Ultrasound-assisted extraction of bioactive compounds from goji berries: Optimization, bioactivity, and intestinal permeability assessment.

Lycium barbarum L. berries have a remarkable chemical composition and extensive biological activities, being a valuable component of health and nutraceutical practices. Nevertheless, a deep insight on the intestinal permeation of the pro-healthy bioactive compounds is urgently needed to predict the real effects on human body. This study attempted, for the first time, to optimize the Ultrasound-Assisted Extraction (UAE) of goji berries using a Response Surface Methodology approach and establish the intestinal permeation of the principal pro-healthy compounds. The optimal extraction conditions were a solid:liquid ratio of 8.75 % for 56.21 min, using an intensity of 59.05 W/m2. The optimal extract displayed a remarkable antioxidant capacity, with LC/DAD-ESI-MS analysis unveiled a diverse phytochemical profile, encompassing different compounds (e.g. glu-lycibarbarspermidine F, 2-glu-kukoamine, rutin, 3,5-dicaffeoylquinic acid). The intestinal co-culture model demonstrated that glu-lycibarbarspermidine F (isomer 2) (73.70 %), 3,5-dicaffeoylquinic acid (52.66 %), and isorhamnetin-3-O-rutinoside (49.31 %) traversed the intestinal cell layer, exerting beneficial health-promoting effects.

Insights into PCSK9-LDLR Regulation and Trafficking via the Differential Functions of MHC-I Proteins HFE and HLA-C.

PCSK9 is implicated in familial hypercholesterolemia via targeting the cell surface PCSK9-LDLR complex toward lysosomal degradation. The M2 repeat in the PCSK9's C-terminal domain is essential for its extracellular function, potentially through its interaction with an unidentified "protein X". The M2 repeat was recently shown to bind an R-x-E motif in MHC-class-I proteins (implicated in the immune system), like HLA-C, and causing their lysosomal degradation. These findings suggested a new role of PCSK9 in the immune system and that HLA-like proteins could be "protein X" candidates. However, the participation of each member of the MHC-I protein family in this process and their regulation of PCSK9's function have yet to be determined. Herein, we compared the implication of MHC-I-like proteins such as HFE (involved in iron homeostasis) and HLA-C on the extracellular function of PCSK9. Our data revealed that the M2 domain regulates the intracellular sorting of the PCSK9-LDLR complex to lysosomes, and that HFE is a new target of PCSK9 that inhibits its activity on the LDLR, whereas HLA-C enhances its function. This work suggests the potential modulation of PCSK9's functions through interactions of HFE and HLA-C.

Circular economyeast: Saccharomyces cerevisiae as a sustainable source of glucans and its safety for skincare application.

Glucans, a polysaccharide naturally present in the yeast cell wall that can be obtained from side streams generated during the fermentation process, have gained increasing attention for their potential as a skin ingredient. Therefore, this study focused on the extraction method to isolate and purify water-insoluble glucans from two different Saccharomyces cerevisiae strains: an engineered strain obtained from spent yeast in an industrial fermentation process and a wild strain produced through lab-scale fermentation. Two water-insoluble extracts with a high glucose content (> 90 %) were achieved and further subjected to a chemical modification using carboxymethylation to improve their water solubility. All the glucans' extracts, water-insoluble and carboxymethylated, were structurally and chemically characterized, showing almost no differences between both yeast-type strains. To ensure their safety for skin application, a broad safety assessment was undertaken, and no cytotoxic effect, immunomodulatory capacity (IL-6 and IL-8 regulation), genotoxicity, skin sensitization, and impact on the skin microbiota were observed. These findings highlight the potential of glucans derived from spent yeast as a sustainable and safe ingredient for cosmetic and skincare formulations, contributing to the sustainability and circular economy.

Linguistic Disparities in Diabetes Care Quality in California Community Health Centers Before and During the COVID-19 Pandemic.

