched for NLR genes expressed by normal kidneys and differentially expressed in human CKD transcriptomics databases. For NLRP6, lower kidney expression correlated with decreasing glomerular filtration rate. The role and molecular mechanisms of Nlrp6 in kidney fibrosis were explored in wild-type and Nlrp6-deficient mice and cell cultures. Data mining of single-cell transcriptomics databases identified proximal tubular cells as the main site of Nlrp6 expression in normal human kidneys and tubular cell Nlrp6 was lost in CKD. We confirmed kidney Nlrp6 downregulation following murine unilateral ureteral obstruction. Nlrp6-deficient mice had higher kidney p38 MAPK activation and more severe kidney inflammation and fibrosis. Similar results were obtained in adenine-induced kidney fibrosis. Mechanistically, profibrotic cytokines transforming growth factor beta 1 (TGF-ß1) and TWEAK decreased Nlrp6 expression in cultured tubular cells, and Nlrp6 downregulation resulted in increased TGF-ß1 and CTGF expression through p38 MAPK activation, as well as in downregulation of the antifibrotic factor Klotho, suggesting that loss of Nlrp6 promotes maladaptive tubular cell responses. The pattern of gene expression following Nlrp6 targeting in cultured proximal tubular cells was consistent with maladaptive transitions for proximal tubular cells described in single-cell transcriptomics datasets. In conclusion, endogenous constitutive Nlrp6 dampens sterile kidney inflammation and fibrosis. Loss of Nlrp6 expression by tubular cells may contribute to CKD progression.

Inhibition of BRD4 Attenuates ER Stress-induced Renal Ischemic-Reperfusion Injury.

Renal ischemia-reperfusion injury (IRI) leads to endoplasmic reticulum (ER) stress, thereby initiating the unfolded protein response (UPR). When sustained, this response may trigger the inflammation and tubular cell death that acts to aggravate the damage. Here, we show that knockdown of the BET epigenetic reader BRD4 reduces the expression of ATF4 and XBP1 transcription factors under ER stress activation. BRD4 is recruited to the promoter of these highly acetylated genes, initiating gene transcription. Administration of the BET protein inhibitor, JQ1, one hour after renal damage induced by bilateral IRI, reveals reduced expression of ATF4 and XBP1 genes, low KIM-1 and NGAL levels and recovery of the serum creatinine and blood urea nitrogen levels. To determine the molecular pathways regulated by ATF4 and XBP1, we performed stable knockout of both transcription factors using CRISPR-Cas9 and RNA sequencing. The pathways triggered under ER stress were mainly XBP1-dependent, associated with an adaptive UPR, and partially regulated by JQ1. Meanwhile, treatment with JQ1 downmodulated most of the pathways regulated by ATF4 and related to the pathological processes during exacerbated UPR activation. Thus, BRD4 inhibition could be useful for curbing the maladaptive UPR activation mechanisms, thereby ameliorating the progression of renal disease.

RIPK3 and kidney disease / RIPK3 y nefropatía

Receptor interacting protein kinase 3 (RIPK3) is an intracellular kinase at the crossroads of cell death and inflammation. RIPK3 contains a RIP homotypic interaction motif (RHIM) domain which allows interactions with other RHIM-containing proteins and a kinase domain that allows phosphorylation of target proteins. RIPK3 may be activated through interaction with RHIM-containing proteins such as RIPK1, TRIF and DAI (ZBP1, DLM-1) or through RHIM-independent mechanisms in an alkaline intracellular pH. RIPK3 mediates necroptosis and promotes inflammation, independently of necroptosis, through either activation of NFκB or the inflammasome. There is in vivo preclinical evidence of the contribution of RIPK3 to both acute kidney injury (AKI) and chronic kidney disease (CKD) and to the AKI-to-CKD transition derived from RIPK3 deficient mice or the use of small molecule RIPK3 inhibitors. In these studies, RIPK3 targeting decreased inflammation but kidney injury improved only in some contexts. Clinical translation of these findings has been delayed by the potential of some small molecule inhibitors of RIPK3 kinase activity to trigger apoptotic cell death by inducing conformational changes of the protein. A better understanding of the conformational changes in RIPK3 that trigger apoptosis, dual RIPK3/RIPK1 inhibitors or repurposing of multiple kinase inhibitors such as dabrafenib may facilitate clinical development of the RIPK3 inhibition concept for diverse inflammatory diseases, including kidney diseases (AU)

