Age-related metabolic changes limit efficacy of deoxynucleoside-based therapy in thymidine kinase 2-deficient mice

Thymidine kinase 2 (TK2) catalyses the phosphorylation of deoxythymidine (dThd) and deoxycytidine (dCtd) within mitochondria. TK2 deficiency leads to mtDNA depletion or accumulation of multiple deletions. In patients, TK2 mutations typically manifest as a rapidly progressive myopathy with infantile onset, leading to respiratory insufficiency and encephalopathy in the most severe clinical presentations. TK2-deficient mice develop the most severe form of the disease and die at average postnatal day 16. dThd+dCtd administration delayed disease progression and expanded lifespan of a knockin murine model of the disease. We daily administered TK2 knockout mice (Tk2 KO) from postnatal day 4 with equimolar doses of dThd+dCtd, dTMP+dCMP, dThd alone or dCtd alone. We monitored body weight and survival and studied different variables at 12 or 29 days of age. We determined metabolite levels in plasma and target tissues, mtDNA copy number in tissues, and the expression and activities of enzymes with a relevant role in mitochondrial dNTP anabolism or catabolism. dThd+dCtd treatment extended average lifespan of Tk2 KO mice from 16 to 34 days, attenuated growth retardation, and rescued mtDNA depletion in skeletal muscle and other target tissues of 12-day-old mice, except in brain. However, the treatment was ineffective in 29-day-old mice that still died prematurely. Bioavailability of dThd and dCtd markedly decreased during mouse development. Activity of enzymes catabolizing dThd and dCtd increased with age in small intestine. Conversely, the activity of the anabolic enzymes decreased in target tissues during mouse development. We also found that administration of dThd alone had the same impact on survival to that of dThd+dCtd, whereas dCtd alone had no influence on lifespan. dThd+dCtd treatment recruits alternative cytosolic salvage pathways for dNTP synthesis, suggesting that this therapy would be of benefit for any Tk2 mutation. dThd accounts for the therapeutic effect of the combined treatment in mice. During the first weeks after birth, mice experience marked tissue-specific metabolic regulations and ontogenetic changes in dNTP metabolism-related enzymes that limit therapeutic efficacy to early developmental stages. This study was funded by grants from the Spanish Ministry of Industry, Economy and Competitiveness, the Spanish Instituto de Salud Carlos III, the Fundación Inocente, Inocente, AFM Téléthon and the Generalitat de Catalunya. The disclosed funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Assessment of in vivo versus in vitro biofilm formation of clinical methicillin-resistant Staphylococcus aureus isolates from endotracheal tubes

Our aim was to demonstrate that biofilm formation in a clinical strain of methicillin-resistant Staphylococcus aureus (MRSA) can be enhanced by environment exposure in an endotracheal tube (ETT) and to determine how it is affected by systemic treatment and atmospheric conditions. Second, we aimed to assess biofilm production dynamics after extubation. We prospectively analyzed 70 ETT samples obtained from pigs randomized to be untreated (controls, n = 20), or treated with vancomycin (n = 32) or linezolid (n = 18). A clinical MRSA strain (MRSA-in) was inoculated in pigs to create a pneumonia model, before treating with antibiotics. Tracheally intubated pigs with MRSA severe pneumonia, were mechanically ventilated for 69 ± 16 hours. All MRSA isolates retrieved from ETTs (ETT-MRSA) were tested for their in vitro biofilm production by microtiter plate assay. In vitro biofilm production of MRSA isolates was sequentially studied over the next 8 days post-extubation to assess biofilm capability dynamics over time. All experiments were performed under ambient air (O2) or ambient air supplemented with 5% CO2. We collected 52 ETT-MRSA isolates (placebo N = 19, linezolid N = 11, and vancomycin N = 22) that were clonally identical to the MRSA-in. Among the ETT-MRSA isolates, biofilm production more than doubled after extubation in 40% and 50% under 5% CO2 and O2, respectively. Systemic antibiotic treatment during intubation did not affect this outcome. Under both atmospheric conditions, biofilm production for MRSA-in was at least doubled for 9 ETT-MRSA isolates, and assessment of these showed that biofilm production decreased progressively over a 4-day period after extubation. In conclusion, a weak biofilm producer MRSA strain significantly enhances its biofilm production within an ETT, but it is influenced by the ETT environment rather than by the systemic treatment used during intubation or by the atmospheric conditions used for bacterial growth

Protein Supplementation with Short Peptides Prevents Early Muscle Mass Loss after Roux-en-Y-Gastric Bypass