BACKGROUND: Disparities in diabetes care quality may have increased for patients with limited English language proficiency (LEP) compared to non-LEP patients during the COVID-19 pandemic. Changes in diabetes care quality for adult LEP and non-LEP patients of community health centers (CHCs) were examined from 2019 to 2020. METHODS: Adults with Type 2 diabetes (n = 15 965) of 88 CHC sites in California and with 1+ visit/year in 2019 and 2020 from OCHIN electronic health record data were included. Multivariable regression models estimated the association of LEP status and changes in diabetes care quality from 2019 to 2020, controlling for patient sociodemographic and clinical characteristics. Interaction terms (LEP × 2020) were used to estimate differential over time changes in (1) blood pressure screening, (2) blood pressure control (<140/90 mm Hg), and (3) hemoglobin A1c control (HbA1c <8%) for LEP versus non-LEP patients. RESULTS: LEP and non-LEP patients with diabetes had comparable blood pressure screening and control in 2019 and in 2020. LEP patients were less likely than non-LEP patients to have their HbA1c under control in 2019 (OR = 0.85, 95% CI = 0.77, 0.96, P = .006) and 2020 (OR = 0.83, 95% CI = 0.75, 0.92, P = .001). There were no differential changes in HbA1c control over time for LEP and non-LEP patients. DISCUSSION: Although LEP patients were less likely than non-LEP patients to have their HbA1c under control, CHCs maintained quality of care equally for LEP and non-LEP patients with diabetes during the early pandemic period.

Macrophage Migration Inhibitory Factor (MIF) is a Key Player in Dry Eye Disease.

PURPOSE: To explore the role of the proinflammatory cytokine, macrophage migration inhibitory factor (MIF), in a murine model of dry eye disease (DED). METHODS: The role of MIF on DED was determined using genetically MIF deficient mice and pharmacological inhibition of MIF. DED was induced with 0.5 mg of scopolamine via subcutaneous injection in wild type (WT) and mice lacking MIF (Mif-/-), three times a day for 21 days. DED signs, tear volume, ferning pattern and cytology impression were evaluated. Also, eye tissues were collected to determine transcripts of key inflammatory mediators and histopathological damage. In a second set of experiments, we neutralized MIF with ISO-1, an isozaxiline-derivative MIF tautomerase activity-inhibiting small molecule in WT mice, following an acute DED model for 10 days. ISO-1 was given starting on day 3 after DED induction and signs were evaluated, including a recovery phase in both experimental approaches. RESULTS: When compared to WT, Mif-/- mice showed attenuated signs of DED like preserved mucin pattern and increased tear volume. Also, Mif-/- mice maintained conjunctival epithelial cells and less corneal damage, associated with lower levels of TNFα and IL-1ß. At recovery phase, Mif-/- mice presented improved signs. Interestingly, in cornea and conjunctiva the absence of MIF selectively downregulated the transcription of inflammatory enzymes like inos and nox4 whereas displayed enhanced transcripts of il-4, il-13, tgfß and cox2. Finally, pharmacological inhibition of MIF using ISO-1, replicated the above findings in the mouse model. CONCLUSION: MIF is a central positive mediator of the inflammatory process in experimental DED, thus, targeting MIF could be used as a novel therapy in ocular surface inflammatory pathologies.

Biological Potential and Bioaccessibility of Encapsulated Curcumin into Cetyltrimethylammonium Bromide Modified Cellulose Nanocrystals.

Curcumin is a natural phenolic compound with important biological functions. Despite its demonstrated efficacy in vitro, curcumin biological activities in vivo are dependent on its bioaccessibility and bioavailability, which have been highlighted as a crucial challenge. Cetyltrimethylammonium bromide-modified cellulose nanocrystals (CNC-CTAB) have been shown to be effective in curcumin encapsulation, as they have the potential to enhance biological outcomes. This study evaluated the biological effects of curcumin encapsulated within CNC-CTAB structures, namely its antioxidant, anti-inflammatory and antimicrobial properties, as well as the release profile under digestion conditions and intestinal permeability. Encapsulated curcumin demonstrated antioxidant and anti-inflammatory properties, effectively reducing reactive oxygen species and cytokine production by intestinal cells. The delivery system exhibited antimicrobial properties against Campylobacter jejuni bacteria, further suggesting its potential in mitigating intestinal inflammation. The system showed the ability to protect curcumin from degradation and facilitate its interaction with the intestinal epithelium, highlighting the potential of CNC-CTAB as carrier to enhance curcumin intestinal biological functions.

