Selection and Application of Aptamer Affinity for Protein Purification.

Aptamers are affinity-based oligonucleotide ligands raised against a target molecule, which might be of proteic or other nature. Aptamers are developed by using a reiterative in vitro selection procedure, named SELEX, in which the target is exposed to a combinatorial oligonucleotide combinatorial library. Target bound oligonucleotides are eluted, and PCR amplified followed by the next SELEX round. The process is repeated until no further increase in target binding affinity and specificity is achieved. Selected aptamers are identified and immobilized for protein purification. In view of their stability against denaturation and capability of renaturation, low costs of production, easiness of modification and stabilization, oligonucleotide aptamers are excellent tools as high-affinity ligands for applications of protein purification.

Evaluación psicométrica de la escala de calidad de servicios de atención en donantes de un banco de sangre en Medellín, Colombia, 2019 / Psychometric assessment of a quality scale perceived in blood bank donors of Medellín, Colombia, 2019 / Avaliação psicométrica de uma escala de qualidade percebida em doadores de um banco de sangue em Medellín, Colômbia, 2019

Resumen Objetivo: Evaluar la adaptabilidad del modelo Service Quality (servqual), con el fin de valorar la calidad del servicio en la atención de los donantes de un banco de sangre en la ciudad de Medellín. Metodología: Se llevó a cabo una evaluación psicométrica en 301 donantes de sangre, para determinar la calidad percibida. Se realizaron pruebas de consistencia interna con alpha de Cronbach e índice de discriminación, y se evaluó la validez de contenido de los dos cuestionarios y la estructura factorial, mediante análisis factorial exploratorio. Resultados: El alpha de Cronbach fue de 0,86; en las subescalas de expectativas, 0,87, y en percepciones, 0,77. Se observó una mayor consistencia interna en el cuestionario de expectativas que de percepciones. Aunque se obtuvo una baja consistencia interna en algunas dimensiones, no se afecta la escala de manera global. Conclusión: La escala servqual presenta un adecuado desempeño psicométrico y, por tanto, es útil para evaluar la calidad percibida en la atención en donantes de sangre, siempre y cuando sea abordada de forma unidimensional.

Abstract Objective: To assess the reproducibility and validity of the scale of perceived quality in donors from a blood bank in the city of Medellín. Methodology: Psychometric evaluation in 301 blood donors through internal consistency with Cronbach's alpha and discrimination index. Content validity of the polls and factorial structure were evaluated by means of exploratory factor analysis. Results: Cronbach's alpha was 0.86. For the expectations subscales, 0.87 and in perceptions 0.77. A greater internal consistency was observed in the poll of expectations rather than in that of perceptions. Although low internal consistency was obtained in some dimensions, this does not affect the scale globally. The use of the instrument should be approached in a unidimensional way. Conclusion: The servqual scale presents an adequate psychometric performance and, therefore, is useful to assess the perceived quality of care in blood donors in the Colombian context, as long as it is approached in a unidimensional way.

Resumo Objetivo: Avaliar a reprodutibilidade e validade da escala de qualidade percebida em doadores de um banco de sangue da cidade de Medellín. Metodologia: Avaliação psicométrica em 301 doadores de sangue, por meio da consistência interna com Alfa de Cronbach e índice de discriminação. A validade do conteúdo dos questionários e a estrutura fatorial foram avaliadas por meio de análise fatorial exploratória. Resultados: O Alfa de Cronbach foi de 0,86, nas subescalas de expectativas 0,87 e nas percepções 0,77. Foi observada maior consistência interna no questionário de expectativas do que de percepções. Embora tenha sido obtida baixa consistência interna em algumas dimensões, isso não afeta a escala globalmente; a utilização do instrumento deve ser abordada de uma forma unidimensional. Conclusão: A escala servqual apresenta um desempenho psicométrico adequado e, portanto, é útil para avaliar a percepção da qualidade do cuidado em doadores de sangue no contexto colombiano, desde que seja abordada de uma forma unidimensional.

FOXN1 compound heterozygous mutations cause selective thymic hypoplasia in humans.