Bariatric surgery; Nutrition supplementation; ProteinCirugía bariátrica; Suplementación nutricional; ProteínaCirurgia bariàtrica; Suplementació nutricional; ProteïnaIntroduction: A significant reduction in fat-free mass (FFM) following bariatric surgery (BS) has been reported, and adequate protein intake is recommended for FFM preservation. Current guidelines of nutritional management after BS recommend complex protein (CP) compounds. However, Roux-en-Y-gastric bypass (RYGB) has a negative impact on CP digestion, leading to protein malabsorption. At present, there is no data regarding the impact of early supplementation with short peptide-based (SPB) or hydroxy methylbutyrate (HMB)-enriched formulas on the evolution of the FFM after the BS. Aim: The aim of this study is to evaluate the effect of nutritional products based on CP, HBM-enriched, or SPB formulas on the FFM of patients that undergo RYGB. Material and methods: This is a prospective interventional study, including three groups of patients (according to the type of protein product) as candidates for BS, recruited between December 2021 and April 2022, matched by age, gender, and BMI. All patients underwent evaluations at baseline and one month post-BS, including: medical history, physical and anthropometric evaluation, bioimpedance, and biochemical analysis. Results: A total of 60 patients were recruited: 63% women, mean age 43.13 ± 9.4 years, and BMI 43.57 ± 4.1 kg/m2. The % of FFM loss from total weight loss (TWL) was significantly lower in the SPB group than CP and HMB groups despite the major %TWL in this group (40.60 ± 17.27 in CP, 34.57 ± 13.15 in HMB, and 19.14 ± 9.38 in SPB, p < 0.001). TWL% was 9.98 ± 1.82 vs. 9.83 ± 2.71 vs. 13.56 ± 4.30, p < 0.001, respectively. Conclusion: In our study, the SPB supplementation prevented almost 50% FFM lost from the TWL than the CP- or HMB-enriched compounds at one month post-BS. These results are significant in the setting of muscle mass preservation after the BS, and have the potential to change the current guidelines for the management of nutritional supplementation after BS.This study was supported by grants from the Instituto de Salud Carlos III (Fondo de Investigacion Sanitaria, PI20/01086). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Sugammadex, a neuromuscular blockade reversal agent, causes neuronal apoptosis in primary cultures.

Sugammadex, a γ-cyclodextrin that encapsulates selectively steroidal neuromuscular blocking agents, such as rocuronium or vecuronium, has changed the face of clinical neuromuscular pharmacology. Sugammadex allows a rapid reversal of muscle paralysis. Sugammadex appears to be safe and well tolerated. Its blood-brain barrier penetration is poor (< 3% in rats), and thus no relevant central nervous toxicity is expected. However the blood brain barrier permeability can be altered under different conditions (i.e. neurodegenerative diseases, trauma, ischemia, infections, or immature nervous system). Using MTT, confocal microscopy, caspase-3 activity, cholesterol quantification and Western-blot we determine toxicity of Sugammadex in neurons in primary culture. Here we show that clinically relevant sugammadex concentrations cause apoptotic/necrosis neuron death in primary cultures. Studies on the underlying mechanism revealed that sugammadex-induced activation of mitochondria- dependent apoptosis associates with depletion of neuronal cholesterol levels. Furthermore SUG increase CytC, AIF, Smac/Diablo and CASP-3 protein expression in cells in culture. Potential association of SUG-induced alteration in cholesterol homeostasis with oxidative stress and apoptosis activation occurs. Furthermore, resistance/sensitivity to oxidative stress differs between neuronal cell types

Rotated domains in selective area epitaxy grown ZnP : formation mechanism and functionality

Zinc phosphide (ZnP) is an ideal absorber candidate for solar cells thanks to its direct bandgap, earth-abundance, and optoelectronic characteristics, albeit it has been insufficiently investigated due to limitations in the fabrication of high-quality material. It is possible to overcome these factors by obtaining the material as nanostructures, e.g. via the selective area epitaxy approach, enabling additional strain relaxation mechanisms and minimizing the interface area. We demonstrate that ZnP nanowires grow mostly defect-free when growth is oriented along the [100] and [110] of the crystal, which is obtained in nanoscale openings along the [110] and [010] on InP(100). We detect the presence of two stable rotated crystal domains that coexist in the structure. They are due to a change in the growth facet, which originates either from the island formation and merging in the initial stages of growth or lateral overgrowth. These domains have been visualized through 3D atomic models and confirmed with image simulations of the atomic scale electron micrographs. Density functional theory simulations describe the rotated domains' formation mechanism and demonstrate their lattice-matched epitaxial relation. In addition, the energies of the shallow states predicted closely agree with transition energies observed by experimental studies and offer a potential origin for these defect transitions. Our study represents an important step forward in the understanding of ZnP and thus for the realisation of solar cells to respond to the present call for sustainable photovoltaic technology