A trial-based cost-effectiveness analysis of antibiotic prescription strategies for non-complicated respiratory tract infections in children.

BACKGROUND: Antibiotic prescription for respiratory tract infections (RTIs) in children attending primary care centres is almost double that predicted according to bacterial prevalence. Delayed antibiotic prescription (DAP) is designed to deploy a more rational use of antibiotics. While studies have evaluated DAP efficacy and safety for children with RTIs, little research has been conducted on the economic implications. METHODS: Our trial compared cost-effectiveness for DAP, immediate antibiotic prescription (IAP), and no antibiotic prescription (NAP) for children aged 2-14 years with acute uncomplicated RTIs attended to in 39 primary care centres in Spain. The main outcome was the incremental cost-effectiveness ratio (ICER), measured in euros per gained quality-adjusted life days (QALDs). Net monetary benefit (NMB) was also calculated as a tool for decision making. The analysis was performed from a societal perspective for a time horizon of 30 days, and included healthcare direct costs, non-healthcare direct and indirect costs, and the antimicrobial resistance (AMR) cost. RESULTS: DAP was the most cost-effective strategy, even when the cost of AMR was included. QALD values for the three strategies were very similar. IAP compared to DAP was more costly (109.68 vs 100.90 euros) and similarly effective (27.88 vs 27.94 QALDs). DAP compared to NAP was more costly (100.90 vs 97.48 euros) and more effective (27.94 vs. 27.82 QALDs). The ICER for DAP compared to NAP was 28.84 euros per QALD. The deterministic sensitivity analysis indicated that non-healthcare indirect costs had the greatest impact on the ICER. The cost-effectiveness acceptability curve showed that DAP was the preferred option in approximately 81.75% of Monte Carlo iterations, assuming a willingness-to-pay value of 82.2 euros per gained QALD. CONCLUSIONS: When clinicians are in doubt about whether an antibiotic is needed for children with RTIs attending PC centres, those treated with the DAP strategy will have slightly better efficiency outcomes than those treated with IAP because its costs are lower than those of IAP. DAP is also the most cost-effective strategy over a time horizon of 30 days if AMR is considered, despite higher short-term costs than NAP. However, if in the long term the costs of AMR are larger than estimated, NAP could also be an alternative strategy. TRIAL REGISTRATION: This trial has been registered at www. CLINICALTRIALS: gov (identifier NCT01800747; Date: 28/02/2013 (retrospectively registered).

Modification of Acorn Starch Structure and Properties by High Hydrostatic Pressure.

Despite being rich in starch, over half of acorn production is undervalued. High hydrostatic pressure was used to modify the properties of Q. pyrenaica (0.1 and 460 MPa for 20 min) and Q. robur (0.1 and 333 MPa for 17.4 min) acorn starches to obtain high-valued ingredients. Pressure significantly altered the span distribution and heterogeneity of the acorn starch granules depending on the species, but their morphology was unaffected. Pressurization increased the amylose/amylopectin ratio and damaged starch contents, but the effect was more prominent in Q. pyrenaica than in Q. robur. However, the polymorphism, relative crystallinity, gelatinization temperatures, and enthalpies were preserved. The pressure effect on the starch properties depended on the property and species. The solubility, swelling power, and acorn gels' resistance towards deformation for both species decreased after pressurization. For Q. pyrenaica starch, the in vitro digestibility increased, but the pseudoplastic behavior decreased after pressurization. No differences were seen for Q. robur. Regarding the commercial starch, acorn starches had lower gelatinization temperatures and enthalpies, lower in vitro digestibility, lower resistance towards deformation, superior pseudoplastic behavior, and overall higher solubility and swelling power until 80 °C. This encourages the usage of acorn starches as a new food ingredient.

Assessing the In Vitro Digestion of Lactoferrin-Curcumin Nanoparticles Using the Realistic Gastric Model.

