RESUMEN
In 2020, STEM training programs across the country were challenged to provide support to students during a nation-wide shutdown of research institutions in response to the COVID-19 pandemic. Five U.S. high school science internship programs funded by the Doris Duke Charitable Foundation, with a history of collaboration, developed innovative strategies for distance-learning (DL) opportunities during the pandemic. Forty under-represented high school and undergraduate students were paired with scientific mentors at one of the programs for a DL scientific internship. Summer training combined synchronous and asynchronous programming with research projects adapted for DL success. Ninety-five percent of students who participated were satisfied with the training experience, nearly identical to exit survey responses from 2019 when our programs were held in-person. More students were interested in pursuing a career in research at the end of the program and credited the DL experience with increasing interest in research careers. Some DL elements were ideal for underrepresented youth, including a more flexible schedule and elimination of cost and time for travel. While the lack of in-person instruction challenged our ability to create a strong student community, we found that preparation, communication, and flexibility were key elements to these successful DL programs. The increased emphasis on interpretation and analysis of data, rather than data collection, enhanced student learning. This manuscript highlights the changes made to our curricula, elements which were most successful, and recommends strategies for future distance-learning programming.
RESUMEN
The p75(NTR) neurotrophin receptor has been implicated in multiple biological and pathological processes. While significant advances have recently been made in understanding the physiologic role of p75(NTR) , many details and aspects remain to be determined. This is in part because the two existing knockout mouse models (Exons 3 or 4 deleted, respectively), both display features that defy definitive conclusions. Here we describe the generation of mice that carry a conditional p75(NTR) (p75(NTR-FX) ) allele made by flanking Exons 4-6, which encode the transmembrane and all cytoplasmic domains, by loxP sites. To validate this novel conditional allele, both neural crest-specific p75(NTR) /Wnt1-Cre mutants and conventional p75(NTR) null mutants were generated. Both mutants displayed abnormal hind limb reflexes, implying that loss of p75(NTR) in neural crest-derived cells causes a peripheral neuropathy similar to that seen in conventional p75(NTR) mutants. This novel conditional p75(NTR) allele will offer new opportunities to investigate the role of p75(NTR) in specific tissues and cells.
Asunto(s)
Alelos , Ratones Noqueados/genética , Receptores de Factor de Crecimiento Nervioso/genética , Animales , Clonación Molecular , Cruzamientos Genéticos , Embrión de Mamíferos/metabolismo , Embrión de Mamíferos/patología , Desarrollo Embrionario , Exones , Femenino , Vectores Genéticos/genética , Vectores Genéticos/metabolismo , Genotipo , Inmunohistoquímica , Extremidad Inferior/fisiología , Masculino , Ratones , Ratones Noqueados/embriología , Ratones Noqueados/metabolismo , Cresta Neural/embriología , Cresta Neural/metabolismo , Cresta Neural/patología , Enfermedades del Sistema Nervioso Periférico/patología , Receptores de Factor de Crecimiento Nervioso/metabolismo , Reflejo AnormalRESUMEN
Neuroblastoma (NB) is a childhood cancer that arises in the adrenal gland and often shows differentiated neuronal and glial elements. The RET receptor signal pathway is functional in most NB, while loss of nerve growth factor (NGF) receptor (trkA) gene expression correlates with an aggressive phenotype. Thus, we hypothesized that the RET and TRKA signal pathways collaborate to instruct NB differentiation, reminiscent of normal neuronal maturation. Here, we demonstrate that activation of the RET receptor by glial cell line-derived neurotrophic factor (GDNF) increases expression of the RET receptor complex in a panel of malignant human NB cell lines, indicative of a positive feedback mechanism. GDNF also induces growth cessation concomitant with an arrest of cells in the G(0)/G(1) phase of the cell cycle. Furthermore, GDNF synergizes with ciliary neurotrophic factor (CNTF) to enhance TRKA receptor expression, thereby strengthening the NGF-mediated differentiation signal. Differentiated NB cells downregulate expression of the amplified N-myc gene, concurrent with the arrest of cell proliferation, while expressing neuron-specific markers (i.e., SCG10). Interestingly, maintenance of differentiated NB cells in culture is independent of the trophic activity of GDNF, but depends on TRKA signaling, thereby re-enacting the differentiation of normal sympathoadrenal (SA) progenitor cells.
