Comparison of the editing patterns and editing efficiencies of TALEN and CRISPR-Cas9 when targeting the human CCR5 gene

Abstract The human C-C chemokine receptor type-5 (CCR5) is the major transmembrane co-receptor that mediates HIV-1 entry into target CD4+ cells. Gene therapy to knock-out the CCR5 gene has shown encouraging results in providing a functional cure for HIV-1 infection. In gene therapy strategies, the initial region of the CCR5 gene is a hotspot for producing functional gene knock-out. Such target gene editing can be done using programmable endonucleases such as transcription activator-like effector nucleases (TALEN) or clustered regularly interspaced short palindromic repeats (CRISPR-Cas9). These two gene editing approaches are the most modern and effective tools for precise gene modification. However, little is known of potential differences in the efficiencies of TALEN and CRISPR-Cas9 for editing the beginning of the CCR5 gene. To examine which of these two methods is best for gene therapy, we compared the patterns and amount of editing at the beginning of the CCR5 gene using TALEN and CRISPR-Cas9 followed by DNA sequencing. This comparison revealed that CRISPR-Cas9 mediated the sorting of cells that contained 4.8 times more gene editing than TALEN+ transfected cells.

Transfusion medicine in medical education: an analysis of curricular grids in Brazil and a review of the current literature

Background Blood transfusions are one of the most performed medical procedures in the world. Thus, as education in transfusion medicine is vital to medical care, it should aim to promote a responsible practice with the rational use of blood by doctors. This study aims to investigate the situation of the teaching of transfusion medicine in medical schools in Brazil. Method The websites of the 249 Brazilian medical schools in operation in June 2015 were visited and the curricula of the medical courses were investigated in respect to the presence or absence of a transfusion medicine discipline. When available, the subject grids were analyzed to verify whether a description of content regarding transfusion medicine was given within other disciplines. Results Of the 249 medical school sites visited, information on the curriculum was obtained from 178. Of the medical schools that published their curriculum, 132 (74.1%) did not have disciplines of transfusion medicine or hematology and only seven (3.9%) had a discipline of transfusion medicine in the curricular grid. Conclusions Education on transfusion medicine is of fundamental importance for safe and efficient transfusion practices. Deficiencies in medical knowledge of this subject have been found worldwide. The results of this study indicate a possible deficiency in teaching the basics of this specialty. Thus, additional prospective studies to assess the knowledge and practice of transfusion medicine in Brazilian medical schools are warranted, which could prompt a discussion on the importance of offering training in transfusion medicine to medical students.

Fibronectin modulates thymocyte-thymic epithelial cell interactions following Trypanosoma cruzi infection

Developing thymocytes interact with thymic epithelial cells (TECs) through cell-cell interactions, TEC-derived secretory moieties and extracellular matrix (ECM)-mediated interactions. These physiological interactions are crucial for normal thymocyte differentiation, but can be disrupted in pathological situations. Indeed, there is severe thymic atrophy in animals acutely infected with Trypanosoma cruzi due to CD4+CD8+ thymocyte depletion secondary to caspase-mediated apoptosis, together with changes in ECM deposition and thymocyte migration. We studied an in vitro model of TEC infection by T. cruzi and found that infected TEC cultures show a reduced number of cells, which was likely associated with decreased proliferative capacity, but not with increased cell death, as demonstrated by bromodeoxyuridine and annexin-V labelling. The infected TEC cultures exhibited increased expression of fibronectin (FN), laminin (LM) and type IV collagen. Importantly, treatment with FN increased the relative number of infected cells, whereas treatment with anti-FN or anti-LM antibodies resulted in lower infection rates. Consistent with these data, we observed increased thymocyte adhesion to infected TEC cultures. Overall, these results suggest that ECM molecules, particularly FN, facilitate infection of the thymic epithelium and that the consequent enhancement of ECM expression might be associated with changes in TEC-thymocyte interactions.