Nanosized delivery systems have been the subject of research and discussion in the scientific community due to their unique properties and functionality. However, studies reporting the behaviour of nanodelivery systems under dynamic in vitro digestion conditions are still very scarce. To address this gap, this study aims to assess the dynamic in vitro gastric digestion of lactoferrin/curcumin nanoparticles in the realistic gastric model (RGM). For this purpose, the INFOGEST standard semi-dynamic digestion protocol was used. The nanosystems were characterized in terms of hydrodynamic size, size distribution, polydispersity index (PdI), and zeta potential using dynamic light scattering (DLS), before and during the digestion process. Confocal laser scanning microscopy (CLSM) was also used to examine particle aggregation. In addition, the release of curcumin was evaluated spectroscopically and the intrinsic fluorescence of lactoferrin was measured throughout the digestion process. The protein hydrolysis was also determined by UV-VIS-SWNIR spectroscopy to estimate, in real-time, the presence of free NH2 groups during gastric digestion. It was possible to observe that lactoferrin/curcumin nanoparticles were destabilized during the dynamic digestion process. It was also possible to conclude that low sample volumes can pose a major challenge in the application of dynamic in vitro digestion models.

Iniciativas de autonomía alimentaria como escenarios para la acción comunitaria en salud basada en activos / Food autonomy initiatives as scenarios for community action in health based on assets

Se expone el desarrollo de un proceso territorial de acción comunitaria para la salud basada en activos, que tuvo como objetivo generar estrategias concretas para combatir el hambre y la malnutrición en un barrio popular de la ciudad de Tunja (Colombia) con altas brechas de desigualdad económica y fragmentación social. A partir de la identificación y la dinamización de diversas iniciativas de autonomías alimentarias se generó una red comunitaria que facilitó la utilización colectiva de recursos, saberes y prácticas propias alrededor del proceso agroalimentario. Con ello se promovió la accesibilidad a alimentos saludables y culturalmente legítimos, a la vez que se configuró un espacio vincular de autonomía, organización, participación y cooperación solidaria entre vecinos. Esto demuestra la potencialidad salutogénica de las acciones locales en salud y de abordar la alimentación de manera participativa, hecho que señalamos como una propuesta político-popular y académica para la promoción de la salud colectiva.(AU)

This paper presents the development of a territorial process of community action for health based on assets. Its objective was to generate concrete strategies to combat hunger and malnutrition in a working-class neighbourhood of the Colombian city of Tunja where there are significant gaps in terms of economic inequality and social fragmentation. Through the identification and dynamization of diverse initiatives of food autonomy, a community network was created which facilitated the collective use of their own resources, knowledge, and practices around the agri-food process. This promoted access to healthy and culturally accepted foods and a space where autonomy, organisation, participation, and cooperation among neighbours converged. The above shows the salutogenic potentiality of local actions in health and of approaching food in a participative way, something that we point out as a political-popular and academic proposal for the promotion of collective health.(AU)

[Food autonomy initiatives as scenarios for community action in health based on assets]. / Iniciativas de autonomía alimentaria como escenarios para la acción comunitaria en salud basada en activos.

This paper presents the development of a territorial process of community action for health based on assets. Its objective was to generate concrete strategies to combat hunger and malnutrition in a working-class neighbourhood of the Colombian city of Tunja where there are significant gaps in terms of economic inequality and social fragmentation. Through the identification and dynamization of diverse initiatives of food autonomy, a community network was created which facilitated the collective use of their own resources, knowledge, and practices around the agri-food process. This promoted access to healthy and culturally accepted foods and a space where autonomy, organisation, participation, and cooperation among neighbours converged. The above shows the salutogenic potentiality of local actions in health and of approaching food in a participative way, something that we point out as a political-popular and academic proposal for the promotion of collective health.

Carboxymethyl Cellulose as a Food Emulsifier: Are Its Days Numbered?