Impacte ambiental dels iots en els ports de Sitges : anàlisi comparativa

Els ports esportius a nivell català han augmentat considerablement a causa de la creixent afició a la navegació i els esports nàutics. L'increment de la demanda d'amarratges al port i la conseqüent concentració d'iots, ha implicat la necessitat d'un estudi dels seus impactes potencials. El Port Garraf té un impacte més reduït degut a que prioritzen les embarcacions de vela, en canvi pel que fa al Port d'Aiguadolç i Port Ginesta la proporció d'embarcacions a motor és superior, per tant tindran un consum més elevat de combustible. Pel que fa a residus especials, concretament olis, els tres ports tenen un impacte sever per la difícil recuperació del medi en cas d'accident, encara que hi ha una bona gestió en els ports. Per acabar el Port d'Aiguadolç és el que té un consum d'aigua superior.Los puertos deportivos a nivel catalán han aumentado considerablemente a causa de la creciente afición de la navegación y los deportes náuticos. El incremento de la demanda de amarres en el puerto y la consecuente concentración de yates, ha implicado la necesidad de un estudio de los posibles impactos potenciales. El Port Garraf tiene un impacto más reducido debido a que se priorizan las embarcaciones de vela, en cambio, en el Port d'Aiguadolç y el Port Ginesta, la proporción de embarcaciones a motor es superior, por lo tanto, tendrán un consumo más elevado de combustible. Por lo que se refiere a los residuos especiales, concretamente aceites, los tres puertos tienen un impacto severo por la difícil recuperación del medio en caso de accidente, aunque haya una buena gestión en los puertos. Para acabar, el Port d'Aiguadolç es el que tiene un consumo de agua superior.The marinas has increased considerably in Catalonia because of the growing hobby of sailing and water sports. The increase in the demand of moorings in the marinas and the consequent concentration of yachts, has involved the need of a study of the potential environmental impacts. The Port of Garraf has a smaller environmental impact because it prioritises sailboats; however, the proportion of motor boats is greater in the Port of Ginesta and in the Port of Aiguadolç. That is why they both have a higher consumption of fuel. Regarding the hazardous waste, particularly oils, the three ports have a severe impact because of the difficult recovery of the environment in case of accident, although there is a good management in the ports. To conclude, the Port of Aiguadolç is the most consumptive of water. According to the comparison that has been made of the different flows, it can be concluded that the Port of Ginesta is the most sustainable. Thanks to the good management of the port and its user awareness. Although there is more surface and more boats, this port is the one which has the lower potential of impacts

Diagnostic Value of Endotracheal Aspirates Sonication on Ventilator-Associated Pneumonia Microbiologic Diagnosis

Microorganisms are able to form biofilms within respiratory secretions. Methods to disaggregate such biofilms before utilizing standard, rapid, or high throughput diagnostic technologies may aid in pathogen detection during ventilator associated pneumonia (VAP) diagnosis. Our aim was to determine if sonication of endotracheal aspirates (ETA) would increase the sensitivity of qualitative, semi-quantitative, and quantitative bacterial cultures in an animal model of pneumonia caused by Pseudomonasaeruginosa or by methicillin resistant Staphylococcusaureus (MRSA). MATERIAL AND METHODS: P.aeruginosa or MRSA was instilled into the lungs or the oropharynx of pigs in order to induce severe VAP. Time point assessments for qualitative and quantitative bacterial cultures of ETA and bronchoalveolar lavage (BAL) samples were performed at 24, 48, and 72 h after bacterial instillation. In addition, at 72 h (autopsy), lung tissue was harvested to perform quantitative bacterial cultures. Each ETA sample was microbiologically processed with and without applying sonication for 5 min at 40 KHz before bacterial cultures. Sensitivity and specificity were determined using BAL as a gold-standard. Correlation with BAL and lung bacterial burden was also determined before and after sonication. Assessment of biofilm clusters and planktonic bacteria was performed through both optical microscopy utilizing Gram staining and Confocal Laser Scanning Microscopy utilizing the LIVE/DEAD®BacLight kit. RESULTS: 33 pigs were included, 27 and 6 from P.aeruginosa and MRSA pneumonia models, respectively. Overall, we obtained 85 ETA, 69 (81.2%) from P.aeruginosa and 16 (18.8%) from MRSA challenged pigs. Qualitative cultures did not significantly change after sonication, whereas quantitative ETA cultures did significantly increase bacterial counting. Indeed, sonication consistently increased bacterial burden in ETAs at 24, 48, and 72 h after bacterial challenge. Sonication also improved sensitivity of ETA quantitative cultures and maintained specificity at levels previously reported and accepted for VAP diagnosis. CONCLUSION: The use of sonication in ETA respiratory samples needs to be clinically validated since sonication could potentially improve pathogen detection before standard, rapid, or high throughput diagnostic methods used in routine microbial diagnostics