Polymeric Nanoparticles that Combine Dexamethasone and Naproxen for the Synergistic Inhibition of Il12b Transcription in Macrophages

Recent studies have demonstrated in vivo synergistic immunosuppressive and anti-inflammatory capacity of dexamethasone (Dx) and naproxen (NAP) in collagen-induced arthritis (CIA) rats. However, the molecular basis of this synergistic effect is barely understood. The low solubility of these drugs and their adverse effects hamper their efficacy on the treatment of inflammatory processes making nanoparticulated systems promising candidates to overcome these drawbacks. The aim of this work is the preparation of polymeric nanoparticles (NPs) that combine NAP and Dx in different concentrations, and the evaluation of the expression of key genes related to autoimmune diseases like CIA. To do so, self-assembled polymeric NPs that incorporate covalently-linked NAP and physically entrapped Dx are designed to have hydrodynamic properties that, according to bibliography, may improve retention and colocalization of both drugs at inflammation sites. The rapid uptake of NPs by macrophages is demonstrated using coumarine-6-loaded NPs. Dx is efficiently encapsulated and in vitro biological studies demonstrate that the Dx-loaded NAP-bearing NPs are noncytotoxic and reduce lipopolysaccharide- induced NO released levels at any of the tested concentrations. Moreover, at the molecular level, a significant synergistic reduction of Il12b transcript gene expression when combining Dx and NAP is demonstrated.Authors would like to thank the Spanish Ministry of Science, Innovation and Universities (MAT2017-84277-R and SAF2017-82223-R) and CIBER-BBN for the financial support of this project. E.E.-C. and Y.P. would like to thank the training program for Academic Staff (FPU15/06109 and FPU15/06170, respectively) of the Spanish Ministry of Education Culture and Sport. The kind support by Alvaro González- Gómez, Rosana Ramírez, and David Gómez, in the synthesis, cell culture and SEM experiments is greatly appreciatedPeer reviewe

Longitudinal change in proteinuria and kidney outcomes in C3 glomerulopathy

11 p.-4 fig.-4 tab.Introduction: The association between a change in proteinuria over time and its impact in kidney prognosis has not been analyzed in C3 glomerulopathy. This study aims to investigate the association between the longitudinal change in proteinuria and the risk of kidney failure.Methods: Retrospective, multicenter observational cohort study in 35 nephrology departments belonging to the Spanish Group for the Study of Glomerular Diseases (GLOSEN). Patients diagnosed with C3 glomerulopathy between 1995 and 2020 were enrolled. A joint modeling of linear mixed-effects models was applied to assess the underlying trajectory of a repeatedly measured proteinuria, and a Cox model to evaluate the association of this trajectory with the risk of kidney failure.Results: The study group consisted of 85 patients, 70 C3 glomerulonephritis and 15 dense deposit disease, with a median age of 26 years (range 13-41). During a median follow-up of 42 months, 25 patients reached kidney failure. The longitudinal change in proteinuria showed a strong association with the risk of this outcome, with a doubling of proteinuria levels resulting in a 2.5-fold increase of the risk. A second model showed that a ≥ 50% proteinuria reduction over time was significantly associated with a lower risk of kidney failure (HR: 0.79; 95% CI : 0.56-0.97; p < 0.001). This association was also found when the ≥50% proteinuria reduction was observed within the first 6 and 12 months of follow-up.Conclusion: The longitudinal change in proteinuria is strongly associated with the risk of kidney failure. The change in proteinuria over time can provide clinicians a dynamic prediction of kidney outcomes.This study was supported by the Instituto de Salud Carlos III/Fondo Europeo de Desarrollo Regional (ISCIII/FEDER) grant PI16/01685 and PI19/1624, and Red de Investigación Renal (RedInRen) (RD12/0021/0029) (to M.P.), the Autonomous Region of Madrid (S2017/BMD-3673) (to M.P.); E.G.d.J. was supported by the Spanish ‘Ministerio de Ciencia, Innovación y Universidades’ (RYC-2013-13395 and RTI2018-095955-B-100); S.R.d.C. was supported by Ministerio de Economía y Competitividad/FEDER grant SAF2015-66287R and Autonomous Region of Madrid grant S2017/BMD3673.Peer reviewe

Spatiotemporal Characteristics of the Largest HIV-1 CRF02_AG Outbreak in Spain: Evidence for Onward Transmissions

