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Abstract Background The physiological relevance of cell-to-cell communication mediated by small extracellular vesicle-encapsulated microRNAs (sEV-miRNAs) remains debated because of the limiting representativity of specific miRNAs within the extracellular pool. We hypothesize that sEV-miRNA non-canonical function consisting of the stimulation of Toll-like receptor 7 (TLR7) may rely on a global shift of the sEV cargo rather than on the induction of one or few specific miRNAs. Psoriasis represents an ideal model to test such hypothesis as it is driven by overt activation of TLR7-expressing plasmacytoid dendritic cells (pDCs) following keratinocyte damage. Methods To mimic the onset of psoriasis, keratinocytes were treated with a cocktail of psoriatic cytokines or UV-irradiated. SmallRNA sequencing was performed on sEVs released by healthy and UV-treated keratinocytes. sEV-miRNAs were analyzed for nucleotide composition as well as for the presence of putative TLR7-binding triplets. Primary human pDCs where stimulated with sEVs +/- inhibitors of TLR7 (Enpatoran), of sEV release (GW4869 + manumycin) and of TLR7-mediated pDC activation (anti-BDCA-2 antibody). Secretion of type I IFNs and activation of CD8+T cells were used as readouts. qPCR on psoriatic and healthy skin biopsies was conducted to identify induced miRNAs. Results sEV-miRNAs released by damaged keratinocytes revealed a significantly higher content of TLR7-activating sequences than healthy cells. As expected, differential expression analysis confirmed the presence of miRNAs upregulated in psoriatic skin, including miR203a. More importantly, 76.5% of induced miRNAs possessed TLR7-binding features and among these we could detect several previously demonstrated TLR7 ligands. In accordance with this in silico analysis, sEVs from damaged keratinocytes recapitulated key events of psoriatic pathogenesis by triggering pDCs to release type I interferon and activate cytotoxic CD8+T cells in a TLR7- and sEV-dependent manner. Discussion Our results demonstrate that miR203a is just one paradigmatic TLR7-activating miRNA among the hundreds released by UV-irradiated keratinocytes, which altogether trigger pDC activation in psoriatic conditions. This represents the first evidence that cell damage shifts the miRNA content of sEVs towards a TLR7-activating cargo capable to propagate inflammation and immunity, offering strong support to the physiological role of systemic miRNA-based cell-to-cell communication.

Investigations on acute carbon monoxide (CO) poisoning struggle to highlight a relevant discriminant criterion related to CO poisoning severity for predicting complications, such as delayed neurological syndromes. In this context, it remains difficult to demonstrate the superiority of one method of oxygen (O2) administration over others or to identify the optimal duration of normobaric 100% oxygen (NBO) treatment. Myoglobin, as hemoglobin, are a potential binding site for CO, which could be a source of extravascular CO storage that impacts the severity of CO poisoning. It is not possible in routine clinical practice to estimate this potential extravascular CO storage. Indirect means of doing so that are available in the first few hours of poisoning could include, for example, the carboxyhemoglobin half-life (COHbt1/2), which seems to be influenced itself by the level and duration of CO exposure affecting this store of CO within the body. However, before the elimination of CO can be assessed, the COHbt1/2 toxicokinetic model must be confirmed: research still debates whether this model mono- or bi-compartmental. The second indirect mean could be the assessment of a potential COHb rebound after COHb has returned to 5% and NBO treatment has stopped. Moreover, a COHb rebound could be considered to justify the duration of NBO treatment. On an experimental swine model exposed to moderate CO poisoning (940 ppm for ±118 min until COHb reached 30%), we first confirm that the COHb half-life follows a bi-compartmental model. Secondly, we observe for the first time a slight COHb rebound when COHb returns to 5% and oxygen therapy is stopped. On the basis of these two toxicokinetic characteristics in favor of extravascular CO storage, we recommend that COHbt1/2 is considered using the bi-compartmental model in future clinical studies that compare treatment effectiveness as a potential severity criterion to homogenize cohorts of the same severity. Moreover, from a general toxicokinetic point of view, we confirm that a treatment lasting less than 6 hours appears to be insufficient for treating moderate CO poisoning.

Background. In islet transplantation, the use of dynamic hypothermic preservation techniques is a current challenge. This study compares the efficacy of 3 pancreas preservation methods: static cold storage, hypothermic machine perfusion (HMP), and oxygenated HMP. Methods. A standardized human pancreas split model was employed using discarded organs from both donation after brain death (n = 15) and donation after circulatory death (DCD) (n = 9) donors. The pancreas head was preserved using static cold storage (control group), whereas the tail was preserved using the 3 different methods (study group). Data on donor characteristics, pancreas histology, isolation outcomes, and functional tests of isolated islets were collected. Results. Insulin secretory function evaluated by calculating stimulation indices and total amount of secreted insulin during high glucose stimulation (area under the curve) through dynamic perifusion experiments was similar across all paired groups from both DCD and donation after brain death donors. In our hands, islet yield (IEQ/g) from the pancreas tails used as study groups was higher than that of the pancreas heads as expected although this difference did not always reach statistical significance because of great variability probably due to suboptimal quality of organs released for research purposes. Moreover, islets from DCD organs had greater purity than controls (P ≤ 0.01) in the HMP study group. Furthermore, our investigation revealed no significant differences in pancreas histology, oxidative stress markers, and apoptosis indicators. Conclusions. For the first time, a comparative analysis was conducted, using a split model, to assess the effects of various preservation methods on islets derived from pancreas donors. Nevertheless, no discernible variances were observed in terms of islet functionality, histological attributes, or isolation efficacy. Further investigations are needed to validate these findings for clinical application.