Carboxymethyl cellulose use in industry is ubiquitous. Though it is recognized as safe by the EFSA and FDA, newer works have raised concerns related to its safety, as in vivo studies showed evidence of gut dysbiosis associated with CMC's presence. Herein lies the question, is CMC a gut pro-inflammatory compound? As no work addressed this question, we sought to understand whether CMC was pro-inflammatory through the immunomodulation of GI tract epithelial cells. The results showed that while CMC was not cytotoxic up to 25 mg/mL towards Caco-2, HT29-MTX and Hep G2 cells, it had an overall pro-inflammatory behavior. In a Caco-2 monolayer, CMC by itself increased IL-6, IL-8 and TNF-α secretion, with the latter increasing by 1924%, and with these increases being 9.7 times superior to the one obtained for the IL-1ß pro-inflammation control. In co-culture models, an increase in secretion in the apical side, particularly for IL-6 (692% increase), was observed, and when RAW 264.7 was added, data showed a more complex scenario as stimulation of pro-inflammatory (IL-6, MCP-1 and TNF-α) and anti-inflammatory (IL-10 and IFN-ß) cytokines in the basal side was observed. Considering these results, CMC may exert a pro-inflammatory effect in the intestinal lumen, and despite more studies being required, the incorporation of CMC in foodstuffs must be carefully considered in the future to minimize potential GI tract dysbiosis.

Design of Innovative Biocompatible Cellulose Nanostructures for the Delivery and Sustained Release of Curcumin.

Poor aqueous solubility, stability and bioavailability of interesting bioactive compounds is a challenge in the development of bioactive formulations. Cellulose nanostructures are promising and sustainable carriers with unique features that may be used in enabling delivery strategies. In this work, cellulose nanocrystals (CNC) and cellulose nanofibers were investigated as carriers for the delivery of curcumin, a model liposoluble compound. Nanocellulose modification with the surfactant cetyltrimethylammonium bromide (CTAB), tannic acid and decylamine (TADA), and by TEMPO-mediated oxidation were also tested and compared. The carrier materials were characterized in terms of structural properties and surface charge, while the delivery systems were evaluated for their encapsulation and release properties. The release profile was assessed in conditions that mimic the gastric and intestinal fluids, and cytotoxicity studies were performed in intestinal cells to confirm safe application. Modification with CTAB and TADA resulted in high curcumin encapsulation efficiencies of 90 and 99%, respectively. While no curcumin was released from TADA-modified nanocellulose in simulated gastrointestinal conditions, CNC-CTAB allowed for a curcumin-sustained release of ca. 50% over 8 h. Furthermore, the CNC-CTAB delivery system showed no cytotoxic effects on Caco-2 intestinal cells up to 0.125 g/L, meaning that up to this concentration the system is safe to use. Overall, the use of the delivery systems allowed for the reduction in the cytotoxicity associated with higher curcumin concentrations, highlighting the potential of nanocellulose encapsulation systems.

Comparative Study of Green and Traditional Routes for Cellulose Extraction from a Sugarcane By-Product.

Sugarcane bagasse (SCB) is the main residue of the sugarcane industry and a promising renewable and sustainable lignocellulosic material. The cellulose component of SCB, present at 40-50%, can be used to produce value-added products for various applications. Herein, we present a comprehensive and comparative study of green and traditional approaches for cellulose extraction from the by-product SCB. Green methods of extraction (deep eutectic solvents, organosolv, and hydrothermal processing) were compared to traditional methods (acid and alkaline hydrolyses). The impact of the treatments was evaluated by considering the extract yield, chemical profile, and structural properties. In addition, an evaluation of the sustainability aspects of the most promising cellulose extraction methods was performed. Among the proposed methods, autohydrolysis was the most promising approach in cellulose extraction, yielding 63.5% of a solid fraction with ca. 70% cellulose. The solid fraction showed a crystallinity index of 60.4% and typical cellulose functional groups. This approach was demonstrated to be environmentally friendly, as indicated by the green metrics assessed (E(nvironmental)-factor = 0.30 and Process Mass Intensity (PMI) = 20.5). Autohydrolysis was shown to be the most cost-effective and sustainable approach for the extraction of a cellulose-rich extract from SCB, which is extremely relevant for aiming the valorization of the most abundant by-product of the sugarcane industry.

SKI-1/S1P Facilitates SARS-CoV-2 Spike Induced Cell-to-Cell Fusion via Activation of SREBP-2 and Metalloproteases, Whereas PCSK9 Enhances the Degradation of ACE2.

