Translation and assembly of HLA-DR antigens in Xenopus oocytes injected with mRNA from a human B-cell line.

HLA-DR antigens are polymorphic cell surface glycoproteins, expressed primarily in B lymphocytes and macrophages, which are thought to play an important role in the immune response. Two polypeptide chains, alpha and beta, are associated at the cell surface, and a third chain associates with alpha and beta intracellularly. RNA isolated from the human B-cell line Raji was injected in Xenopus laevis oocytes. Immunoprecipitates of translation products with several monoclonal antibodies revealed the presence of HLA-DR antigens similar to those synthesized in Raji cells. One monoclonal antibody was able to bind the beta chain after dissociation of the three polypeptide chains with detergent. The presence of all three chains was confirmed by two-dimensional gel electrophoresis. The glycosylation pattern of the three chains was identical to that observed in vivo, as evidenced in studies using tunicamycin, an inhibitor of N-linked glycosylation. The presence of alpha chains assembled with beta chains in equimolar ratio was further demonstrated by amino-terminal sequencing. An RNA fraction enriched for the three mRNAs, encoding alpha, beta, and intracellular chains, was isolated. This translation-assembly system and the availability of monoclonal antibodies make it possible to assay for mRNA encoding specific molecules among the multiple human Ia-like antigens.

Abrogation of HMX1 function causes rare oculoauricular syndrome associated with congenital cataract, anterior segment dysgenesis, and retinal dystrophy.

PURPOSE: To define the phenotypic manifestation, confirm the genetic basis, and delineate the pathogenic mechanisms underlying an oculoauricular syndrome (OAS). METHODS: Two individuals from a consanguineous family underwent comprehensive clinical phenotyping and electrodiagnostic testing (EDT). Genome-wide microarray analysis and Sanger sequencing of the candidate gene were used to identify the likely causal variant. Protein modelling, Western blotting, and dual luciferase assays were used to assess the pathogenic effect of the variant in vitro. RESULTS: Complex developmental ocular abnormalities of congenital cataract, anterior segment dysgenesis, iris coloboma, early-onset retinal dystrophy, and abnormal external ear cartilage presented in the affected family members. Genetic analyses identified a homozygous c.650A>C; p.(Gln217Pro) missense mutation within the highly conserved homeodomain of the H6 family homeobox 1 (HMX1) gene. Protein modelling predicts that the variant may have a detrimental effect on protein folding and/or stability. In vitro analyses were able to demonstrate that the mutation has no effect on protein expression but adversely alters function. CONCLUSIONS: Oculoauricular syndrome is an autosomal recessive condition that has a profound effect on the development of the external ear, anterior segment, and retina, leading to significant visual loss at an early age. This study has delineated the phenotype and confirmed HMX1 as the gene causative of OAS, enabling the description of only the second family with the condition. HMX1 is a key player in ocular development, possibly in both the pathway responsible for lens and retina development, and via the gene network integral to optic fissure closure.

Peripheral Nerve Repair: Multimodal Comparison of the Long-Term Regenerative Potential of Adipose Tissue-Derived Cells in a Biodegradable Conduit.

Tissue engineering is a popular topic in peripheral nerve repair. Combining a nerve conduit with supporting adipose-derived cells could offer an opportunity to prevent time-consuming Schwann cell culture or the use of an autograft with its donor site morbidity and eventually improve clinical outcome. The aim of this study was to provide a broad overview over promising transplantable cells under equal experimental conditions over a long-term period. A 10-mm gap in the sciatic nerve of female Sprague-Dawley rats (7 groups of 7 animals, 8 weeks old) was bridged through a biodegradable fibrin conduit filled with rat adipose-derived stem cells (rASCs), differentiated rASCs (drASCs), human (h)ASCs from the superficial and deep abdominal layer, human stromal vascular fraction (SVF), or rat Schwann cells, respectively. As a control, we resutured a nerve segment as an autograft. Long-term evaluation was carried out after 12 weeks comprising walking track, morphometric, and MRI analyses. The sciatic functional index was calculated. Cross sections of the nerve, proximal, distal, and in between the two sutures, were analyzed for re-/myelination and axon count. Gastrocnemius muscle weights were compared. MRI proved biodegradation of the conduit. Differentiated rat ASCs performed significantly better than undifferentiated rASCs with less muscle atrophy and superior functional results. Superficial hASCs supported regeneration better than deep hASCs, in line with published in vitro data. The best regeneration potential was achieved by the drASC group when compared with other adipose tissue-derived cells. Considering the ease of procedure from harvesting to transplanting, we conclude that comparison of promising cells for nerve regeneration revealed that particularly differentiated ASCs could be a clinically translatable route toward new methods to enhance peripheral nerve repair.

Decreasing Proportion of Recent Infections among Newly Diagnosed HIV-1 Cases in Switzerland, 2008 to 2013 Based on Line-Immunoassay-Based Algorithms.

