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1.
Pigment Cell Melanoma Res ; 35(6): 613-621, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-35920064

RESUMO

The skin of adult mammals protects from radiation, physical and chemical insults. While melanocytes and melanocyte-producing stem cells contribute to proper skin function in healthy organisms, dysfunction of these cells can lead to the generation of malignant melanoma-the deadliest type of skin cancer. Addressing cells of the melanocyte lineage in vivo represents a prerequisite for the understanding of melanoma on cellular level and the development of preventive and treatment strategies. Here, the inducible Cre-loxP-system has emerged as a promising tool to specifically target, monitor, and modulate cells in adult mice. Re-analysis of existing sequencing data sets of melanocytic cells revealed that genes with a known function in neural cells, including neural stem cells (Aldh1L1 and Nestin), are also expressed in melanocytic cells. Therefore, in this study, we explored whether the promoter activity of Nestin and Aldh1L1 can serve to target cells of the melanocyte lineage using the inducible CreERT2 -loxP-system. Using an immunohistochemical approach and different time points of analysis, we were able to map the melanocytic fate of recombined stem cells in the adult hair follicle of Nestin-CreERT2 and Aldh1L1-CreERT2 transgenic mice. Thus, we here present two new mouse models and propose their use to study and putatively modulate adult melanocytic cells in vivo.


Assuntos
Melanoma , Neoplasias Cutâneas , Animais , Camundongos , Integrases/genética , Melanócitos/patologia , Melanoma/patologia , Camundongos Transgênicos , Nestina/genética , Neoplasias Cutâneas/genética
2.
Methods Mol Biol ; 2045: 187-199, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30888667

RESUMO

The use of inducible transgenic Nestin-CreERT2 mice has proved to be an essential research tool for gene targeting and studying the molecular pathways implicated in adult neurogenesis, namely, inside the adult subgranular zone (SGZ) of the dentate gyrus and the adult subventricular zone (SVZ) lining the lateral ventricles. Several lines of Nestin-CreER-expressing mice were generated and used in adult neurogenesis research in the past two decades; however, their suitability for studying neurogenesis in aged mice remains elusive. Here, we assessed the efficiency of Cre-loxP genetic recombination in the aging SVZ using the Nestin-CreERT2/Rosa26YFP line designed by Lagace et al. (J Neurosci 27(46):12623-12629, 2007). This analysis was performed in 12-month-old (middle-aged) mice and 20-month-old (old) mice compared to 2-month-old (young adult) mice. To evaluate successful recombination, our approach relies on the histological assessment of Cre mRNA level of expression and the YFP reporter gene's expression inside the aging SVZ by combining in situ hybridization and immunohistochemistry. Using co-immunolabeling, this approach also provides the advantage of estimating the percentage of recombined progeny [(GFP+Nestin+)/Nestin+] and the rate of cell proliferation [(GFP+Ki67+)/GFP+] inside the aging SVZ niche.


Assuntos
Imunofluorescência/métodos , Ventrículos Laterais/metabolismo , Nestina/genética , Células-Tronco Neurais/citologia , Neurogênese , Neurônios/citologia , Recombinação Genética , Células-Tronco Adultas/citologia , Células-Tronco Adultas/metabolismo , Envelhecimento , Animais , Linhagem da Célula , DNA Complementar/genética , Genes Reporter/genética , Hibridização In Situ , Integrases/genética , Integrases/metabolismo , Ventrículos Laterais/fisiologia , Proteínas Luminescentes/análise , Proteínas Luminescentes/química , Proteínas Luminescentes/genética , Camundongos , Camundongos Transgênicos , Células-Tronco Neurais/metabolismo , Neurônios/metabolismo , Oligorribonucleotídeos/genética , Recombinação Genética/efeitos dos fármacos , Fluxo de Trabalho
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