RÉSUMÉ
Ischemia-reperfusion injury (IRI) is a major event in renal transplantation, leading to adverse outcomes. Bone marrow mesenchymal stem cells (BMSCs) are novel promising therapeutics for repairing kidney injuries. The therapeutic efficacy of BMSCs with ISL1 overexpression in renal IRI and its underlying mechanism need to be investigated. The unilateral renal IRI rat model was established to mimic clinical acute kidney injury. Rats were injected with PBS, BMSCs-Scrambled or BMSCs-ISL1 via the tail vein at the timepoint of reperfusion, and then sacrificed after 24 h of reperfusion. The administration of BMSCs-ISL1 significantly improved renal function, inhibited tubular cells apoptosis, inflammation, oxidative stress in rats. In vitro, HKC cells subjected to H2O2 stimulation were pretreated with the conditioned medium (CM) of BMSCs-Scrambled or BMSCs-ISL1. The pretreatment of ISL1-CM attenuated apoptosis and oxidative stress induced by H2O2 in HKC cells. Our proteomic data suggested that haptoglobin (Hp) was one of the secretory proteins in ISL1-CM. Subsequent experiments confirmed that Hp was the important paracrine factor from BMSCs-ISL1 that exerted anti-apoptotic and antioxidant functions. Mechanistically, Hp played a cytoprotective role via the inhibition of ERK signaling pathway, which could be abrogated by Ro 67-7476, the ERK phosphorylation agonist. The results suggested that paracrine action may be the main mechanism for BMSCs-ISL1 to exert protective effects. As an important anti-apoptotic and antioxidant factor in ISL1-CM, Hp may serve as a new therapeutic agent for treating IRI, providing new insights for overcoming the long-term adverse effects of stem cell therapy.
Sujet(s)
Apoptose , Protéines à homéodomaine LIM , Cellules souches mésenchymateuses , Stress oxydatif , Communication paracrine , Lésion d'ischémie-reperfusion , Facteurs de transcription , Animaux , Lésion d'ischémie-reperfusion/métabolisme , Lésion d'ischémie-reperfusion/anatomopathologie , Lésion d'ischémie-reperfusion/thérapie , Stress oxydatif/effets des médicaments et des substances chimiques , Apoptose/effets des médicaments et des substances chimiques , Cellules souches mésenchymateuses/métabolisme , Cellules souches mésenchymateuses/cytologie , Protéines à homéodomaine LIM/métabolisme , Protéines à homéodomaine LIM/génétique , Rats , Mâle , Facteurs de transcription/métabolisme , Facteurs de transcription/génétique , Rat Sprague-Dawley , Rein/métabolisme , Rein/anatomopathologie , Humains , Peroxyde d'hydrogène/pharmacologie , Peroxyde d'hydrogène/métabolisme , Transplantation de cellules souches mésenchymateuses/méthodes , Atteinte rénale aigüe/métabolisme , Atteinte rénale aigüe/anatomopathologie , Atteinte rénale aigüe/thérapie , Milieux de culture conditionnés/pharmacologie , Lignée cellulaireRÉSUMÉ
Optimizing the efficiency of definitive endoderm (DE) differentiation is necessary for the generation of diverse organ-like structures. In this study, we used the small molecule inhibitor saracatinib (SAR) to enhance DE differentiation of human embryonic stem cells and induced pluripotent stem cells. SAR significantly improved DE differentiation efficiency at low concentrations. The interaction between SAR and Focal Adhesion Kinase (FAK) was explored through RNA-seq and molecular docking simulations, which further supported the inhibition of DE differentiation by p-FAK overexpression in SAR-treated cells. In addition, we found that SAR inhibited the nuclear translocation of Yes-associated protein (YAP), a downstream effector of FAK, which promoted DE differentiation. Moreover, the addition of SAR enabled a significant reduction in activin A (AA) from 50 to 10 ng/mL without compromising DE differentiation efficiency. For induction of the pancreatic lineage, 10 ng/ml AA combined with SAR at the DE differentiation stage yielded a comparative number of PDX1+/NKX6.1+ pancreatic progenitor cells to those obtained by 50 ng/ml AA treatment. Our study highlights SAR as a potential modulator that facilitates the cost-effective generation of DE cells and provides insight into the orchestration of cell fate determination.
Sujet(s)
Benzodioxoles , Différenciation cellulaire , Endoderme , Quinazolines , Transduction du signal , Humains , Différenciation cellulaire/effets des médicaments et des substances chimiques , Endoderme/effets des médicaments et des substances chimiques , Endoderme/cytologie , Endoderme/métabolisme , Benzodioxoles/pharmacologie , Transduction du signal/effets des médicaments et des substances chimiques , Quinazolines/pharmacologie , Facteurs de transcription/métabolisme , Cellules souches pluripotentes induites/effets des médicaments et des substances chimiques , Cellules souches pluripotentes induites/métabolisme , Cellules souches pluripotentes induites/cytologie , Protéines adaptatrices de la transduction du signal/métabolisme , Protéines de signalisation YAP/métabolisme , Focal adhesion kinase 1/métabolisme , Focal adhesion kinase 1/génétique , Cellules souches embryonnaires humaines/effets des médicaments et des substances chimiques , Cellules souches embryonnaires humaines/métabolisme , Cellules souches embryonnaires humaines/cytologie , Activines/métabolisme , Simulation de docking moléculaireRÉSUMÉ
Ischemia-reperfusion injury (IRI) is an inevitable pathophysiological phenomenon in kidney transplantation. Necroptosis is an undoubtedly important contributing mechanism in renal IRI. We first screened differentially expressed necroptosis-related genes (DENRGs) from public databases. Eight DENRGs were validated by independent datasets and verified by qRT-PCR in a rat IRI model. We used univariate and multivariate Cox regression analyses to establish a prognostic signature, and graft survival analysis was performed. Immune infiltrating landscape analysis and gene set enrichment analysis (GSEA) were performed to understand the underlying mechanisms of graft loss, which suggested that necroptosis may aggravate the immune response, resulting in graft loss. Subsequently, a delayed graft function (DGF) diagnostic signature was constructed using the Least Absolute Shrinkage and Selection Operator (LASSO) and exhibited robust efficacy in validation datasets. After comprehensively analyzing DENRGs during IRI, we successfully constructed a prognostic signature and DGF predictive signature, which may provide clinical insights for kidney transplant.