Chromophoric dissolved organic matter (CDOM) is crucial in aquatic ecosystems, influencing light penetration and biogeochemical processes. This study investigates the CDOM variability in seven oligotrophic lakes of North Andean Patagonia using Landsat 8 imagery. An empirical band ratio model was calibrated and validated for the estimation of CDOM concentrations in surface lake water as the absorption coefficient at 440 nm (acdom440, m−1). Of the five atmospheric corrections evaluated, the QUAC (Quick Atmospheric Correction) method demonstrated the highest accuracy for the remote estimation of CDOM. The application of separate models for deep and shallow lakes yielded superior results compared to a combined model, with R2 values of 0.76 and 0.82 and mean absolute percentage errors (MAPEs) of 14% and 22% for deep and shallow lakes, respectively. The spatio-temporal variability of CDOM was characterized over a five-year period using satellite-derived acdom440 values. CDOM concentrations varied widely, with very low values in deep lakes and moderate values in shallow lakes. Additionally, significant seasonal fluctuations were evident. Lower CDOM concentrations were observed during the summer to early autumn period, while higher concentrations were observed in the winter to spring period. A gradient boosting regression tree analysis revealed that inter-lake differences were primarily influenced by the lake perimeter to lake area ratio, mean lake depth, and watershed area to lake volume ratio. However, seasonal CDOM variation was largely influenced by Lake Nahuel Huapi water storage (a proxy for water level variability at a regional scale), followed by precipitation, air temperature, and wind. This research presents a robust method for estimating low to moderate CDOM concentrations, improving environmental monitoring of North Andean Patagonian Lake ecosystems. The results deepen the understanding of CDOM dynamics in low-impact lakes and its main environmental drivers, enhance the ability to estimate lacustrine carbon stocks on a regional scale, and help to predict the effects of climate change on this important variable.

Background and aimsHuman islet preparations designated for research exhibit diverse insulin-secretory profiles. This study aims to assess the impact of donor- and isolation-related factors on in vitro islet secretory function.MethodsA retrospective analysis of 46 isolations from 23 pancreata discarded for clinical transplantation was conducted. In vitro islet secretory function tests were performed on Day 1 and Day 7 of culture. Linear mixed-effects models (LMMs) were employed to investigate the relationships between various predictors characterizing the patient and donor characteristics as well as the isolation effectiveness and two functional outcomes including the islet stimulation index (SI) and area under the insulin curve (AUC). Fixed effects were introduced to represent the main effects of each predictor, and backward elimination was utilized to select the most significant fixed effects for the final model. Interaction effects between the timepoint (Day 7 vs. Day 1) and the predictors were also evaluated to assess whether predictors were associated with the temporal evolution of SI and AUC. Fold-change (Fc) values associated with each predictor were obtained by exponentiating the corresponding coefficients of the models, which were built on log-transformed outcomes.ResultsAnalysis using LMMs revealed that donor body mass index (BMI) (Fc = 0.961, 95% CI = 0.927–0.996, p = 0.05), donor gender (female vs. male, Fc = 0.702, 95% CI = 0.524–0.942, p = 0.04), and donor hypertension (Fc = 0.623, 95% CI = 0.466–0.832, p=

Introduction: Nipple–areolar complex (NAC) reconstruction after breast cancer surgery is challenging and does not always provide optimal long-term esthetic results. Therefore, generating a NAC using tissue engineering techniques, such as a decellularization–recellularization process, is an alternative option to recreate a specific 3D NAC morphological unit, which is then covered with an in vitro regenerated epidermis and, thereafter, skin-grafted on the reconstructed breast.Materials and methods: Human NACs were harvested from cadaveric donors and decellularized using sequential detergent baths. Cellular clearance and extracellular matrix (ECM) preservation were analyzed by histology, as well as by DNA, ECM proteins, growth factors, and residual sodium dodecyl sulfate (SDS) quantification. In vivo biocompatibility was evaluated 30 days after the subcutaneous implantation of native and decellularized human NACs in rats. In vitro scaffold cytocompatibility was assessed by static seeding of human fibroblasts on their hypodermal side for 7 days, while human keratinocytes were seeded on the scaffold epidermal side for 10 days by using the reconstructed human epidermis (RHE) technique to investigate the regeneration of a new epidermis.Results: The decellularized NAC showed a preserved 3D morphology and appeared white. After decellularization, a DNA reduction of 98.3% and the absence of nuclear and HLA staining in histological sections confirmed complete cellular clearance. The ECM architecture and main ECM proteins were preserved, associated with the detection and decrease in growth factors, while a very low amount of residual SDS was detected after decellularization. The decellularized scaffolds were in vivo biocompatible, fully revascularized, and did not induce the production of rat anti-human antibodies after 30 days of subcutaneous implantation. Scaffold in vitro cytocompatibility was confirmed by the increasing proliferation of seeded human fibroblasts during 7 days of culture, associated with a high number of living cells and a similar viability compared to the control cells after 7 days of static culture. Moreover, the RHE technique allowed us to recreate a keratinized pluristratified epithelium after 10 days of culture.Conclusion: Tissue engineering allowed us to create an acellular and biocompatible NAC with a preserved morphology, microarchitecture, and matrix proteins while maintaining their cell growth potential and ability to regenerate the skin epidermis. Thus, tissue engineering could provide a novel alternative to personalized and natural NAC reconstruction.