Asunto(s)
Neuroblastoma/patología , Proteínas Proto-Oncogénicas/fisiología , Proteínas Tirosina Quinasas Receptoras/fisiología , Receptor trkA/fisiología , Diferenciación Celular , Línea Celular Tumoral , Factor Neurotrófico Ciliar/farmacología , Relación Dosis-Respuesta a Droga , Factor Neurotrófico Derivado de la Línea Celular Glial , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial , Humanos , Factor de Crecimiento Nervioso/farmacología , Factores de Crecimiento Nervioso/farmacología , Proteínas Proto-Oncogénicas c-myc/análisis , Proteínas Proto-Oncogénicas c-ret , Transducción de SeñalRESUMEN
Brain injuries by physical trauma, epileptic seizures, or microbial infection upset the osmotic homeostasis resulting in cell swelling (cerebral edema), inflammation, and apoptosis. Expression of the neurotrophin receptor p75NTR is increased in the injured tissue and axon regeneration is repressed by the Nogo receptor using p75NTR as the signal transducer. Hence, p75NTR seems central to the injury response and we wished to determine the signals that regulate its expression. Here, we demonstrate that tonicity mediated cell swelling rapidly activates transcription of the endogenous p75NTR gene and of a p75NTR promoter-reporter gene in various cell types. Transcription activation is independent of de novo protein synthesis and requires the activities of phospholipase C, protein kinase C, and nitric-oxide synthase. Hence, p75NTR is a nitric oxide effector gene regulated by osmotic swelling, thereby providing a strategy for therapeutic intervention to modulate p75NTR functions following injury.
Asunto(s)
Ácido Egtácico/análogos & derivados , Proteínas de la Mielina/metabolismo , Óxido Nítrico/metabolismo , Ósmosis , Receptores de Factor de Crecimiento Nervioso/metabolismo , Western Blotting , Calcio/metabolismo , Reactivos de Enlaces Cruzados/farmacología , Relación Dosis-Respuesta a Droga , Ácido Egtácico/metabolismo , Electroforesis en Gel de Poliacrilamida , Citometría de Flujo , Regulación de la Expresión Génica , Humanos , Immunoblotting , NG-Nitroarginina Metil Éster/farmacología , Neuronas/metabolismo , Óxido Nítrico Sintasa/metabolismo , Proteínas Nogo , Regiones Promotoras Genéticas , Proteína Quinasa C/metabolismo , Receptor de Factor de Crecimiento Nervioso , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Factores de Tiempo , Transcripción Genética , Activación Transcripcional , Transfección , Células Tumorales Cultivadas , Fosfolipasas de Tipo C/metabolismoRESUMEN
Neuroblastoma (NB) is the most common extracranial solid tumor of childhood. In advanced disease stages, prognosis is poor and treatments have limited efficacy, thus novel strategies are warranted. The synthetic retinoid Fenretinide (HPR) induces apoptosis in NB and melanoma cell lines. We reported an in vitro potentiation of HPR effects on melanoma cells when the drug is incorporated into GD2-targeted immunoliposomes (anti-GD2-SIL-HPR). We investigated the antitumor activity of anti-GD2-SIL-HPR against NB cells, both in vitro and in vivo. Anti-GD2-SIL showed specific, competitive binding to and uptake by, various NB cell lines. In in vitro cytotoxicity studies, NB cells, incubated with 30 microM HPR entrapped in anti-GD2-immunoliposomes, showed a significant reduction in cellular growth compared to free HPR, HPR entrapped in Ab-free liposomes or anti-GD2 empty liposomes. In an in vivo NB metastatic model, we demonstrated that anti-GD2-SIL-HPR completely inhibited the development of macroscopic and microscopic metastases in comparison to controls. Similar, but significantly less potent, antitumor effect was observed also in mice treated with anti-GD2 immunoliposomes without HPR (anti-GD2-SIL-blank) or anti-GD2 MAb alone (p = 0.0297 and p = 0.0294, respectively, vs. anti-GD2-SIL-HPR). Moreover, our results clearly demonstrated that although anti-GD2 MAb had a strong antitumor effect in this in vivo NB model, 100% curability was obtained only after treatment with anti-GD2-SIL-HPR (p < 0.0001). Anti-GD2 liposomal HPR should receive clinical evaluation as adjuvant therapy of neuroblastoma.