Mesenchymal stromal cells induced regulatory B cells are enriched in extracellular matrix genes and IL-10 independent modulators.

Regulatory B cells (Breg) are essential players in tolerance and immune homeostasis. However, lack of specific Breg markers limit their potential in clinical settings. Mesenchymal stromal cells (MSC) modulate B cell responses and are described to induce Breg in vitro. The aim of this work was to characterize MSC induced Breg (iBreg) and identify specific Breg biomarkers by RNAseq. After 7-day coculture with adipose tissue-derived MSC, B cells were enriched in transitional B cell populations, with increased expression and secretion of IL-10 and no TNFα. In addition, iBreg showed potential to modulate T cell proliferation at 2 to 1 cell ratios and their phenotype remained stable for 72h. RNAseq analysis of sorted IL-10 positive and negative iBreg populations identified over 1500 differentially expressed genes (DEG) among both populations. Analysis of biological processes of DEG highlighted an enrichment of immune regulation and extracellular matrix genes in IL-10- iBreg populations, while IL-10+ iBreg DEG were mostly associated with cell activation. This was supported by T cells modulation assays performed in the presence of anti-IL-10 neutralizing antibodies showing the non-essential role of IL-10 in the immunomodulatory capacity of iBregs on T cells. However, based on RNAseq results we explored the role of TGF-ß and found out that it plays a major role on iBreg induction and iBreg immunomodulatory properties. Therefore, we report that MSC induce B cell populations characterized by the generation of extracellular matrix and immune modulation independently of IL-10.

Modulation of the autophagic pathway inhibits HIV-1 infection in human lymphoid tissue cultured ex vivo.

A complex link exists between HIV-1 and autophagy, and discordant results have been reported in different in vitro models regarding the way HIV and autophagy modulate each other. Despite this, there is very limited knowledge about the interplay between HIV and autophagy in vivo in lymphoid tissue, due in part by the lack of cell models that recapitulate the in vivo setting. Here, we evaluate the interrelationship between HIV and autophagy using human ex vivo lymphoid tissue cultures as an HIV infection model. Our results showed that human lymphoid aggregated cultures (HLACs) from tonsillar tissue displayed fully functional autophagic activity. In this system, HIV infection resulted in an increase in autophagy. Notably, we observed that both, autophagy-enhancing (rapamycin) or blocking drugs (3-methyladenine, chloroquine and bafilomycin), were able to decrease HIV-DNA levels and HIV replication. Therefore, efficient HIV-1 replication requires a fine-tuned level of autophagy, so modifications of this balance will have a negative impact on its replication. Therefore, targeting the autophagic pathway could be a new therapeutic approach to be explored to treat HIV-1 infection. Ex vivo cultures of human lymphoid tissue are a suitable model to obtain further insights into HIV and its intricate relationship with autophagy.

In Vitro Characterization of Human CD24hiCD38hi Regulatory B Cells Shows CD9 Is Not a Stable Breg Cell Marker.

Regulatory B (Breg) cells are endowed with immune suppressive functions. Various human and murine Breg subtypes have been reported. While interleukin (IL)-10 intracellular staining remains the most reliable way to identify Breg cells, this technique hinders further essential functional studies. Recent findings suggest that CD9 is an effective surface marker of murine IL-10 competent Breg cells. However, the stability of CD9 and its relevance as a unique marker for human Breg cells, which have been widely characterized as CD24hiCD38hi, have not been investigated. Here, we demonstrate that CD9 expression is sensitive to in vitro B cell stimulations. CD9 expression could either be re-expressed or downregulated in purified CD9-negative B cells and CD9-positive B cells, respectively. We found no significant differences in the Breg differentiation capacity of the CD9-negative and CD9-positive B cells. Furthermore, CD9-positive B cells co-express CD40 and CD86, suggesting their nature as B cell activation or co-stimulatory molecules, rather than regulatory ones. Therefore, we report the relatively unstable CD9 as a distinct surface molecule, indicating the need for further research for a more reliable marker to purify human Breg cells.

Anti-Siglec-1 antibodies block Ebola viral uptake and decrease cytoplasmic viral entry.

Several Ebola viruses cause outbreaks of lethal haemorrhagic fever in humans, but developing therapies tackle only Zaire Ebola virus. Dendritic cells (DCs) are targets of this infection in vivo. Here, we found that Ebola virus entry into activated DCs requires the sialic acid-binding Ig-like lectin 1 (Siglec-1/CD169), which recognizes sialylated gangliosides anchored to viral membranes. Blockage of the Siglec-1 receptor by anti-Siglec-1 monoclonal antibodies halted Ebola viral uptake and cytoplasmic entry, offering cross-protection against other ganglioside-containing viruses such as human immunodeficiency virus type 1.