We report on 2 patients with compound heterozygous mutations in forkhead box N1 (FOXN1), a transcription factor essential for thymic epithelial cell (TEC) differentiation. TECs are critical for T cell development. Both patients had a presentation consistent with T-/loB+NK+ SCID, with normal hair and nails, distinct from the classic nude/SCID phenotype in individuals with autosomal-recessive FOXN1 mutations. To understand the basis of this phenotype and the effects of the mutations on FOXN1, we generated mice using CRISPR-Cas9 technology to genocopy mutations in 1 of the patients. The mice with the Foxn1 compound heterozygous mutations had thymic hypoplasia, causing a T-B+NK+ SCID phenotype, whereas the hair and nails of these mice were normal. Characterization of the functional changes due to the Foxn1 mutations revealed a 5-amino acid segment at the end of the DNA-binding domain essential for the development of TECs but not keratinocytes. The transcriptional activity of this Foxn1 mutant was partly retained, indicating a region that specifies TEC functions. Analysis of an additional 9 FOXN1 mutations identified in multiple unrelated patients revealed distinct functional consequences contingent on the impact of the mutation on the DNA-binding and transactivation domains of FOXN1.

MIR205HG Is a Long Noncoding RNA that Regulates Growth Hormone and Prolactin Production in the Anterior Pituitary.

MicroRNAs (miRNAs) are processed from primary miRNA transcripts (pri-miRNAs), many of which are annotated as long noncoding RNAs (lncRNAs). We assessed whether MIR205HG, the host gene for miR-205, has independent functions as an lncRNA. Comparing mice with targeted deletions of MIR205HG and miR-205 revealed a functional role for the lncRNA in the anterior pituitary. Mice lacking MIR205HG had a temporal reduction in Pit1, growth hormone, and prolactin. This was mediated, in part, through the ability of this lncRNA to bind and regulate the transcriptional activity of Pit1 in conjunction with Zbtb20. Knockdown of MIR205HG in lactotropes decreased the expression of Pit1, Zbtb20, prolactin, and growth hormone, while its overexpression enhanced the levels of these transcripts. The effects of MIR205HG on the pituitary were independent of miR-205. The data support a role for MIR205HG as an lncRNA that regulates growth hormone and prolactin production in the anterior pituitary.

Enhanced Proteolytic Processing of Recombinant Human Coagulation Factor VIII B-Domain Variants by Recombinant Furins.

Recombinant human factor VIII (rFVIII) is used in replacement therapy for hemophilia A. Current research efforts are focused on bioengineering rFVIII molecules to improve its secretion efficiency and stability, limiting factors for its efficient production. However, high expression yield in mammalian cells of these rFVIII variants is generally associated with limited proteolytic processing. Non-processed single-chain polypeptides constitute non-natural FVIII molecule configurations with unpredictable toxicity and/or antigenicity. Our main objective was to demonstrate the feasibility of promoting full-proteolytic processing of an rFVIII variant retaining a portion of the B-domain, converting it into the smallest natural activatable form of rFVIII, while keeping its main advantage, i.e., improved secretion efficiency. We generated and employed a CHO-DG44 cell clone producing an rFVIII variant retaining a portion of the B-domain and the FVIII native cleavage site between Arg(1648) and Glu(1649). By bioengineering CHO-DG44 cells to express stably the recombinant human endoproteases PACE, PACE-SOL, PCSK5, PCSK6, or PCKS7, we were able to achieve complete intra- or extracellular proteolytic processing of this rFVIII variant. Additionally, our quantitative data indicated that removal of the B-domain segment by intracellular proteolytic processing does not interfere with this rFVIII variant secretion efficiency. This work also provides the first direct evidence of (1) intracellular cleavage at the Arg(1648) FVIII processing site promoted by wild-type PACE and PCSK7 and (2) proteolytic processing at the Arg(1648) FVIII processing site by PCSK6.

Manejo de los síntomas dolorosos de la endometriosis con dispositivo intrauterino de levonorgestrel en una paciente con pelvis congelada: reporte de caso / Managing endometriosis pain symptoms with a levonorgestrel intrauterine device in a patient suffering from frozen pelvis: a case report

El manejo de los síntomas dolorosos de la endometriosis es un tema controvertido en nuestra especialidad. La mayoría de los tratamientos utilizado ofrecen diferentes grados de mejoría a las pacientes mientras los reciben, tienen múltiples efectos secundarios y es previsible una recaída pronto en el tiempo siguiente a su descontinuación. Muchos de los medicamentos adversos no deben ser utilizados por mucho tiempo. En algunas pacientes con paridad satisfecha, la anexo-histerectomía es una alternativa terapéutica para mejorar la calidad de vida cuando han sufrido la enfermedad durante toda su etapa reproductiva. En casos avanzados de endometriosis, dicho tipo de cirugía puede ser difícil y poner en riesgo de daño a los órganos pélvicos. Describimos el caso de una paciente en quien fue imposible practicar la cirugía optándose por manejar sus síntomas con un dispositivo intrauterino de levonorgestrel (Mirena) con un resultado satisfactorio.