Proprotein convertases activate various envelope glycoproteins and participate in cellular entry of many viruses. We recently showed that the convertase furin is critical for the infectivity of SARS-CoV-2, which requires cleavage of its spike protein (S) at two sites: S1/S2 and S2'. This study investigates the implication of the two cholesterol-regulating convertases SKI-1 and PCSK9 in SARS-CoV-2 entry. The assays used were cell-to-cell fusion in HeLa cells and pseudoparticle entry into Calu-3 cells. SKI-1 increased cell-to-cell fusion by enhancing the activation of SREBP-2, whereas PCSK9 reduced cell-to-cell fusion by promoting the cellular degradation of ACE2. SKI-1 activity led to enhanced S2' formation, which was attributed to increased metalloprotease activity as a response to enhanced cholesterol levels via activated SREBP-2. However, high metalloprotease activity resulted in the shedding of S2' into a new C-terminal fragment (S2″), leading to reduced cell-to-cell fusion. Indeed, S-mutants that increase S2″ formation abolished S2' and cell-to-cell fusion, as well as pseudoparticle entry, indicating that the formation of S2″ prevents SARS-CoV-2 cell-to-cell fusion and entry. We next demonstrated that PCSK9 enhanced the cellular degradation of ACE2, thereby reducing cell-to-cell fusion. However, different from the LDLR, a canonical target of PCSK9, the C-terminal CHRD domain of PCSK9 is dispensable for the PCSK9-induced degradation of ACE2. Molecular modeling suggested the binding of ACE2 to the Pro/Catalytic domains of mature PCSK9. Thus, both cholesterol-regulating convertases SKI-1 and PCSK9 can modulate SARS-CoV-2 entry via two independent mechanisms.

Spent Yeast Waste Streams as a Sustainable Source of Bioactive Peptides for Skin Applications.

Spent yeast waste streams are a byproduct obtained from fermentation process and have been shown to be a rich secondary source of bioactive compounds such as phenolic compounds and peptides. The latter are of particular interest for skin care and cosmetics as they have been shown to be safe and hypoallergenic while simultaneously being able to exert various effects upon the epidermis modulating immune response and targeting skin metabolites, such as collagen production. As the potential of spent yeast's peptides has been mainly explored for food-related applications, this work sought to understand if peptide fractions previously extracted from fermentation engineered spent yeast (Saccharomyces cerevisiae) waste streams possess biological potential for skin-related applications. To that end, cytotoxic effects on HaCat and HDFa cells and whether they were capable of exerting a positive effect upon the production of skin metabolites relevant for skin health, such as collagen, hyaluronic acid, fibronectin and elastin, were evaluated. The results showed that the peptide fractions assayed were not cytotoxic up to the highest concentration tested (500 µg/mL) for both cell lines tested. Furthermore, all peptide fractions showed a capacity to modulate the various target metabolites production with an overall positive effect being observed for the four fractions over the six selected targets (pro-collagen IαI, hyaluronic acid, fibronectin, cytokeratin-14, elastin, and aquaporin-9). Concerning the evaluated fractions, the overall best performance (Gpep > 1 kDa) was of an average promotion of 41.25% over the six metabolites and two cell lines assessed at a concentration of 100 µg/mL. These results showed that the peptide fractions assayed in this work have potential for future applications in skin-related products at relatively low concentrations, thus providing an alternative solution for one of the fermentation industry's waste streams and creating a novel and highly valuable bioactive ingredient with encompassing activity to be applied in future skin care formulations.

Molecular interactions of PCSK9 with an inhibitory nanobody, CAP1 and HLA-C: Functional regulation of LDLR levels.