La proteína quinasa 3 que interactúa con el receptor (RIPK3) es una quinasa intracelular que se encuentra a medio camino entre la muerte celular y la inflamación. La RIPK3 contiene un dominio motivo de interacción homotípica de RIP (RHIM), que permite las interacciones con otras proteínas que contienen RHIM, y un dominio de quinasa que permite la fosforilación de las proteínas diana. La RIPK3 puede ser activada a través de la interacción con las proteínas que contienen RHIM tales como RIPK1, TRIF y DAI (ZBP1, DLM-1), o a través de mecanismos independientes de RHIM en un pH intracelular alcalino. La RIPK3 media en la necroptosis y promueve la inflamación, independientemente de la necroptosis, bien a través de la activación de NFκB, o del inflamasoma. Existe evidencia preclínica in vivo de la contribución de RIPK3 a la insuficiencia renal aguda (IRA) y la enfermedad renal crónica (ERC), así como a la transición IRA-ERC derivada de ratones con deficiencia de RIPK3 o del uso de pequeñas moléculas inhibidoras de RIPK3. En dichos estudios, el tener a RIPK3 como objetivo redujo la inflamación, pero la nefropatía mejoró solo en algunos contextos. La traducción clínica de estos hallazgos se ha demorado debido al potencial de ciertas pequeñas moléculas inhibidoras de la actividad de la quinasa RIPK3 para activar la muerte celular induciendo cambios conformacionales de la proteína. Comprender mejor los cambios conformacionales de RIPK3 activadores de la apoptosis, los inhibidores duales RIPK3/RIPK1 o la reconversión de múltiples inhibidores de la quinasa tales como dabrafenib podría facilitar el desarrollo clínico del concepto de la inhibición de RIPK3 para diversas enfermedades inflamatorias, incluyendo las enfermedades renales (AU)

Ferroptosis and kidney disease / Ferroptosis y nefropatía

Cell death is a finely regulated process occurring through different pathways. Regulated cell death, either through apoptosis or regulated necrosis offers the possibility of therapeutic intervention. Necroptosis and ferroptosis are among the best studied forms of regulated necrosis in the context of kidney disease. We now review the current evidence supporting a role for ferroptosis in kidney disease and the implications of this knowledge for the design of novel therapeutic strategies. Ferroptosis is defined functionally, as a cell modality characterized by peroxidation of certain lipids, constitutively suppressed by GPX4 and inhibited by iron chelators and lipophilic antioxidants. There is functional evidence of the involvement of ferroptosis in diverse forms of kidneys disease. In a well characterized nephrotoxic acute kidney injury model, ferroptosis caused an initial wave of death, triggering an inflammatory response that in turn promoted necroptotic cell death that perpetuated kidney dysfunction. This suggests that ferroptosis inhibitors may be explored as prophylactic agents in clinical nephrotoxicity or ischemia-reperfusion injury such as during kidney transplantation. Transplantation offers the unique opportunity of using anti-ferroptosis agent ex vivo, thus avoiding bioavailability and in vivo pharmacokinetics and pharmacodynamics issues

La muerte celular es un proceso minuciosamente regulado que se desarrolla a través de diferentes vías. La muerte celular regulada, ya sea mediante apoptosis o necrosis regulada, ofrece la posibilidad de introducir una intervención terapéutica. La necroptosis y la ferroptosis se encuentran entre las formas mejor estudiadas de necrosis regulada en el contexto de la nefropatía. Revisamos los datos actuales que avalan que la ferroptosis desempeña una función en la nefropatía y las repercusiones que tiene este conocimiento en el diseño de nuevas estrategias terapéuticas. La ferroptosis se define de forma funcional como una modalidad celular caracterizada por la peroxidación de ciertos lípidos, constitutivamente suprimida por GPX4 e inhibida por quelantes férricos y antioxidantes lipofílicos. Existen datos probatorios funcionales de la implicación de la ferroptosis en diversas formas de nefropatía. En un modelo de lesión renal aguda nefrotóxica bien caracterizado, la ferroptosis provocó una ola inicial de muerte, la cual desencadenó una respuesta inflamatoria que a su vez promovió la muerte celular necroptótica que perpetuó la disfunción renal. Esto sugiere que los inhibidores de la ferroptosis pueden explorarse como agentes profilácticos en la nefrotoxicidad clínica o en la lesión por isquemia-reperfusión, como durante un trasplante de riñón. Los trasplantes ofrecen una oportunidad única para el uso de agentes inhibidores de la ferroptosis ex vivo, con lo que se evitarían los problemas de biodisponibilidad y los problemas de farmacocinética y farmacodinámica in vivo