HIV-1 immune activation induces Siglec-1 expression and enhances viral trans-infection in blood and tissue myeloid cells.

BACKGROUND: Myeloid cells are key players in the recognition and response of the host against invading viruses. Paradoxically, upon HIV-1 infection, myeloid cells might also promote viral pathogenesis through trans-infection, a mechanism that promotes HIV-1 transmission to target cells via viral capture and storage. The receptor Siglec-1 (CD169) potently enhances HIV-1 trans-infection and is regulated by immune activating signals present throughout the course of HIV-1 infection, such as interferon α (IFNα). RESULTS: Here we show that IFNα-activated dendritic cells, monocytes and macrophages have an enhanced ability to capture and trans-infect HIV-1 via Siglec-1 recognition of viral membrane gangliosides. Monocytes from untreated HIV-1-infected individuals trans-infect HIV-1 via Siglec-1, but this capacity diminishes after effective antiretroviral treatment. Furthermore, Siglec-1 is expressed on myeloid cells residing in lymphoid tissues, where it can mediate viral trans-infection. CONCLUSIONS: Siglec-1 on myeloid cells could fuel novel CD4(+) T-cell infections and contribute to HIV-1 dissemination in vivo.

Orthopaedic Residents Improve Confidence and Knot-Tying Speed With a Skills Course.

PURPOSE: To determine the effect of a knot-tying module, within an arthroscopic training course, on resident speed, resident confidence, and biomechanical quality of arthroscopically tied knots. METHODS: Sixty-four participants (8 postgraduate year [PGY]-3 and 8 PGY-4 orthopaedic residents annually for 4 years) were enrolled in a 5-day training course, which included a daily knot-tying module. Self-assessed confidence was obtained by pre-course (day 1) and post-course (day 5) questionnaire. Each participant tied 5 sequential knots using an arthroscopic knot-tying station. Time per knot was recorded in seconds. Knots were later preloaded, cycled, and tested for peak load to failure and displacement change. Mean peak load to failure, displacement change, speed, and confidence were compared before and after training. RESULTS: The mean time to complete 5 knots was significantly faster after training (12.8 minutes before the course [day 1] v 9.39 minutes after the course [day 5]) (P < .0001). Confidence improved from pre-course (mean, 3.3) to post-course (mean, 7.8) questionnaires (P < .0001). No statistically significant difference was found between peak force for pre-course (mean, 136 N) and post-course (mean, 138 N) knots (P = .076). No statistically significant difference was detected in mean displacement change (mean, 3.51 mm before the course v 3.57 mm after the course) (P = .61). Comparison of PGY-3 and PGY-4 residents was significant only for a higher pre-course confidence in PGY-4 residents (P = .02). CONCLUSIONS: Participation in an arthroscopic knot-tying module improves resident speed and confidence in tying arthroscopic knots. Our data did not show a significant change in peak load to failure or loop security with training. These findings suggest that participation in a knot-tying module improves efficiency regarding arthroscopic knot tying by residents. CLINICAL RELEVANCE: Residents who practice arthroscopic knot tying 5 days per year as part of an arthroscopic training course may be more efficient in the operating room.

Arthroscopic versus open comparison of long head of biceps tendon visualization and pathology in patients requiring tenodesis.

PURPOSE: The purpose of this study was to compare arthroscopic versus open examination of the proximal long head of the biceps tendon (LHB) in patients undergoing open, subpectoral tenodesis. METHODS: Eighty consecutive patients were prospectively enrolled, of whom 62 were included in the study. During arthroscopy, the most distal extent of the LHB visualized was marked with a Bovie device. The tendon was pulled into the joint with an arthroscopic grasper, showing additional LHB and was again marked with the device. LHB fraying, flattening, redness, and degeneration were graded as absent, mild, moderate, or severe. During open subpectoral tenodesis, the grossly visualized LHB was graded in the same manner and the locations of both marks plus the total length of the LHB observed during open visualization were measured and recorded. After subpectoral tenodesis, the excised portion of the LHB was histologically graded as normal, fibrosis/tendinosis, or inflamed. RESULTS: On average, during open tenodesis, 95 mm (range, 75 to 130 mm) of LHB was visualized. This was greater than the length visualized during diagnostic arthroscopy of 16 mm (range, 5 to 28 mm), or 17%, and the length visualized while pulling the tendon into the joint with an arthroscopic grasper of 30 mm (range, 15 to 45 mm), or 32%. The difference in LHB length observed during open versus arthroscopic examination with a grasper was statistically significant (P < .0001). In addition, when compared with LHB pathology observed in an open manner, arthroscopic visualization showed only 67% of pathology, underestimated noted pathology in 56% of patients, and overestimated noted pathology in 11% of patients. Histologic evaluation showed fibrosis/tendinosis in 100% of cases but inflammation in only 5%. CONCLUSIONS: When compared with open inspection during subpectoral tenodesis, arthroscopic examination of the LHB visualizes only 32% of the tendon and may underestimate pathology. LEVEL OF EVIDENCE: Level II, diagnostic study-development of diagnostic criteria based on consecutive patients with universally applied gold standard.