BACKGROUND: HIV surveillance requires monitoring of new HIV diagnoses and differentiation of incident and older infections. In 2008, Switzerland implemented a system for monitoring incident HIV infections based on the results of a line immunoassay (Inno-Lia) mandatorily conducted for HIV confirmation and type differentiation (HIV-1, HIV-2) of all newly diagnosed patients. Based on this system, we assessed the proportion of incident HIV infection among newly diagnosed cases in Switzerland during 2008-2013. METHODS AND RESULTS: Inno-Lia antibody reaction patterns recorded in anonymous HIV notifications to the federal health authority were classified by 10 published algorithms into incident (up to 12 months) or older infections. Utilizing these data, annual incident infection estimates were obtained in two ways, (i) based on the diagnostic performance of the algorithms and utilizing the relationship 'incident = true incident + false incident', (ii) based on the window-periods of the algorithms and utilizing the relationship 'Prevalence = Incidence x Duration'. From 2008-2013, 3'851 HIV notifications were received. Adult HIV-1 infections amounted to 3'809 cases, and 3'636 of them (95.5%) contained Inno-Lia data. Incident infection totals calculated were similar for the performance- and window-based methods, amounting on average to 1'755 (95% confidence interval, 1588-1923) and 1'790 cases (95% CI, 1679-1900), respectively. More than half of these were among men who had sex with men. Both methods showed a continuous decline of annual incident infections 2008-2013, totaling -59.5% and -50.2%, respectively. The decline of incident infections continued even in 2012, when a 15% increase in HIV notifications had been observed. This increase was entirely due to older infections. Overall declines 2008-2013 were of similar extent among the major transmission groups. CONCLUSIONS: Inno-Lia based incident HIV-1 infection surveillance proved useful and reliable. It represents a free, additional public health benefit of the use of this relatively costly test for HIV confirmation and type differentiation.

Length of the Mitral Isthmus But Not Anatomical Location of Ablation Line Predicts Bidirectional Mitral Isthmus Block in Patients Undergoing Catheter Ablation of Persistent Atrial Fibrillation: A Randomized Controlled Trial.

INTRODUCTION: Mitral isthmus (MI) ablation is an effective option in patients undergoing ablation for persistent atrial fibrillation (AF). Achieving bidirectional conduction block across the MI is challenging, and predictors of MI ablation success remain incompletely understood. We sought to determine the impact of anatomical location of the ablation line on the efficacy of MI ablation. METHODS AND RESULTS: A total of 40 consecutive patients (87% male; 54 ± 10 years) undergoing stepwise AF ablation were included. MI ablation was performed in sinus rhythm. MI ablation was performed from the left inferior PV to either the posterior (group 1) or the anterolateral (group 2) mitral annulus depending on randomization. The length of the MI line (measured with the 3D mapping system) and the amplitude of the EGMs at 3 positions on the MI were measured in each patient. MI block was achieved in 14/19 (74%) patients in group 1 and 15/21 (71%) patients in group 2 (P = NS). Total MI radiofrequency time (18 ± 7 min vs. 17 ± 8 min; P = NS) was similar between groups. Patients with incomplete MI block had a longer MI length (34 ± 6 mm vs. 24 ± 5 mm; P < 0.001), a higher bipolar voltage along the MI (1.75 ± 0.74 mV vs. 1.05 ± 0.69 mV; P < 0.01), and a longer history of continuous AF (19 ± 17 months vs. 10 ± 10 months; P < 0.05). In multivariate analysis, decreased length of the MI was an independent predictor of successful MI block (OR 1.5; 95% CI 1.1-2.1; P < 0.05). CONCLUSIONS: Increased length but not anatomical location of the MI predicts failure to achieve bidirectional MI block during ablation of persistent AF.

Is the Front Line Prepared for the Changing Faces of Patients? Predictors of Cross-Cultural Preparedness Among Clinical Nurses and Resident Physicians in Lausanne, Switzerland.

UNLABELLED: Phenomenon: Assuring quality medical care for all persons requires that healthcare providers understand how sociocultural factors affect a patient's health beliefs/behaviors. Switzerland's changing demographics highlight the importance of provider cross-cultural preparedness for all patients-especially those at risk for social/health precarity. We evaluated healthcare provider cross-cultural preparedness for commonly encountered vulnerable patient profiles. APPROACH: A survey on cross-cultural care was mailed to Lausanne University hospital's "front-line healthcare providers": clinical nurses and resident physicians at our institution. Preparedness items asked "How prepared do you feel to care for ... ?" (referring to example patient profiles) on an ascending 5-point Likert scale. We examined proportions of "4 - well/5 - very well prepared" and the mean composite score for preparedness. We used linear regression to examine the adjusted effect of demographics, work context, cultural-competence training, and cross-cultural care problem awareness, on preparedness. FINDINGS: Of 885 questionnaires, 368 (41.2%) were returned: 124 (33.6%) physicians and 244 (66.4%) nurses. Mean preparedness composite was 3.30 (SD = 0.70), with the lowest proportion of healthcare providers feeling prepared for patients "whose religious beliefs affect treatment" (22%). After adjustment, working in a sensitized department (β = 0.21, p = .01), training on the history/culture of a specific group (β = 0.25, p = .03), and awareness regarding (a) a lack of practical experience caring for diverse populations (β = 0.25, p = .004) and (b) inadequate cross-cultural training (β = 0.18, p = .04) were associated with higher preparedness. Speaking French as a dominant language and physician role (vs. nurse) were negatively associated with preparedness (β = -0.26, p = .01; β = -0.22, p = .01). Insights: The state of cross-cultural care preparedness among Lausanne's front-line healthcare providers leaves room for improvement. Our study points toward institutional strategies to improve preparedness: notably, making sure departments are sensitized to cross-cultural care resources and increasing provider diversity to reflect the changing Swiss demographic.