The tasks that workers in university restaurants (URs) perform involves repetitive movements of the back, lifting excessive weight, remaining in a standing posture for prolonged periods of time, and, thus, an evaluation of the lumbar region of these workers is indicated. Therefore, the objective of this study was to outline the profile of lower back pain in workers from the URs of a Public University in the South of Brazil. This is a cross-sectional, observational, quantitative study, in which participants aged 18 to 59 years, of both sexes, who performed some function within the restaurants of a university in southern Brazil and who reported the presence of lower back pain were selected. Participants were evaluated by a sociodemographic questionnaire, a visual analogue scale (VAS) for pain, a range of motion (ROM) of the lumbar spine evaluation, and the Brazilian Start Back Screening Tool (SBST) questionnaire. Results are presented as absolute and relative frequencies, with a Spearman correlation, and numerical variables as mean and standard deviation or median, minimum, and maximum. Twenty-eight participants were included in the study, 71.4% of whom were women, with a mean age of 36.3 (±10.8) years old. The report of pain above 4 in the VAS was 71.4% of the participants and all had a decrease in ROM in all movements evaluated in the lumbar spine. Most (71.4%) had low risk for psychosocial factors by SBST. The profile of restaurant workers at the evaluated university is characterized by being primarily performed by women, and these workers presented severe pain and limited joint movements in their lumbar region.

Background The development of new metabolic diagnostic imaging PET is changing the history of metastatic prostate cancer (pCa), identifying situations of progression with a low disease burden; radiation treatment of metabolically active oligometastatic sites has been shown to be effective to prolong patient survival. In the scenario of oligometastatic disease to pelvic lymph nodes there is no uniform consensus on treatment volumes and radiotherapy doses to use. Methods We retrospectively assessed a series of 50 patients treated from 2015 to 2021 at our center who presented with recurrent pelvic lymph node pCa disease with 1-3 lymph-nodes lateralized to one side of the pelvis. Patients were treated with intensity modulated Rapid Arc radiotherapy (IMRT), limiting the treatment volume to the chain of the affected side only. During the follow-up, the patients who presented a biochemical recurrence of the disease were evaluated by PET. Results The biochemical progression-free survival and the metastatic progression-free survival were respectively 36% and 49% at 5 years. 22/50 patients presented a documented recurrence on PET. Only one patient presented a relapse within the irradiated volume and no patient presented a relapse on the contralateral pelvic lymph node chain. No patient had gastrointestinal toxicity > grade 1 RTOG. Conclusion Treatment of patients with oligorecurrent (1-3 lesions) pelvic pCa, limiting the volume of irradiation to only one side of the affected lymph node chain, results in good biochemical disease control and presents a low risk of neoplastic contralateral progression.

BackgroundThe COBE 2991 cell processor, commonly used for pancreatic islet isolation, is no longer distributed in Europe, leading to a search for alternative purification procedures with equivalent efficacy. The aim of this study was to evaluate the efficacy of an alternative method based on the discontinuous purification of islets.MethodsThe conventional isolation procedure using a standard continuous islet purification with COBE 2991 of n = 4 human pancreas was compared to n = 8 procedures using a discontinuous purification with a “bottle” method from donors of similar characteristics. Islet equivalents, purity, and dynamic glucose-stimulated insulin secretion were evaluated.ResultsA similar islet yield was obtained using continuous vs. discontinuous purification methods (76,292.5 ± 40,550.44 vs. 79,625 ± 41,484.46 islet equivalents, p = 0.89). Islets from both groups had similar purity (78.75% ± 19.73% vs. 55% ± 18.16%, p = 0.08) and functionality both in terms of stimulation index (3.31 ± 0.83 vs. 5.58 ± 3.38, p = 0.22) and insulin secretion (1.26 ± 0.83 vs. 1.53 ± 1.40 mean AUC, p = 0.73). Moreover, the size of the islets was significantly larger in the discontinuous vs. continuous purification group (19.2% ± 10.3% vs. 45.4% ± 18.8% of islets less than 100 µm, p = 0.0097 and 23.7% ± 5.3% vs. 15.6% ± 5.8% of 200–250 µm islet size, p = 0.03).ConclusionCompared to the conventional purification procedure, discontinuous purification with a bottle method shows similar results with regard to isolation yield and islet secretory function. Furthermore, this alternative technique allows for obtaining larger islets.