OBJECTIVE: The liver-derived circulating PCSK9 enhances the degradation of the LDL receptor (LDLR) in endosomes/lysosomes. PCSK9 inhibition or silencing is presently used in clinics worldwide to reduce LDL-cholesterol, resulting in lower incidence of cardiovascular disease and possibly cancer/metastasis. The mechanism by which the PCSK9-LDLR complex is sorted to degradation compartments is not fully understood. We previously suggested that out of the three M1, M2 and M3 subdomains of the C-terminal Cys/His-rich-domain (CHRD) of PCSK9, only M2 is critical for the activity of extracellular of PCSK9 on cell surface LDLR. This likely implicates the binding of M2 to an unknown membrane-associated "protein X" that would escort the complex to endosomes/lysosomes for degradation. We reported that a nanobody P1.40 binds the M1 and M3 domains of the CHRD and inhibits the function of PCSK9. It was also reported that the cytosolic adenylyl cyclase-associated protein 1 (CAP1) could bind M1 and M3 subdomains and enhance the activity of PCSK9. In this study, we determined the 3-dimensional structure of the CHRD-P1.40 complex to understand the intricate interplay between P1.40, CAP1 and PCSK9 and how they regulate LDLR degradation. METHODS: X-ray diffraction of the CHRD-P1.40 complex was analyzed with a 2.2 Å resolution. The affinity and interaction of PCSK9 or CHRD with P1.40 or CAP1 was analyzed by atomic modeling, site-directed mutagenesis, bio-layer interferometry, expression in hepatic cell lines and immunocytochemistry to monitor LDLR degradation. The CHRD-P1.40 interaction was further analyzed by deep mutational scanning and binding assays to validate the role of predicted critical residues. Conformational changes and atomic models were obtained by small angle X-ray scattering (SAXS). RESULTS: We demonstrate that PCSK9 exists in a closed or open conformation and that P1.40 favors the latter by binding key residues in the M1 and M3 subdomains of the CHRD. Our data show that CAP1 is well secreted by hepatic cells and binds extracellular PCSK9 at distinct residues in the M1 and M3 modules and in the acidic prodomain. CAP1 stabilizes the closed conformation of PCSK9 and prevents P1.40 binding. However, CAP1 siRNA only partially inhibited PCSK9 activity on the LDLR. By modeling the previously reported interaction between M2 and an R-X-E motif in HLA-C, we identified Glu567 and Arg549 as critical M2 residues binding HLA-C. Amazingly, these two residues are also required for the PCSK9-induced LDLR degradation. CONCLUSIONS: The present study reveals that CAP1 enhances the function of PCSK9, likely by twisting the protein into a closed configuration that exposes the M2 subdomain needed for targeting the PCSK9-LDLR complex to degradation compartments. We hypothesize that "protein X", which is expected to guide the LDLR-PCSK9-CAP1 complex to these compartments after endocytosis into clathrin-coated vesicles, is HLA-C or a similar MHC-I family member. This conclusion is supported by the PCSK9 natural loss-of-function Q554E and gain-of-function H553R M2 variants, whose consequences are anticipated by our modeling.

Hipoacusia súbita y COVID-19. Revisión sistemática / Sudden Sensorineural Hearing Loss and COVID-19. Systematic Review

Introducción: Se han incrementado los casos de sordera súbita, acúfeno y neuritis vestibular relacionados con la infección por SARS-CoV-2. Objetivo: Realizar una revisión sistemática de los casos publicados de hipoacusia súbita en el contexto de una infección por SARS-CoV-2 y discutir la potencial asociación de la infección como posible agente etiológico. Material y método: Se realizó una búsqueda sistemática en PubMed, EMBASE, Cochrane Library y Scielo, sin restricción de idioma, incluyendo todos los artículos publicados desde marzo del 2020 hasta el 31 de enero de 2022. Resultados: Un total de 16 artículos cumplieron los criterios de inclusión, obteniendo 30 pacientes con sordera súbita y COVID-19. El 53% fueron mujeres y el 47% hombres con una edad media de 46,75 años. 4 pacientes presentaron factores de riesgo cardiovascular, uno asma, otro artritis reumatoide y 10 no presentaron antecedentes. 25 tuvieron una sordera unilateral y en 5 casos fue bilateral. Se encontraron todos los grados de hipoacusia, desde leves a cofosis. El acúfeno fue el síntoma más comúnmente asociado la mitad de los pacientes, acompañado de vértigo en un 10%. 14 pacientes precisaron ingreso en UCI, 9 no recibieron tratamiento para la infección, un paciente recibió hidroxicloroquina, 2 favipavir y enoxaparina y 2 azitromicina y prednisona. El implante coclear fue el tratamiento de la hipoacusia en 2 casos, uno fue tratado con hidroxicloroquina y 23 con corticoides intravenosos y/o orales y/o intratimpánicos. Conclusiones: La hipoacusia súbita es un síntoma recientemente asociado a la infección por SARS-CoV-2 y de prevalencia incierta. Se debe considerar la presencia de hipoacusia como parte de la evaluación clínica de estos pacientes para garantizar un diagnóstico y tratamiento tempranos. (AU)

Effect of Coronavirus 19 on Maintaining Balance in Skilled Athletes.

