RESUMEN
Despite the well-known hepatoprotective role of the epidermal growth factor receptor (EGFR) pathway upon acute damage, its specific actions during chronic liver disease, particularly cholestatic injury, remain ambiguous and unresolved. Here, we analyzed the consequences of inactivating EGFR signaling in the liver on the regenerative response following cholestatic injury. For that, transgenic mice overexpressing a dominant negative mutant human EGFR lacking tyrosine kinase activity (ΔEGFR) in albumin-positive cells were submitted to liver damage induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC), an experimental model resembling human primary sclerosing cholangitis. Our results show an early activation of EGFR after 1-2 days of a DDC-supplemented diet, followed by a signaling switch-off. Furthermore, ΔEGFR mice showed less liver damage and a more efficient regeneration following DDC injury. Analysis of the mechanisms driving this effect revealed an enhanced activation of mitogenic/survival signals, AKT and ERK1/2-MAPKs, and changes in cell turnover consistent with a quicker resolution of damage in response to DDC. These changes were concomitant with profound differences in the profile of intrahepatic immune cells, consisting of a shift in the M1/M2 balance towards M2 polarity, and the Cd4/Cd8 ratio in favor of Cd4 lymphocytes, overall supporting an immune cell switch into a pro-restorative phenotype. Interestingly, ΔEGFR livers also displayed an amplified ductular reaction, with increased expression of EPCAM and an increased number of CK19-positive ductular structures in portal areas, demonstrating an overexpansion of ductular progenitor cells. In summary, our work supports the notion that hepatocyte-specific EGFR activity acts as a key player in the crosstalk between parenchymal and non-parenchymal hepatic cells, promoting the pro-inflammatory response activated during cholestatic injury and therefore contributing to the pathogenesis of cholestatic liver disease. © 2022 The Pathological Society of Great Britain and Ireland.
Asunto(s)
Hepatopatías , Regeneración Hepática , Albúminas/metabolismo , Albúminas/farmacología , Animales , Descarboxilasas de Aminoácido-L-Aromático/metabolismo , Descarboxilasas de Aminoácido-L-Aromático/farmacología , Molécula de Adhesión Celular Epitelial/metabolismo , Molécula de Adhesión Celular Epitelial/farmacología , Receptores ErbB/metabolismo , Hepatocitos/patología , Humanos , Hígado/patología , Hepatopatías/patología , Regeneración Hepática/fisiología , Ratones , Ratones Transgénicos , Proteínas Tirosina Quinasas , Proteínas Proto-Oncogénicas c-akt/metabolismoRESUMEN
Ubiquitylation of receptor tyrosine kinases (RTKs) regulates both the levels and functions of these receptors. The neurotrophin receptor TrkB (also known as NTRK2), a RTK, is ubiquitylated upon activation by brain-derived neurotrophic factor (BDNF) binding. Although TrkB ubiquitylation has been demonstrated, there is a lack of knowledge regarding the precise repertoire of proteins that regulates TrkB ubiquitylation. Here, we provide mechanistic evidence indicating that ubiquitin carboxyl-terminal hydrolase 8 (USP8) modulates BDNF- and TrkB-dependent neuronal differentiation. USP8 binds to the C-terminus of TrkB using its microtubule-interacting domain (MIT). Immunopurified USP8 deubiquitylates TrkB in vitro, whereas knockdown of USP8 results in enhanced ubiquitylation of TrkB upon BDNF treatment in neurons. As a consequence of USP8 depletion, TrkB levels and its activation are reduced. Moreover, USP8 protein regulates the differentiation and correct BDNF-dependent dendritic formation of hippocampal neurons in vitro and in vivo We conclude that USP8 positively regulates the levels and activation of TrkB, modulating BDNF-dependent neuronal differentiation.This article has an associated First Person interview with the first author of the paper.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo , Receptor trkB , Factor Neurotrófico Derivado del Encéfalo/genética , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Células Cultivadas , Endopeptidasas , Complejos de Clasificación Endosomal Requeridos para el Transporte , Hipocampo/metabolismo , Humanos , Glicoproteínas de Membrana , Neuronas/metabolismo , Receptor trkB/genética , Receptor trkB/metabolismo , Transducción de Señal , Ubiquitina Tiolesterasa/genéticaRESUMEN
Ubiquitination of the TrkA neurotrophin receptor in response to NGF is critical in the regulation of TrkA activation and functions. TrkA is ubiquitinated, among other E3 ubiquitin ligases, by Nedd4-2. To understand mechanistically how TrkA ubiquitination is regulated, we performed a siRNA screening to identify deubiquitinating enzymes and found that USP36 acts as an important regulator of TrkA activation kinetics and ubiquitination. However, USP36 action on TrkA was indirect because it does not deubiquitinate TrkA. Instead, USP36 binds to Nedd4-2 and regulates the association of TrkA and Nedd4-2. In addition, depletion of USP36 increases TrkA·Nedd4-2 complex formation, whereas USP36 expression disrupts the complex, resulting in an enhancement or impairment of Nedd4-2-dependent TrkA ubiquitination, respectively. Moreover, USP36 depletion leads to enhanced total and surface TrkA expression that results in increased NGF-mediated TrkA activation and signaling that augments PC12 cell differentiation. USP36 actions extend beyond TrkA because the presence of USP36 interferes with Nedd4-2-dependent Kv7.2/3 channel regulation. Our results demonstrate that USP36 binds to and regulates the actions of Nedd4-2 over different substrates affecting their expression and functions.
