RESUMEN
There has been growing interest in telemedicine for cystic fibrosis over recent years based largely on convenience for patients and/or increasing the frequency of surveillance and early detection which, it is assumed, could improve treatment outcomes. During 2020, the covid-19 pandemic catalysed the pace of development of this field, as CF patients were presumed to be at high risk of infection. Most clinics adapted to digital platforms with provision of lung function monitoring and sample collection systems. Here, we present the views of multidisciplinary team members at a large paediatric CF centre on what has worked well and what requires further optimisation in the future. In response to the question posed, 'Do we still need face to face clinics?' our answer is 'Yes, but not every time, and not for everyone'.
Asunto(s)
Fibrosis Quística , Telemedicina , COVID-19 , Niño , Fibrosis Quística/diagnóstico , Fibrosis Quística/epidemiología , Fibrosis Quística/terapia , Humanos , PandemiasRESUMEN
p21 (WAF1/Cip1) is the only member of the CIP/KIP family which has a well-characterized PCNA-binding domain. p21 is known to have an important function in the coordination of the cellular pathways which are activated in response to DNA damage, though the significance of the p21-PCNA interaction is not completely clear. We have analyzed the effects of expressing a miniprotein containing the PCNA-binding domain of p21 upon the cell cycle and upon the proliferation of various cell types. We have compared this with the effect of expressing a mutant form which is defective in PCNA-binding, but which retains the secondary cyclin-CDK-inhibitory site. No PCNA-dependent effects were seen in the short term upon cell cycle distribution. However, clonogenic assays show that the GFP-peptide miniprotein can significantly suppress proliferation in a PCNA-dependent manner. In some cell types, however, the suppression of proliferation was not PCNA-dependent, suggesting that cellular environment is a contributory factor to the effect of this miniprotein. The capacity of this peptide sequence to suppress cell proliferation in vivo is of interest as the basis for the design of potential antiproliferative therapeutic agents.
Asunto(s)
División Celular/fisiología , Ciclinas/metabolismo , Inhibidores Enzimáticos/metabolismo , Péptidos/metabolismo , Antígeno Nuclear de Célula en Proliferación/metabolismo , Western Blotting , División Celular/genética , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Ciclinas/genética , Colorantes Fluorescentes/metabolismo , Proteínas Fluorescentes Verdes , Humanos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Microscopía Fluorescente , Péptidos/genética , Pruebas de Precipitina , Antígeno Nuclear de Célula en Proliferación/genética , Estructura Terciaria de Proteína , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Transfección , Células Tumorales Cultivadas , Ensayo de Tumor de Célula MadreRESUMEN
Cell-free systems derived from unfertilized Xenopus eggs have been particularly informative in the study of the regulation and biochemistry of DNA replication. We have developed a Xenopus-based system to analyze proliferating cell nuclear antigen (PCNA)-specific effects on the functional properties of egg extracts. To do this, we have coupled peptides derived from p21 (Waf1/Cip1) to beads and used these to deplete PCNA from Xenopus egg extracts. The effect on various aspects of DNA replication can be analyzed after the readdition of PCNA and other purified proteins. Using this system, we have shown that replication of single-stranded M13 DNA is entirely dependent upon PCNA. By adding exogenous T7 DNA polymerase to PCNA-depleted extracts, we have uncoupled processive DNA replication from PCNA activity and so created an experimental system to analyze the dependence of postreplicative processes on PCNA function. We have shown that successful chromatin assembly is specifically dependent on PCNA. However, systems for analyzing the far more complex mechanisms required for the replication of nuclear double-stranded DNA have proved so far to be refractory to specific PCNA depletion.
Asunto(s)
Cromatina/fisiología , Ciclinas/metabolismo , Replicación del ADN/efectos de los fármacos , Inhibidores Enzimáticos/metabolismo , Antígeno Nuclear de Célula en Proliferación/fisiología , Animales , Antimaláricos/farmacología , Western Blotting , Cloroquina/farmacología , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Ciclinas/química , Ciclinas/genética , Replicación del ADN/fisiología , ADN Polimerasa Dirigida por ADN/genética , ADN Polimerasa Dirigida por ADN/metabolismo , Inhibidores Enzimáticos/química , Femenino , Humanos , Masculino , Oocitos/efectos de los fármacos , Oocitos/fisiología , Péptidos/química , Péptidos/metabolismo , Antígeno Nuclear de Célula en Proliferación/farmacología , Estructura Terciaria de Proteína , Proteínas Recombinantes/metabolismo , Espermatozoides/citología , Espermatozoides/fisiología , Extractos de Tejidos/química , Extractos de Tejidos/metabolismo , Xenopus laevisRESUMEN
As an aid to the management of the Pyrenean population of the brown bear Ursus arctos, a sexing method based on the amplification of a Y chromosome specific sequence has been developed, and tested using hairs found in the field as a source of DNA. This method involves a two-step polymerase chain reaction (PCR) which allows the detection of a very small amount of DNA, probably a single SRY gene molecule. The sex can reliably be identified using about 50pg of DNA extract as template. It is possible that this approach could, with adjustments, be used to identify the sex of other species of eutherian mammals.