Background and Aim: The circulating recombinant form 02_AG (CRF02_AG) is the predominant clade among the human immunodeficiency virus type-1 (HIV-1) non-Bs with a prevalence of 5.97% (95% Confidence Interval-CI: 5.41–6.57%) across Spain. Our aim was to estimate the levels of regional clustering for CRF02_AG and the spatiotemporal characteristics of the largest CRF02_AG subepidemic in Spain.Methods: We studied 396 CRF02_AG sequences obtained from HIV-1 diagnosed patients during 2000–2014 from 10 autonomous communities of Spain. Phylogenetic analysis was performed on the 391 CRF02_AG sequences along with all globally sampled CRF02_AG sequences (N = 3,302) as references. Phylodynamic and phylogeographic analysis was performed to the largest CRF02_AG monophyletic cluster by a Bayesian method in BEAST v1.8.0 and by reconstructing ancestral states using the criterion of parsimony in Mesquite v3.4, respectively.Results: The HIV-1 CRF02_AG prevalence differed across Spanish autonomous communities we sampled from (p &lt; 0.001). Phylogenetic analysis revealed that 52.7% of the CRF02_AG sequences formed 56 monophyletic clusters, with a range of 2–79 sequences. The CRF02_AG regional dispersal differed across Spain (p = 0.003), as suggested by monophyletic clustering. For the largest monophyletic cluster (subepidemic) (N = 79), 49.4% of the clustered sequences originated from Madrid, while most sequences (51.9%) had been obtained from men having sex with men (MSM). Molecular clock analysis suggested that the origin (tMRCA) of the CRF02_AG subepidemic was in 2002 (median estimate; 95% Highest Posterior Density-HPD interval: 1999–2004). Additionally, we found significant clustering within the CRF02_AG subepidemic according to the ethnic origin.Conclusion: CRF02_AG has been introduced as a result of multiple introductions in Spain, following regional dispersal in several cases. We showed that CRF02_AG transmissions were mostly due to regional dispersal in Spain. The hot-spot for the largest CRF02_AG regional subepidemic in Spain was in Madrid associated with MSM transmission risk group. The existence of subepidemics suggest that several spillovers occurred from Madrid to other areas. CRF02_AG sequences from Hispanics were clustered in a separate subclade suggesting no linkage between the local and Hispanic subepidemics

Prevalence, associated factors and outcomes of pressure injuries in adult intensive care unit patients: the DecubICUs study

Funder: European Society of Intensive Care Medicine; doi: http://dx.doi.org/10.13039/501100013347Funder: Flemish Society for Critical Care NursesAbstract: Purpose: Intensive care unit (ICU) patients are particularly susceptible to developing pressure injuries. Epidemiologic data is however unavailable. We aimed to provide an international picture of the extent of pressure injuries and factors associated with ICU-acquired pressure injuries in adult ICU patients. Methods: International 1-day point-prevalence study; follow-up for outcome assessment until hospital discharge (maximum 12 weeks). Factors associated with ICU-acquired pressure injury and hospital mortality were assessed by generalised linear mixed-effects regression analysis. Results: Data from 13,254 patients in 1117 ICUs (90 countries) revealed 6747 pressure injuries; 3997 (59.2%) were ICU-acquired. Overall prevalence was 26.6% (95% confidence interval [CI] 25.9–27.3). ICU-acquired prevalence was 16.2% (95% CI 15.6–16.8). Sacrum (37%) and heels (19.5%) were most affected. Factors independently associated with ICU-acquired pressure injuries were older age, male sex, being underweight, emergency surgery, higher Simplified Acute Physiology Score II, Braden score 3 days, comorbidities (chronic obstructive pulmonary disease, immunodeficiency), organ support (renal replacement, mechanical ventilation on ICU admission), and being in a low or lower-middle income-economy. Gradually increasing associations with mortality were identified for increasing severity of pressure injury: stage I (odds ratio [OR] 1.5; 95% CI 1.2–1.8), stage II (OR 1.6; 95% CI 1.4–1.9), and stage III or worse (OR 2.8; 95% CI 2.3–3.3). Conclusion: Pressure injuries are common in adult ICU patients. ICU-acquired pressure injuries are associated with mainly intrinsic factors and mortality. Optimal care standards, increased awareness, appropriate resource allocation, and further research into optimal prevention are pivotal to tackle this important patient safety threat

Prevalence, associated factors and outcomes of pressure injuries in adult intensive care unit patients: the DecubICUs study