Introduction: Durable reconstruction of critical size bone defects is still a surgical challenge despite the availability of numerous autologous and substitute bone options. In this paper, we have investigated the possibility of creating a living bone allograft, using the perfusion/decellularization/recellularization (PDR) technique, which was applied to an original model of vascularized porcine bone graft.Materials and Methods: 11 porcine bone forelimbs, including radius and ulna, were harvested along with their vasculature including the interosseous artery and then decellularized using a sequential detergent perfusion protocol. Cellular clearance, vasculature, extracellular matrix (ECM), and preservation of biomechanical properties were evaluated. The cytocompatibility and in vitro osteoinductive potential of acellular extracellular matrix were studied by static seeding of NIH-3T3 cells and porcine adipose mesenchymal stem cells (pAMSC), respectively.Results: The vascularized bone grafts were successfully decellularized, with an excellent preservation of the 3D morphology and ECM microarchitecture. Measurements of DNA and ECM components revealed complete cellular clearance and preservation of ECM’s major proteins. Bone mineral density (BMD) acquisitions revealed a slight, yet non-significant, decrease after decellularization, while biomechanical testing was unmodified. Cone beam computed tomography (CBCT) acquisitions after vascular injection of barium sulphate confirmed the preservation of the vascular network throughout the whole graft. The non-toxicity of the scaffold was proven by the very low amount of residual sodium dodecyl sulfate (SDS) in the ECM and confirmed by the high live/dead ratio of fibroblasts seeded on periosteum and bone ECM-grafts after 3, 7, and 16 days of culture. Moreover, cell proliferation tests showed a significant multiplication of seeded cell populations at the same endpoints. Lastly, the differentiation study using pAMSC confirmed the ECM graft’s potential to promote osteogenic differentiation. An osteoid-like deposition occurred when pAMSC were cultured on bone ECM in both proliferative and osteogenic differentiation media.Conclusion: Fully decellularized bone grafts can be obtained by perfusion decellularization, thereby preserving ECM architecture and their vascular network, while promoting cell growth and differentiation. These vascularized decellularized bone shaft allografts thus present a true potential for future in vivo reimplantation. Therefore, they may offer new perspectives for repairing large bone defects and for bone tissue engineering.

Premature birth is unpredictable and can result from social, economic and/or health-related factors for women and newborns. This study aimed to analyze the sociodemographic profile of parents of premature newborns admitted to a Neonatal Intensive Care Unit (NICU). Furthermore, it is a descriptive, exploratory, and quantitative study, carried out from March to August 2019 in an NICU. Demographic, socioeconomic, and data related to the newborn and the pregnancy process were collected with a questionnaire containing closed and open questions, which were analyzed and compared with related studies. 31 mothers and 19 fathers participated. Most of them were young and completed high school. Almost all fathers had paying jobs, while only half of mothers worked, and not all of them contributed to social security (INSS). Fathers received 30% more than mothers, even with similar functions and working hours. 47% of the families lived with an income of up to 2 minimum wages per month. Half of the pregnancies were unplanned. Births with gestational ages (GA) ≥ 32 weeks predominated. On average, the mothers had more than 8 prenatal consultations and displayed hypertension as the most prevalent maternal risk factor. Knowing the families of newborns can support measures to promote maternal and child health.

The potential use of porcine islets for transplantation in humans has triggered interest in understanding porcine islet physiology. However, the number of studies dedicated to this topic has remained limited, as most islet physiologists prefer to use the less time-consuming rodent model or the more clinically relevant human islet. An often-overlooked aspect of pig islet physiology is its alpha cell activity and regulation of its glucagon secretion. In vitro islet perifusion is a reliable method to study the dynamics of hormone secretion in response to different stimuli. We thus used this method to quantify and study glucagon secretion from pig islets. Pancreatic islets were isolated from 20 neonatal (14 to 21-day old) and 5 adult (>2 years) pigs and cultured in appropriate media. Islet perifusion experiments were performed 8 to 10 days post-isolation for neonatal islets and 1 to 2 days post-isolation for adult islets. Insulin and glucagon were quantified in perifusion effluent fractions as well as in islet extracts by RIA. Increasing glucose concentration from 1 mM to 15 mM markedly inhibited glucagon secretion independently of animal age. Interestingly, the effect of high glucose was more drastic on glucagon secretion compared to its effect on insulin secretion. In vivo, glucose injection during IVGTT initiated a quick (2-10 minutes) 3-fold decrease of plasmatic glucagon whereas the increase of plasmatic insulin took 20 minutes to become significant. These results suggest that regulation of glucagon secretion significantly contributes to glucose homeostasis in pigs and might compensate for the mild changes in insulin secretion in response to changes in glucose concentration.

OBJETIVO: Avaliar a função musculoesquelética e prevalência de lesões musculoesqueléticas em corredores amadores de rua. MÉTODOS: Selecionados 70 corredores amadores de rua, do sexo masculino, com idade entre 18 e 49 anos, divididos em G10km e G21km. Para identificar as lesões musculoesqueléticas foram avaliados por meio de um questionário semiesetruturado e a função musculoesquelética pelos seguintes testes: força de preensão manual (FPM)para estimar a força muscular global, fleximetria, teste de elevação da perna retificada, teste de sentar e alcançar, mensuração do comprimento de membro inferior, Functional Movement Screen (FMS), Crossover Hop Test for Distance (CHT), Y test, posição de primeira resistência detectada, weight bearing lunge test (WBLT) e flexibilidade da banda iliotibial. RESULTADOS: Em ambos os grupos encontrou-se baixa frequência de lesões, FPM adequada, amplitude de movimento (ADM) de flexão de joelho (fleximetria) e de dorsiflexão (WBLT) diminuídas, encurtamento de músculos isquiotibiais, discrepância de comprimento de membro inferior. Além desses achados, no G21km verificou-se equilíbrio dinâmico alterado e risco de lesão (Y test). CONCLUSÃO: Os corredores de rua apresentaram função musculoesquelética adequada, corroborando com a baixa prevalência de lesões musculoesqueléticas observada.

