RESUMEN
Bietti's crystalline dystrophy (BCD) is an incurable retinal disorder caused by the polypeptide 2 of cytochrome P450 family 4 subfamily V (CYP4V2) mutations. Patients with BCD present degeneration of retinal pigmented epithelial (RPE) cells and consequent blindness. The lack of appropriate disease models and patients' RPE cells limits our understanding of the pathological mechanism of RPE degeneration. In this study, using CYP4V2 mutant pluripotent stem cells as disease models, we demonstrated that RPE cells with CYP4V2 mutations presented a disrupted fatty acid homeostasis, which were characterized with excessive accumulation of poly-unsaturated fatty acid (PUFA), including arachidonic acid (AA) and eicosapentaenoic acid (EPA). The PUFA overload increased mitochondrial reactive oxygen species, impaired mitochondrial respiratory functions, and triggered mitochondrial stress-activated p53-independent apoptosis in CYP4V2 mutant RPE cells. Restoration of the mutant CYP4V2 using adeno-associated virus 2 (AAV2) can effectively reduce PUFA deposition, alleviate mitochondria oxidative stresses, and rescue RPE cell death in BCD RPE cells. Taken together, our results highlight a role of PUFA-induced mitochondrial damage as a central node to potentiate RPE degeneration in BCD patients. AAV2-mediated gene therapy may represent a feasible strategy for the treatment of BCD.
Asunto(s)
Distrofias Hereditarias de la Córnea/metabolismo , Células Epiteliales/metabolismo , Ácidos Grasos Insaturados/farmacología , Mitocondrias/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Células Madre Pluripotentes/metabolismo , Degeneración Retiniana/metabolismo , Enfermedades de la Retina/metabolismo , Epitelio Pigmentado de la Retina/metabolismo , Animales , Células Cultivadas , Distrofias Hereditarias de la Córnea/genética , Distrofias Hereditarias de la Córnea/patología , Familia 4 del Citocromo P450/deficiencia , Familia 4 del Citocromo P450/genética , Células Epiteliales/patología , Femenino , Técnicas de Inactivación de Genes , Humanos , Ratones , Ratones SCID , Mitocondrias/metabolismo , Mutación , Células Madre Pluripotentes/efectos de los fármacos , Degeneración Retiniana/genética , Degeneración Retiniana/patología , Enfermedades de la Retina/genética , Enfermedades de la Retina/patología , Epitelio Pigmentado de la Retina/patologíaRESUMEN
BACKGROUND/AIMS: Cyp4a14 is a member of cytochrome P450 (Cyp450) enzyme superfamily that possesses NADPH monooxygenase activity, which catalyzes omega-hydroxylation of medium-chain fatty acids and arachidonic acid. Study suggests that down-regulation of Cyp4a14 has an anti-inflammatory response in intestine. The present study was to test the function of Cyp4a14 in dextran sulfate sodium (DSS)-induced colitis. METHODS: Female Cyp4a14-knockout (KO) and wild-type (WT) mice were treated with DSS for 6 days to induce colitis. The colon of mice was histologically observed by hematoxylin and eosin (H&E) and periodic acid Schiff (PAS) staining. The serum malondialdehyde (MDA), an endogenous indicator of oxidative stress, was chemically measured. Proinflammatory and NADPH oxidase genes were examined by quantitative polymerase chain reaction (qPCR). RESULTS: Cyp4a14-KO mice had a significantly higher number of goblet cells in the colon and were more resistant to DSS-induced colitis compared with the WT mice. The DSS-treated KO mice had lower levels of MDA. Consistent with the milder inflammatory pathological changes, DSS-treated KO mice had lower levels of IL-1ß, IL-6 and TNF-α mRNA in the liver and the colon. Moreover, the colon of DSS-treated Cyp4a14-KO and WT mice had higher mRNA levels of two members of NADPH oxidases, Nox2 and Nox4, suggesting that both Nox2 and Nox4 are inflammatory markers. By contrast, DSS-treated WT and KO mice had drastically decreased epithelium-localized Nox1 and dual oxidase (Duox) 2 mRNA levels, coinciding with the erosion of the mucosa induced by DSS. CONCLUSION: These results suggests a hypothesis that the increased goblet cell in the colon of Cyp4a14-KO mice provides protection from mucosal injury and Cyp4a14-increased oxidative stress exacerbates DSS-induced colitis. Therefore, Cyp4a14 may represent a potential target for treating colitis.
Asunto(s)
Colitis/patología , Familia 4 del Citocromo P450/genética , Animales , Colitis/inducido químicamente , Colitis/veterinaria , Colon/metabolismo , Colon/patología , Familia 4 del Citocromo P450/deficiencia , Sulfato de Dextran/toxicidad , Femenino , Expresión Génica/efectos de los fármacos , Células Caliciformes/citología , Células Caliciformes/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Peroxidación de Lípido/efectos de los fármacos , Hígado/metabolismo , Malondialdehído/sangre , Ratones , Ratones Noqueados , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Índice de Severidad de la Enfermedad , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Diversity in the genetic lesions that cause cancer is extreme. In consequence, a pressing challenge is the development of drugs that target patient-specific disease mechanisms. To address this challenge, we employed a chemistry-first discovery paradigm for de novo identification of druggable targets linked to robust patient selection hypotheses. In particular, a 200,000 compound diversity-oriented chemical library was profiled across a heavily annotated test-bed of >100 cellular models representative of the diverse and characteristic somatic lesions for lung cancer. This approach led to the delineation of 171 chemical-genetic associations, shedding light on the targetability of mechanistic vulnerabilities corresponding to a range of oncogenotypes present in patient populations lacking effective therapy. Chemically addressable addictions to ciliogenesis in TTC21B mutants and GLUT8-dependent serine biosynthesis in KRAS/KEAP1 double mutants are prominent examples. These observations indicate a wealth of actionable opportunities within the complex molecular etiology of cancer.