Funder: European Society of Intensive Care Medicine; doi: http://dx.doi.org/10.13039/501100013347Funder: Flemish Society for Critical Care NursesAbstract: Purpose: Intensive care unit (ICU) patients are particularly susceptible to developing pressure injuries. Epidemiologic data is however unavailable. We aimed to provide an international picture of the extent of pressure injuries and factors associated with ICU-acquired pressure injuries in adult ICU patients. Methods: International 1-day point-prevalence study; follow-up for outcome assessment until hospital discharge (maximum 12 weeks). Factors associated with ICU-acquired pressure injury and hospital mortality were assessed by generalised linear mixed-effects regression analysis. Results: Data from 13,254 patients in 1117 ICUs (90 countries) revealed 6747 pressure injuries; 3997 (59.2%) were ICU-acquired. Overall prevalence was 26.6% (95% confidence interval [CI] 25.9–27.3). ICU-acquired prevalence was 16.2% (95% CI 15.6–16.8). Sacrum (37%) and heels (19.5%) were most affected. Factors independently associated with ICU-acquired pressure injuries were older age, male sex, being underweight, emergency surgery, higher Simplified Acute Physiology Score II, Braden score 3 days, comorbidities (chronic obstructive pulmonary disease, immunodeficiency), organ support (renal replacement, mechanical ventilation on ICU admission), and being in a low or lower-middle income-economy. Gradually increasing associations with mortality were identified for increasing severity of pressure injury: stage I (odds ratio [OR] 1.5; 95% CI 1.2–1.8), stage II (OR 1.6; 95% CI 1.4–1.9), and stage III or worse (OR 2.8; 95% CI 2.3–3.3). Conclusion: Pressure injuries are common in adult ICU patients. ICU-acquired pressure injuries are associated with mainly intrinsic factors and mortality. Optimal care standards, increased awareness, appropriate resource allocation, and further research into optimal prevention are pivotal to tackle this important patient safety threat

Correction to: Prevalence, associated factors and outcomes of pressure injuries in adult intensive care unit patients: the DecubICUs study

The original version of this article unfortunately contained a mistake

Nanopartículas poliméricas para el tratamiento de procesos inflamatorios

Mención Internacional en el título de doctorLa inflamación, en su forma controlada, es un mecanismo crucial para la supervivencia de un organismo. Sin embargo, alteraciones en esta respuesta biológica provocan un estado de inflamación crónica, el cual es el principal componente de las denominadas enfermedades crónicas inflamatorias. Estas enfermedades son la principal causa de mortalidad en el mundo; causando también discapacidad, pérdida de calidad de vida y altos costes económicos. Los fármacos actuales para el tratamiento de estas enfermedades tienen efectos adversos importantes que limitan su uso prolongado. Además, su hidrofobicidad, baja biodisponibilidad y falta de vectorización activa reducen su efectividad. Para resolver este problema, los investigadores se han centrado en la preparación de derivados más eficientes de estos fármacos, terapias combinadas con efecto sinérgico, o sistemas de liberación controlada nanométricos. Estos últimos, son la alternativa más adecuada a las formulaciones tradicionales ya que permite reducir las dosis aumentando el efecto terapéutico de los fármacos. Además, los protegen de ambientes hostiles, facilitan su liberación a través de distintas barreras anatómicas y permiten una co-liberación vectorizada y controlada de los mismos. Esta tesis se centra en el diseño y preparación de sistemas de liberación controlada catiónicos basados en anti-inflamatorios no esteroideos (AINEs) para mejorar las actuales terapias contra diferentes enfermedades inflamatorias crónicas. Los capítulos 3 y 4 se centran en testar la capacidad de nuevos sistemas de co-liberación controlada de AINEs, naproxeno o ketoprofeno, anclado covalentemente, y dexametasona, encapsulada físicamente, para controlar la respuesta inflamatoria y, más específicamente, reducir la expresión génica de marcadores de enfermedades inflamatorias autoinmunes (i.e. IL12, IL23 y TNF-α). Con este objetivo, estos capítulos describen la síntesis de dos derivados metacrílicos de AINEs y su copolimerización con un monómero hidrofílico, 1-vinyl-imidazol, para obtener estructuras anfifílicas de pseudo-gradiente que se autoensamblen formando nanoparticulas en medio acuoso. La citotoxicidad y capacidad de reducir la expresión de marcadores de inflamación de los sistemas se evaluó in vitro, utilizando macrófagos como célula modelo del sistema inmune. De estos capítulos concluimos que la terapia combinada con naproxeno y dexametasona tiene efecto sinérgico en la reducción de la expresión génica de la interleuquina (IL)12-p40 en macrófagos mediante mecanismos independientes de ciclooxigenasas (COX), ya que este efecto no se observó para la combinación de la dexametasona con el ketoprofeno, el cual es un inhibidor de COX más potente. Este sistema ofrece una alternativa más económica a las actuales terapias anti-IL12-p40 para el tratamiento de enfermedades inflamatorias autoinmunes. El capítulo 5 se centra en la preparación de recubrimientos anti-inflamatorios para modular la respuesta de los macrófagos a un cuerpo extraño como, por ejemplo, un implante. Los recubrimientos se prepararon mediante la técnica de deposición capa a capa combinando heparina (Hep, polianión), un glicosaminoglicano con capacidad anti-inflamatoria inmediata demostrada, con las nanopartículas catiónicas basadas en naproxeno previamente descritas. Se realizó una extensa caracterización físico-química de los sistemas para correlacionar sus propiedades (mojabilidad, comportamiento elástico, carga superficial…) con el comportamiento celular. Además, se estudió in vitro, a corto plazo, el efecto en la adhesión de macrófagos y la producción de IL1β y, a largo plazo, en la formación de células gigantes a partir de macrófagos. Las nanopartículas se inmovilizaron de forma efectiva dando lugar a una superficie hidrofílica y aniónica que comprometía la adhesión de macrófagos a corto plazo. Se atribuyó a la heparina la capacidad de los recubrimientos de reducir la producción de IL1β a corto plazo, mientras que la liberación controlada en el tiempo de las nanopartículas permitió reducir la formación de células gigantes a largo plazo. En definitiva, este capítulo sirve como prueba de concepto para establecer nuevos recubrimientos que puedan suprimir la respuesta inflamatoria a cuerpo extraño y modular la reacción de los macrófagos a los biomateriales a más largo plazo que las terapias actuales. El capítulo 6 se centra en la preparación de nanopartículas basadas en naproxeno recubiertas con ácido hialurónico para su vectorización activa a las células madre cancerígenas de mama que sobre expresen el CD44 mejorando la capacidad anticancerígena del fármaco libre y la hemocompatibilidad del sistema. Con este objetivo, describimos el recubrimiento electroestático de las nanopartículas con ácido hialurónico mediante la técnica de deposición capa a capa. Se obtuvieron partículas recubiertas y sin recubrir de distintos tamaños (i.e. 300 y 350 nm ó 100 y 130 nm de diámetro, respectivamente). El recubrimiento mejoraba la hemocompatibilidad, estabilidad, vectorización y actividad anti-cancerígena del sistema. Mientras que, la reducción del diámetro de los sistemas en 50 ó 30 nm, también aceleró la internalización de los mismos. Además, el sistema demostró inducir la apoptosis y reducir la migración de las células cancerígenas de mama de forma más efectiva que el fármaco libre mediante mecanismos COX-independientes. Por tanto, esta estrategia podría mejorar la terapia actual contra el cáncer de mama donde las células madre son el principal objetivo. Como conclusión, esta tesis describe la preparación de fármacos poliméricos basados en AINEs, su copolimerización con 1-vinyl-imidazol para la obtención de una familia de copolímeros anfifílicos con microestructura de pseudo-gradiente que se autoensamblen en medio acuoso formando sistemas de liberación controlada nanométricos. Además, en los distintos capítulos se describen tres estrategias diferentes para su utilización en el tratamiento de: enfermedades inflamatorias autoinmunes, la respuesta a cuerpo extraño y el cáncer de mama, respectivamente. Su gran potencial en el campo de la inflamación queda patente en el amplio abanico de aplicaciones y proyectos en los que están actualmente implicadas estas nanopartículas (Anexo I)Programa de Doctorado en Ciencia e Ingeniería de Materiales por la Universidad Carlos III de MadridPresidente: Julia Buján Varela.- Secretario: José Carlos Rodríguez Cabello.- Vocal: Martijn Van Griensve