BackgroundEarly allograft dysfunction (EAD) following liver transplantation (LT) remains a major threat to the survival of liver grafts and recipients. In animal models, it is shown that hepatic ischemia-reperfusion injury (IRI) triggers phosphorylation of Mixed Lineage Kinase domain-like protein (pMLKL) inducing necroptotic cell death. However, the clinical implication of pMLKL-mediated cell death in human hepatic IRI remains largely unexplored. In this study, we aimed to investigate the expression of pMLKL in human liver grafts and its association with EAD after LT.MethodsThe expression of pMLKL was determined by immunohistochemistry in liver biopsies obtained from both human and rat LT. Human liver biopsies were obtained at the end of preservation (T0) and ~1 hour after reperfusion (T1). The positivity of pMLKL was quantified electronically and compared in rat and human livers and post-LT outcomes. Multiplex immunofluorescence staining was performed to characterize the pMLKL-expressing cells.ResultsIn the rat LT model, significant pMLKL expression was observed in livers after IRI as compared to livers of sham-operation animals. Similarly, the pMLKL score was highest after IRI in human liver grafts (in T1 biopsies). Both in rats and humans, the pMLKL expression is mostly observed in the portal triads. In grafts who developed EAD after LT (n=24), the pMLKL score at T1 was significantly higher as compared to non-EAD grafts (n=40). ROC curve revealed a high predictive value of pMLKL score at T1 (AUC 0.70) and the ratio of pMLKL score at T1 and T0 (pMLKL-index, AUC 0.82) for EAD. Liver grafts with a high pMLKL index (>1.64) had significantly higher levels of serum ALT, AST, and LDH 24 hours after LT compared to grafts with a low pMLKL index. Multivariate logistical regression analysis identified the pMLKL-index (Odds ratio=1.3, 95% CI 1.1-1.7) as a predictor of EAD development. Immunohistochemistry on serial sections and multiplex staining identified the periportal pMLKL-positive cells as portal fibroblasts, fibrocytes, and a minority of cholangiocytes.ConclusionPeriportal pMLKL expression increased significantly after IRI in both rat and human LT. The histological score of pMLKL is predictive of post-transplant EAD and is associated with early liver injury after LT. Periportal non-parenchymal cells (i.e. fibroblasts) appear most susceptible to pMLKL-mediated cell death during hepatic IRI.

ABSTRACT The aim of this study was to evaluate the evidence of application of short-wave diathermy (SWD) in individuals with chronic low back pain (CLBP) and its prescription parameters. The data sources (MEDLINE, PubMed, LILACS, DARE, PsycINFO, AusportMed, SciELO, PEDro and the Cochrane Central Register of Controlled Trials) were systematically searched for articles published up to December 2017. Randomized and non-randomized clinical trial studies that investigated the effect of the application of SWD on CLBP were selected. Two independent reviewers assessed the risk of bias in studies using the Jadad and the Downs & Black scales. Five studies (731 patients) were included, all of whom presented improvements in CLBP. The majority used continuous mode SWD, with only one using pulsed mode (82 and 200 Hz). SWD produced improvements in CLBP, but there is limited evidence for its application, and a lack of standardization of the parameters used. Furthermore, the results that address this topic should be interpreted with caution due to their low methodological quality and limited number. Level of evidence II; Systematic review.

The lack of viability of massive bone allografts for critical-size bone defect treatment remains a challenge in orthopedic surgery. The literature has reviewed the advantages of a multi-combined treatment with the synergy of an osteoconductive extracellular matrix (ECM), osteogenic stem cells, and growth factors (GFs). Questions are still open about the need for ECM components, the influence of the decellularization process on the latter, the related potential loss of function, and the necessity of using pre-differentiated cells. In order to fill in this gap, a bone allograft surrounded by an osteogenic membrane made of a decellularized collagen matrix from human fascia lata and seeded with periosteal mesenchymal stem cells (PMSCs) was analyzed in terms of de-/recellularization, osteogenic properties, PMSC self-differentiation, and angiogenic potential. While the decellularization processes altered the ECM content differently, the main GF content was decreased in soft tissues but relatively increased in hard bone tissues. The spontaneous osteogenic differentiation was necessarily obtained through contact with a mineralized bone matrix. Trying to deepen the knowledge on the complex matrix–cell interplay could further propel these tissue engineering concepts and lead us to provide the biological elements that allow bone integration in vivo.

This essay focuses on the use of preliminary reference to the European Court of Justice pursuant to art. 267 TFEU in order to protect the independence and autonomy of National judiciaries, in particular within Member States such as Hungary and Poland. In the first part, the author looks at two specific cases in which the possibility for national judges to refer to the Court of Justice was limited by the action of the two illiberal governments. Secondly, also in relation to the decisions of the Brussels Court on the subject, the analysis focuses on the need to protect the preliminary reference in the context of judicial cooperation and on the role of European institutions, in particular the Commission, in doing so.