Monitoring breathing rate at home allows early identification of COPD exacerbations.

BACKGROUND: Respiratory frequency increases during exacerbations of COPD (ECOPD). We hypothesized that this increase can be detected at home before ECOPD hospitalization. METHODS: To test this hypothesis, respiratory frequency was monitored at home daily for 3 months in 89 patients with COPD (FEV1, 42.3% ± 14.0%; reference) who were receiving domiciliary oxygen therapy (9.6 ± 4.0 h/d). RESULTS: During follow-up, 30 patients (33.7%) required hospitalization because of ECOPD. In 21 of them (70%), mean respiratory frequency increased (vs baseline) during the 5 days that preceded it (from 15.2 ± 4.3/min to 19.1 ± 5.9/min, P < .05). This was not the case in patients without ECOPD (16.1 ± 4.8/min vs 15.9 ± 4.9/min). Receiver operating characteristic analysis showed that 24 h before hospitalization, a mean increase of 4.4/min (30% from baseline) provided the best combination of sensitivity (66%) and specificity (93%) (area under the curve [AUC] = 0.79, P < .05). Two days before hospitalization, a mean increase of 2.3/min (15% change from baseline) was associated with a sensitivity of 72% and a specificity of 77% (AUC = 0.76, P < .05). CONCLUSIONS: Respiratory frequency can be monitored daily at home in patients with COPD receiving domiciliary oxygen therapy. In these patients, breathing rate increases significantly days before they require hospitalization because of ECOPD. This may offer a window of opportunity for early intervention.

Identification and diversity of multiresistant Corynebacterium striatum clinical isolates by MALDI-TOF mass spectrometry and by a multigene sequencing approach.

BACKGROUND: The genus Corynebacterium is composed of gram-positive bacteria that are widely distributed throughout the environment; these bacteria are also part of the normal microbiota of human skin and mucous membranes. Multiple studies have shown that species of this genus, including C. striatum, become pathogenic to humans under special conditions. Our aim was to determine the characteristics of clinical multiresistant strains of C. striatum that were isolated in our geographical region, to determine their diversity, and to compare them with the type strain and with related species. We studied fifty-two strains of C. striatum isolated from different hospitals from Mallorca, Spain, mainly from the Hospital Joan March in Bunyola, Mallorca. Most of the strains were isolated from sputum cultures of respiratory samples from patients with chronic obstructive pulmonary disease. To gain further insight into the genetic diversity of the strains, we analysed several housekeeping genes and other genes associated with antibiotic resistance. Strains were also characterised phenotypically by their antibiotic resistance profiles and by MALDI-TOF mass spectrometry analysis. RESULTS: The ITS1 region, gyrA and rpoB were chosen as the appropriate genes in the C. striatum genome to study the genetic diversity of C. striatum species and to discriminate between strains. After analysing these three genes, four sequence types (ST2, ST4, ST1 and ST11) were found to be the most abundant. Splits tree analysis of the strains demonstrated that these clinical isolates did not share any alleles with the type strain of the species. Recombination was detected within all of the C. striatum isolates, and different clonal populations were detected within the samples. CONCLUSIONS: Our results demonstrate that the isolates were best identified using gene-based molecular methods; using these methods, the isolated strains were determined to be different from the type strain of C. striatum. The ITS1 region and the gyrA and rpoB genes were selected because of their variability and were the most useful tools for discriminating between strains. The phenotype and antibiotype characteristics of the strains did not seem suitable for typing purposes. MALDI-TOF mass spectrometry can be a useful method for identifying and discriminating between C. striatum strains.

Spirometrically-defined restrictive ventilatory defect: population variability and individual determinants.