Anti-Inflammatory Polymeric Nanoparticles Based on Ketoprofen and Dexamethasone

Polymeric nanoparticles that combine dexamethasone and naproxen reduce inflammation and synergistically inhibit Interleukin-12b (Il12b) transcription in macrophages. This effect can be the result of a cyclooxygenase-dependent or a cyclooxygenase-independent mechanism. The aim of this work is to obtain potent anti-inflammatory polymeric nanoparticles by the combination of dexamethasone and ketoprofen, one of the most efficient cyclooxygenase-inhibitors among non-steroidal anti-inflammatory drugs, with appropriate hydrodynamic properties to facilitate accumulation and co-release of drugs in inflamed tissue. Nanoparticles are spherical with hydrodynamic diameter (117 &plusmn; 1 nm), polydispersity (0.139 &plusmn; 0.004), and surface charge (+30 &plusmn; 1 mV), which confer them with high stability and facilitate both macrophage uptake and internalization pathways to favor their retention at the inflamed areas and lysosomal degradation and drug release, respectively. In vitro biological studies concluded that the dexamethasone-loaded ketoprofen-bearing system is non-cytotoxic and efficiently reduces lipopolysaccharide-induced nitric oxide release. The RT-qPCR analysis shows that the ketoprofen nanoparticles were able to reduce to almost basal levels the expression of tested pro-inflammatory markers and increase the gene expression of anti-inflammatory cytokines under inflammatory conditions. However, the synergistic inhibition of Il12b observed in nanoparticles that combine dexamethasone and naproxen was not observed in nanoparticles that combine dexamethasone and ketoprofen, suggesting that the synergistic trans-repression of Il12b observed in the first case was not mediated by cyclooxygenase-dependent pathways