RESUMO Objetivo: Verificar a ocorrência e as características de atelectasias, opacidades, hipotransparências e infiltrados pulmonares evidenciados ao raio X de tórax dos recém-nascidos prematuros, de uma unidade de terapia intensiva neonatal. Métodos: Trata-se de estudo observacional transversal. No período de agosto a dezembro de 2017 foram analisadas todas as radiografias de tórax de recém-nascidos. Foram incluídas no estudo as radiografias de tórax de recém-nascidos prematuros com idade gestacional até 36 semanas, no período neonatal que apresentassem alterações evidentes na imagem ou suspeita de alterações, que fossem confirmadas após laudo do médico radiologista. As alterações radiológicas foram associadas com possíveis fatores predisponentes. Resultados: No período, foram realizadas 450 radiografias nos recém-nascidos prematuros, sendo que, em 37, foram descritas quatro alterações: 12 (2,66%) descritas como opacidades, 11 (2,44%) como atelectasias, 10 (2,22%) como infiltrados pulmonares e 4 (0,88%) como hipotransparências. Observou-se maior ocorrência das atelectasias no pulmão direito (81,8%). Dentre as radiografias com alterações, 25 (67,6%) recém-nascidos estavam sob o uso da ventilação mecânica invasiva. Conclusão: Considerando o laudo radiológico, as alterações observadas têm ocorrências sem diferença estatisticamente significante. A atelectasia não foi a alteração mais encontrada. Os fatores que podem ter predisposto ao aparecimento das alterações foram a prematuridade extrema, o baixo peso, o sexo masculino, o mal posicionamento da cânula endotraqueal e o uso de ventilação mecânica invasiva.

Background. The aim of this feasibility study was to determine an alternative oxygenation technique (easy, cheap, and compatible with air transport) for membrane oxygenation during hypothermic machine perfusion (HMP) to improve early graft function in a porcine ischemia-reperfusion autotransplant model. Methods. The left kidney of a ±40- kg pig was exposed to 30 minutes of warm ischemia before 22 hours of preservation and autotransplantation. In the experimental group, oxygenation of the perfusate during HMP was obtained by direct bubble and 30-minute surface oxygenation at start and 1-hour end ischemic (n = 4) and outcome measures compared with historical HMP without active oxygenation (n = 6), 22-hour continuous oxygenated HMP (HMPO2) (n = 8), and 2-hour HMPO2 + 20-hour HMP (n = 6) using membrane oxygenation in both historical oxygenated control groups. Results. Brief bubble and 30-minute surface oxygenation of the perfusate effectively maintained supraphysiological Po2 levels during the first 2 hours of HMP with improved flow dynamics. Although the metabolic profile of the perfusate (ie, flavin mononucleotide) and tissue (ie, glutamate, ATP) after brief O2 uploading at the start of HMP seemed to be slightly better with the use of a membrane oxygenator compared with bubble and interrupted surface oxygenation, both techniques yielded similar, superior early graft function when compared with HMP without active oxygenation. Conclusions. The data presented in this feasibility study support the conclusion that brief bubble and intermittent surface oxygenation could be an alternative oxygenation technique during HMP to achieve an improved kidney graft function compared with HMP without active oxygenation and similar functional outcome when compared with membrane HMPO2.

Cell encapsulation could overcome limitations of free islets transplantation but is currently limited by inefficient cells immune protection and hypoxia. As a response to these challenges, we tested in vitro and in vivo the safety and efficacy of a new macroencapsulation device named MailPan ® . Membranes of MailPan ® device were tested in vitro in static conditions. Its bio-integration and level of oxygenation was assessed after implantation in non-diabetic rats. Immune protection properties were also assessed in rat with injection in the device of allogeneic islets with incompatible Major Histocompatibility Complex. Finally, function was assessed in diabetic rats with a Beta cell line injected in MailPan ® . In vitro, membranes of the device showed high permeability to glucose, insulin, and rejected IgG. In rat, the device displayed good bio-integration, efficient vascularization, and satisfactory oxygenation (>5%), while positron emission tomography (PET)-scan and angiography also highlighted rapid exchanges between blood circulation and the MailPan ® . The device showed its immune protection properties by preventing formation, by the rat recipient, of antibodies against encapsulated allogenic islets. Injection of a rat beta cell line into the device normalized fasting glycemia of diabetic rat with retrieval of viable cell clusters after 2 months. These data suggest that MailPan ® constitutes a promising encapsulation device for widespread use of cell therapy for type 1 diabetes.

Passive daytime radiative cooling presents a promising low-cost refrigeration solution but has thus far relied on specialized nanophotonic structures. Here Bhatia et al. show a directional approach that decouples solar reflectance and infrared emission to achieve superior cooling performance.

A saúde é um fator essencial para o desenvolvimento do ser humano, desta forma os projetos de promoção à saúde do adolescente buscam o desenvolvimento de hábitos saudáveis no sentido do auto-cuidado. Objetivo: promover à saúde de um grupo de atletas de 11 a 14 anos do Colégio Militar do Paraná, envolvendo temas como prática de exercício físico, alimentação saudável, qualidade do sono e saúde ambiental. Métodos: o estudo foi desenvolvido no período de 2016, consistindo em 4 encontros com duração de 30 minutos, envolvendo práticas de promoção e educação em saúde subdividas em conceitual teórico e prático. Resultados: os estudantes praticavam exercício físico e a maioria acreditava possuir uma alimentação saudável e uma boa qualidade de sono, porém, após a realização das atividades, relataram ter aprendido mais sobre a sua própria saúde. Conclusão: existe a necessidade de programas de promoção à saúde do adolescente dentro das escolas.