Acute kidney injury transcriptomics unveils a relationship between inflammation and ageing

There are no pathophysiolgical therapeutic approaches to acute kidney injury (AKI) and the mortality remains high. In addition chronic kidney disease (CKD) predisposes to AKI and AKI contributes to progression of CKD. Recently a transcriptomics approach unveiled a relationship between AKI, inflammation and the regulation of ageing. A transcriptomics analysis of experimental AKI revealed increased kidney expression of Fn14 and transmembrane chemokine CXCL16, as well as a decreased expression of the kidney-secreted anti-ageing hormone Klotho. Fn14 is the receptor for tumor necrosis factor-like weak inducer of apoptosis (TWEAK), a member of the TNF superfamily. In AKI kidneys there was a positive correlation between Fn14 and CXCL16 mRNA expression and an inverse correlation between Fn14 and Klotho mRNA. Tubular cells were the site of Fn14, CXCL16 and Klotho expression in vivo. Research on the relationships between these three molecules disclosed that TWEAK activation of Fn14 promoted inflammation through secretion of chemokines such as CXL16 in tubular cells in culture and in vivo. Furthermore, TWEAK activation of Fn14 decreased expression of Klotho mRNA and protein in culture and in vivo. Interestingly, both TWEAK activation of CXCL16 mRNA transcription and suppression of Klotho mRNA transcription were mediated by the NFκB transcription factor. In conclusion, TWEAK engagement of Fn14 is a central event promoting NFκB-mediated activation of inflammation pathways and suppression of anti-inflammatory/anti-ageing pathways. This information may influence future therapeutic approaches to AKI and inflammation/aging (AU)

No existen estrategias terapéuticas y fisiopatológicas para el fracaso renal agudo (FRA), por lo que los niveles de mortalidad continúan siendo elevados. Además, la enfermedad renal crónica (ERC) predispone a sufrir FRA y el FRA, a su vez, contribuye a que la ERC avance. Recientemente, una estrategia transcriptómica reveló una relación entre el FRA, la inflamación y la regulación del envejecimiento. Un análisis transcriptómico de modelos experimentales de FRA reveló un aumento de la expresión renal de Fn14 y la quimiocina transmembrana CXCL16, así como un descenso en la expresión de la hormona Klotho antienvejecimiento secretada por el riñón. Fn14 es el receptor de la citoquina tumor necrosis factor-like weak inducer of apoptosis (TWEAK), miembro de la superfamilia de factor de necrosis tumoral. En los riñones con FRA, existía una correlación positiva entre Fn14 y la expresión de ARNm de CXCL16 y una correlación inversa entre Fn14 y el ARNm de Klotho. El lugar donde se da la expresión in vivo de Fn14, CXCL16 y Klotho es las células tubulares. La investigación en las relaciones entre estas tres moléculas reveló que la activación de Fn14 por TWEAK provocó la inflamación mediante la secreción de quimiocinas como la CXCL16 en células tubulares, tanto en cultivo como in vivo. Además, la activación de Fn14 por TWEAK disminuyó la expresión de ARNm de Klotho y de proteína, en cultivo y in vivo. Curiosamente, tanto la activación TWEAK de la trascripción de ARNm de CXCL16 y la supresión de la trascripción de ARNm de Klotho estuvieron mediadas por el factor de transcripción NF-κB. Como conclusión, la unión de TWEAK y Fn14 es un elemento clave en promover de la activación mediada por NF-kB de las vías de inflamación y en la supresión de las vías antiinflamatorias y antienvejecimiento. Esta información puede influir en las futuras estrategias terapéuticas para el FRA y la inflamación/envejecimiento (AU)