Asunto(s)
Diferenciación Celular/fisiología , Complejos de Clasificación Endosomal Requeridos para el Transporte/metabolismo , Regulación de la Expresión Génica/fisiología , Canal de Potasio KCNQ2/biosíntesis , Canal de Potasio KCNQ3/biosíntesis , Células-Madre Neurales/metabolismo , Receptor trkA/metabolismo , Ubiquitina Tiolesterasa/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Animales , Complejos de Clasificación Endosomal Requeridos para el Transporte/genética , Células HEK293 , Humanos , Canal de Potasio KCNQ2/genética , Canal de Potasio KCNQ3/genética , Ratones , Ubiquitina-Proteína Ligasas Nedd4 , Células-Madre Neurales/citología , Células PC12 , Unión Proteica , Ratas , Receptor trkA/genética , Ubiquitina Tiolesterasa/genética , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
The development of the nervous system is a temporally and spatially coordinated process that relies on the proper regulation of the genes involved. Neurotrophins and their receptors are directly responsible for the survival and differentiation of sensory and sympathetic neurons; however, it is not fully understood how genes encoding Trk neurotrophin receptors are regulated. Here, we show that rat Bex3 protein specifically regulates TrkA expression by acting at the trkA gene promoter level. Bex3 dimerization and shuttling to the nucleus regulate the transcription of the trkA promoter under basal conditions and also enhance nerve growth factor (NGF)-mediated trkA promoter activation. Moreover, qChIP assays indicate that Bex3 associates with the trkA promoter within a 150 bp sequence, immediately upstream from the transcription start site, which is sufficient to mediate the effects of Bex3. Consequently, the downregulation of Bex3 using shRNA increases neuronal apoptosis in NGF-dependent sensory neurons deprived of NGF and compromises PC12 cell differentiation in response to NGF. Our results support an important role for Bex3 in the regulation of TrkA expression and in NGF-mediated functions through modulation of the trkA promoter.
Asunto(s)
Proteínas Reguladoras de la Apoptosis/fisiología , Diferenciación Celular/fisiología , Factor de Crecimiento Nervioso/farmacología , Multimerización de Proteína/fisiología , Receptor trkA/biosíntesis , Transcripción Genética/fisiología , Animales , Diferenciación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Células Cultivadas , Femenino , Células HEK293 , Humanos , Masculino , Ratones , Factor de Crecimiento Nervioso/fisiología , Neuronas/efectos de los fármacos , Neuronas/fisiología , Multimerización de Proteína/efectos de los fármacos , Ratas , Transcripción Genética/efectos de los fármacosRESUMEN
This multicenter study sponsored by GETH-TC aimed to investigate the incidence and predictors of early (within the first 100 days) and late cardiac events (CE) (ECE and LCE) following allo-HCT in AML patients treated with anthracyclines, focusing on exploring the impact of PTCY on cardiac complications and the impact of CE on overall survival (OS) and non-relapse mortality (NRM). 1020 AML patients were included. PTCY was given to 450 (44.1%) adults. Overall, 94 (9.2) patients experienced CE and being arrythmias, pericardial complications, and heart failure the most prevalent ones. ECE occurred in 49 (4.8%) patients in a median of 13 days after allo-HCT, while LCE were diagnosed in 45 (4.4%) patients in a median of 3.6 years after transplant. Using PTCY increased the risk for ECE in multivariate analysis (HR 2.86, P=0.007), but did not not significantly affect the risk for LCE (HR 1.06, P=0.892). The impact of variables on outcomes revealed was investigated using multivariate regression analyses and revealed that the diagnosis of CE significantly decreased the likelihood of OS (HR 1.66, P=0.005) and increased the likelihood of NRM (HR 2.88, P<0.001). Furthermore, despite using PTCY increased the risk for ECE, its administration was found to be beneficial for OS (HR 0.71, P=0.026). The study suggests that while the incidence of CE was relatively low, it significantly impacted mortality. Standard doses of PTCY increased ECE risk but were associated with improved OS. Therefore, implementing protocols to prevent cardiac complications is recommended, considering the widespread adoption of PTCY in allo-HCT.