El objetivo de este trabajo fue estudiar la estructura del ensamblaje de macroinvertebrados bentónicos de la laguna del Parque Unzué y discutir sobre su calidad de agua. El área de estudio se encuentra en la ciudad de Gualeguaychú (Entre Ríos, Argentina), sobre la margen izquierda del río homónimo. La laguna tiene un área de 2750 m2 y una profundidad máxima de 1,70 m. Durante 2016-2017, se muestreó en 3 puntos (n=10), se midieron parámetros in situ y se recolectaron muestras de bentos, se caracterizaron los sedimentos y los macroinvertebrados fueron identificados hasta el menor nivel taxonómico posible y se los agrupó en grupos funcionales alimenticios (GFA). Se calculó la diversidad, la diversidad máxima esperada (Hmax) y los números de Hill. No se hallaron diferencias estadísticamente significativas (p

Abstract Background Glutaraldehyde fixed xenogeneic heart valve prosthesis are hindered by calcification and lack of growth potential. The aim of tissue decellularization is to remove tissue antigenicity, avoiding the use of glutaraldehyde and improve valve integration with low inflammation and host cell recolonization. In this preliminary study, we investigated the efficacy of a NaOH-based process for decellularization and biocompatibility improvement of porcine pulmonary heart valves in comparison to a detergent-based process (SDS-SDC0, 5%). Methods Native cryopreserved porcine pulmonary heart valves were treated with detergent and NaOH-based processes. Decellularization was assessed by Hematoxylin and eosin/DAPI/alpha-gal/SLA-I staining and DNA quantification of native and processed leaflets, walls and muscles. Elongation stress test investigated mechanical integrity of leaflets and walls (n = 3 tests/valve component) of valves in the native and treated groups (n = 4/group). Biochemical integrity (collagen/elastin/glycosaminoglycans content) of leaflet-wall and muscle of the valves (n = 4/group) was assessed and compared between groups with trichrome staining (Sirius Red/Miller/Alcian blue). Secondly, a preliminary in vivo study assessed biocompatibility (CD3 and CD68 immunostaining) and remodeling (Hematoxylin and eosin/CD31 and ASMA immunofluorescent staining) of NaOH processed valves implanted in orthotopic position in young Landrace pigs, at 1 (n = 1) and 3 months (n = 2). Results Decellularization was better achieved with the NaOH-based process (92% vs 69% DNA reduction in the wall). Both treatments did not significantly alter mechanical properties. The detergent-based process induced a significant loss of glycosaminoglycans (p

Abstract: The objective of this work was to evaluate remaining P compared with soil clay content as a P buffer index to classify P extracted by the Mehlich-1 (M1) and Mehlich-3 (M3) methods in soils from the state of Rio Grande do Sul, Brazil. The experiment was carried out in a completely randomized design with five P2O5 rates (0, 50, 100, 200, and 400 mg kg-1) and two sucessive corn crops, and three replicates, in 20 representative soils of the state. P extracted by M1 and M3 before crop planting was adjusted to P contents in biomass, considering soil buffer capacity. The division of soils into different buffering classes, based on soil clay or remaining P, improved the capacity of estimating soil available P of both methods. However, there was no difference among the correlation coefficients obtained by classifying soils according to the evaluated indexes (remaining P or soil clay) for both M1 and M3 methods. Remaining P is a viable alternative to replace soil clay content to classify soil P extracted with the M1 and M3 methods.

Knowledge of rhizosphere processes is essential for characterizing soil nutrient availability. Our objective in this controlled-climate study was to evaluate phosphorus (P) dynamics in the rhizosphere of juvenile maize (Zea mays L.) grown in soil amended with compost. Maize seedlings were transplanted at the two-leaf stage, and grown for five days. Soil (550 g) collected from the 0-5 cm layer of field plots was placed in one chamber of a vertically divided mini-rhizotron, and soil from the 5-10 cm layer was placed in the other chamber. Micro-suction cups (15 per chamber) were used to collect rhizosphere soil solution that was subsequently analyzed for P and pH. Initially, P concentrations in rhizosphere solution did not differ with distance from the root surface and were similar in the two soil layers. These results reflect the high levels of available P in both soil layers. Solution pH also was similar among the soil layers and distances from the root surface. The soil used in this study was calcareous and strongly buffered against changes in solution pH. After five days, both soil solution P and pH had decreased in both soil layers. However, only soil solution P was affected by distance from the root surface, confirming the pH buffering of the soil. Maize shoot dry matter production was similar in the two soil layers (0.65 g plant-1 in the 0-5 cm layer and 0.60 g plant-1 in the 5-10 cm layer), suggesting that soil P stratification in this soil under no-tillage had little effect on plant growth.

In the context of experimental development of vascularized composite tissue allotransplantation (VCA), the auricular model has been described in rats and humans, but not in pigs. A porcine ear transplant, however, represents an interesting experimental composite facial subunit allotransplant because of its reduced morbidity and its translational nature but mostly because it is composed of very different tissues. In this perspective, we studied the anatomical and surgical aspects of an ear subunit VCA in pigs. Our study was performed on 18 pigs: auricular and cervical regions were dissected without preparation (n = 12) or after latex injections (n = 2) in the common carotid artery (CCA). The angiosomes of the caudal auricular artery and superficial temporal artery were studied with selective injections (n = 2). The surgical harvesting protocol was established using the caudal auricular artery as arterial pedicle, and tissue perfusion was studied with injection of Indian ink (n = 1) and angio-CT (n = 1). Finally, two in vivo orthotopic allotransplantations in four pigs were performed, followed by a short observation period. The caudal auricular artery was shown to be the dominant artery to the auricle ensure complete ear perfusion. Venous drainage relied on the caudal and rostral auricular veins, dissected down to the maxillary and external jugular veins. In vivo allotransplantations confirmed proper auricular vascularization on the sole caudal auricular artery under physiological conditions. We have described a new subunit model for experimental face VCA in large animals. Our study reports a reliable harvesting method and easily performed transplantation with a single-based arterial pedicle.