BACKGROUND: A spirometrically-defined restrictive ventilatory defect is a common finding when performing spirometry. AIMS: We aimed to determine the frequency, geographical variation, individual consequences, and 'severity' of the restrictive ventilatory defect. METHODS: A population-based study was conducted in Spain. The response rate from 11 participating centres was 88.9%, totalling 3,802 participants. A restrictive ventilatory defect was defined according to pre-bronchodilator spirometry as forced expiratory volume in 1s/forced vital capacity (FEV1/FVC) >0.70 and a predicted FVC <80%, in accordance with current American Thoracic Society/European Respiratory Society guidelines. RESULTS: The prevalence of a restrictive ventilatory defect was 12.7% (95% CI 9.7% to 15.7%), with the highest in Seville (19.4%) and Burgos (18.5%) and the lowest in Oviedo (5.2%) and Madrid-La Princesa (5.7%) (p<0.000). Although most of the participants (97.1%) with a restrictive ventilatory defect were objectively considered 'mild' by spirometry (%predicted FVC 50-80%), they reported more phlegm, dyspnoea, and wheezing than healthy control participants (p<0.000), and scored worse in all St George's Respiratory Questionnaire domains of quality of life and activities of daily living (p<0.000). Interestingly, they scored similarly to participants with chronic obstructive pulmonary disease (COPD) in both (p=0.102 and p=0.217). In a multivariate analysis, older age, male gender, heavy smoking, low education, and high body mass index were independently associated with having a restrictive ventilatory defect. CONCLUSIONS: A restrictive ventilatory defect in spirometry is a common finding (12.7%) with a highly variable geographical distribution (range 3.7) whose population burden is important in terms of quality of life and activities of daily living and similar to that of an obstructive pattern compatible with COPD.

High prevalence of undiagnosed airflow limitation in patients with cardiovascular disease.

BACKGROUND: The prevalence of airflow limitation (AL) in patients with cardiovascular disease (CVD) is unknown, and whether AL is adequately diagnosed and treated in these patients has not been investigated before, to our knowledge. METHODS: We compared clinical and spirometric data in three groups of individuals. Two of them were participants in the follow-up of an ongoing population-based study according to the presence or absence of CVD. The third group included patients with coronary artery disease (CAD) confirmed by coronariography regularly visited at a tertiary referral university hospital. AL was defined according to the Global Initiative for Obstructive Lung Disease guidelines. RESULTS: We studied 450 population participants without CVD, 52 population participants with CVD, and 119 hospital patients with CAD. The prevalence of AL in these three groups was 17.5% (95% CI, 14.0-21.0), 19.2% (95% CI, 8.1-30.7), and 33.6% (95% CI, 25.0-42.2), respectively (P < .05). Underdiagnosis of AL ranged from 60% in population participants with CVD up to 87.2% in hospital patients with CAD. Sixty percent of those with spirometrically confirmed AL (in all three groups) did not receive any respiratory treatment. CONCLUSIONS: AL is frequent in individuals with CVD, particularly in those with CAD attended in the hospital, is largely underdiagnosed and therefore is highly undertreated. TRIAL REGISTRATION: Clinicaltrials.gov; Identifier: NCT00787748.

Principales características de los pacientes que demandan atención en el centro de calificación de enfermedades cardiovasculares y que fueron diagnosticados de fiebre reumática. De enero 1977 a diciembre de 1979. Quito enero 1986 / Main characteristics of the patients who demand attention in the center of qualification of cardiovascular diseases and who were diagnosed of rheumatic fever. Of January 1977 to December of 1979. Exempt January 1986

La presente investigación correspondiente a principales características de los pacientes con fiebre reumática, se realizó en el Centro de Calificación de Enfermedades Cardiovasculares debido a los subregistros existentes en la institució0 donde el grupo inició sus práctivcas, La institución mencionada tiene un programa bastante definido en el que abarca el control y tratamiento de pacientes con Argina Extreptocícica, fiebre reumática e hipertensión arterial sin embargo en sus historias clínicas no constan datos relacionados con características socioeconomicas de los usuarios por lo que se propuso el tema mencionado anteriormente, En la realización de la investigación intervinieron profesionales de éste centro y también a la División Nacional de Epidemiología quienes con su colaboración nos permitieron diseñar en forma completa el Marco de Refertencia. Nuestro estudio se centro en 152 pacientes diagnosticados con fiebre reumática en los mismos que identificamos catracterísticas socioeconómicas, de salud así como las complicaciones en los años correspondientes de 1977 a 1979. Constituyendo una expoeriancia muy agradable, a la vez muy interesante encontrar en su gran mayoría a niños convertidos en adultos quienes mediante visitas domiciliarias recormamos tiempos anteriores...