Preliminary studies have reported that motor control is negatively impacted following an infection of COVID-19. The purpose of this study was to evaluate the effect of COVID-19 on maintaining balance in highly skilled athletes. As part of a larger investigation that was initiated in 2019, twelve professional handball players were recruited to participate in a study that was designed to measure static balance performance. Following the initial pre-test, six participants (body height 184.8 ± 4.7 cm; body weight 85.5 ± 3.3 kg; age 21.3 ± 1.2 years) were infected with COVID-19. The remaining six participants (body height 188.7 ± 2.6 cm; body weight 92.3 ± 3.7 kg; age 26.3 ± 3.3 years) never tested positive for COVID-19 and were presumably not infected with the virus. The experimental design required all the participants to complete an initial balance assessment (pre-test) and a later balance assessment (post-test). To fully analyze our data, we conducted a 2 (condition: COVID, no-COVID) X 2 (test: pre-test, post-test) ANOVA with repeated measures on the second factor. Our analysis revealed that the skilled athletes who contracted COVID-19 had a significant decrease in balance performance from the pre-test that occurred prior to being infected with COVID-19 relative to the post-test that occurred following the COVID-19 infection. Additionally, the skilled athletes who were not infected with COVID-19 did not demonstrate the same deterioration in balance performance in the same period. This study highlights the impact COVID-19 has on static balance performance in a group of highly skilled handball players. Longitudinal studies are needed to fully understand the lasting impacts COVID-19 has on motor behavior.

JUN Regulation of Injury-Induced Enhancers in Schwann Cells.

Schwann cells play a critical role after peripheral nerve injury by clearing myelin debris, forming axon-guiding bands of Büngner, and remyelinating regenerating axons. Schwann cells undergo epigenomic remodeling to differentiate into a repair state that expresses unique genes, some of which are not expressed at other stages of Schwann cell development. We previously identified a set of enhancers that are activated in Schwann cells after nerve injury, and we determined whether these enhancers are preprogrammed into the Schwann cell epigenome as poised enhancers before injury. Poised enhancers share many attributes of active enhancers, such as open chromatin, but are marked by repressive histone H3 lysine 27 (H3K27) trimethylation rather than H3K27 acetylation. We find that most injury-induced enhancers are not marked as poised enhancers before injury indicating that injury-induced enhancers are not preprogrammed in the Schwann cell epigenome. Injury-induced enhancers are enriched with AP-1 binding motifs, and the c-JUN subunit of AP-1 had been shown to be critical to drive the transcriptional response of Schwann cells after injury. Using in vivo chromatin immunoprecipitation sequencing analysis in rat, we find that c-JUN binds to a subset of injury-induced enhancers. To test the role of specific injury-induced enhancers, we focused on c-JUN-binding enhancers upstream of the Sonic hedgehog (Shh) gene, which is only upregulated in repair Schwann cells compared with other stages of Schwann cell development. We used targeted deletions in male/female mice to show that the enhancers are required for robust induction of the Shh gene after injury.SIGNIFICANCE STATEMENT The proregenerative actions of Schwann cells after nerve injury depends on profound reprogramming of the epigenome. The repair state is directed by injury-induced transcription factors, like JUN, which is uniquely required after nerve injury. In this study, we test whether the injury program is preprogrammed into the epigenome as poised enhancers and define which enhancers bind JUN. Finally, we test the roles of these enhancers by performing clustered regularly interspaced short palindromic repeat (CRISPR)-mediated deletion of JUN-bound injury enhancers in the Sonic hedgehog gene. Although many long-range enhancers drive expression of Sonic hedgehog at different developmental stages of specific tissues, these studies identify an entirely new set of enhancers that are required for Sonic hedgehog induction in Schwann cells after injury.