La acelerada pérdida de calidad de agua en ríos y arroyos, está asociada con pérdida de biodiversidad, por tanto la determinación de parámetros de calidad de agua en estos sistemas acuáticos es fundamental y urgente. El objetivo del trabajo fue estudiar la calidad de agua de cuatro arroyos de la cuenca del río Gualeguaychú, utilizando bioindicadores. El área de estudio se encuentra en la Pampa Mesopotámica, donde se desarrollan actividades agrícolas y ganaderas, y está sujeta a una profunda transformación de sus ecosistemas desde hace dos décadas. Entre 2013-2015, se estudiaron los arroyos García (GA), Los Perros (LP), Santa Bárbara (SB) y La Cañada (LC). Se realizaron muestreos periódicos determinando variables fisicoquímicas, de ribera, macroinvertebrados bentónicos y de audal. Se aplicaron los índices Biótico de Familias (IBF), Biological Monitoring Working Party (BMWP), el Índice Biótico Pampeano (IBPamp) y el Índice de Calidad de Ribera (ICR). Se recolectaron e identificaron 10007 organismos de 67 taxones. Los índices BMWP e IBF determinaron que existe contaminación de severa a intermedia, el IBPamp determinó escasa contaminación. Según el ICR, los arroyos LP, GA y SB, poseen alteraciones en la ribera; LC, no posee alteraciones. Los índices bióticos de calidad de agua y ribera resultaron una herramienta muy valiosa para determinar el estado ecológico de los cuerpos de agua de la zona, sin embargo, debido a la particularidades de la Pampa Mesopotámica, es necesario realizar ajustes que permitan una mejor interpretación de la diversidad regional y de sus variaciones en el espacio y en el tiempo.

Persistent and excessive cytokine production is a hallmark of autoimmune diseases and may play a role in disease pathogenesis and amplification. Therefore, cytokine neutralization is a useful therapeutic strategy to treat immune-mediated conditions. MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate gene expression in diverse biological processes. Altered miRNA levels are observed in most autoimmune diseases and are recognized to influence autoimmunity through different mechanisms. Here, we review the impact of altered miRNA levels on the expression of cytokines that play a relevant pathogenic role in autoimmunity, namely primary pro-inflammatory cytokines, the IL-17/IL-23 axis, type I interferons and IL-10. Regulation can be either “direct” on the target cytokine, or “indirect,” meaning that one given miRNA post-transcriptionally regulates the expression of a protein that in turn influences the level of the cytokine. In addition, miRNAs associated with extracellular vesicles can regulate cytokine production in neighboring cells, either post-transcriptionally or via the stimulation of innate immune RNA-sensors, such as Toll-like receptors. Because of their tremendous potential as physiological and pathological regulators, miRNAs are in the limelight as promising future biopharmaceuticals. Thus, these studies may lead in the near future to the design and testing of therapeutic miRNAs as next generation drugs to target pathogenic cytokines in autoimmunity.

Abstract The objective of this work was to assess the risk of phosphorus losses by runoff through an index based on the degree of P saturation (DPS), in cropland soils of Southern Brazil. Sixty-five highly representative cropland soils from the region were evaluated. Three labile P forms were measured (Mehlich-1, Mehlich-3, and ammonium oxalate), and four P sorption indexes were tested (phosphorus single sorption point and Fe+Al determined with the three extractors). Water-extractable P (WEP) was used as an index of P susceptibility to losses by surface runoff. The DPS was determined from the ratio between labile P and each sorption index. DPS values obtained from the ratio between Mehlich-1 P and the single P sorption point ranged from 1 to 25%, whereas those from Mehlich-1 P and Fe+Al (ammonium oxalate) ranged from 1 to 55%. All DPS types were highly correlated with WEP. From a practical stand point, the DPS obtained with both P and Fe+Al extracted with Mehlich-1 can be used to estimate the risk of P losses by runoff in soils of Southern Brazil.

ABSTRACT This study analyzed the effects of resistive exercise and/or stretching in the muscle morphology of ovariectomized rats. Seventy-five female Wistar rats (218±22g) were divided into 8 groups: Control (CONTROL, n=5); Ovariectomy/Hysterectomy (OH, n=10): ovariectomized/hysterectomized and then waited 8 weeks for the onset of osteopenia. Stretching (STRET, n=10): manual stretching of the soleus muscle (10 repetitions of 60s), twice weekly; Ovariectomy/Hysterectomy and Stretching (OH+STRET, n=10); Progressive Loading (PL, n=10): 4 sets of 10 repetitions, initial load 50% of body weight, increasing weekly until 70%, three times a week. Progressive Loading and Stretching (PL+STRET, n=10); Ovariectomy/Hysterectomy and Progressive loading exercise (OH+PL, n=10); Ovariectomy/Hysterectomy, Progressive Loading and Stretching (OH+PL+STRET, n=10). Stretching and/or progressive loading were performed for 6 weeks. After 14 weeks, the female rats were euthanized to remove the soleus muscle. The following variables were assessed: body weight; muscle weight and length; number and length of sarcomeres and the muscle fiber cross-sectional area (MFCSA). Comparisons between the groups were performed with ANOVA one-way post hoc Tukey or Kruskall-Wallis (p