Determination of Cardiac Output in a Porcine Model for Ex Vivo Pulmonary Perfusion.

Due to their physiological similarities to humans, pigs are used as experimental models for ex vivo lung perfusion (EVLP). EVLP is a technique that perfuses lungs that are not suitable for transplantation via an extracorporeal circulation pump to improve their function and increase their viability. Existing EVLP protocols are differentiated by the type of perfusion solution and perfusion flow, which varies from 40%-100% of the estimated cardiac output (CO) according to the body surface area (BSA). Devices for measuring CO use simple physical principles and other mathematical models. Thermodilution in animal models continues to be the reference standard for estimating CO because of its simplicity and ease of reproduction. Therefore, the objective of this study was to reproduce the measurement of CO by thermodilution in pigs and compare its precision and accuracy with those obtained by the BSA, weight, and Fick's method, to establish perfusion flow during EVLP. In 23 pigs, a thermodilution catheter was placed in the right jugular vein, and the carotid artery on the same side was cannulated. Blood samples were obtained for gasometry, and CO was estimated by thermodilution, adjusted body surface area, Fick's principle, and per body weight. The CO obtained by the BSA was greater (p = 0.0001, ANOVA, Tukey) than that obtained by the other methods. We conclude that although the methods used in this study to estimate CO are reliable, there are significant differences between them; therefore, each method must be evaluated by the investigator to determine which meets the needs of the protocol.

Pathology results of endoscopic ultrasound-guided tissue acquisition in retroperitoneal masses: a multicenter study.

Background: Malignant etiologies are found in 70-80% of symptomatic retroperitoneal masses. Histology is required for diagnosis and treatment. Information about endoscopic ultrasound (EUS)-guided tissue acquisition (EUS-GTA) is scant for retroperitoneal masses. This study aimed to assess the pathology results of EUS-GTA for diagnosing retroperitoneal masses. Methods: This retrospective, multicenter study involved patients from 5 care centers. All patients with retroperitoneal masses who underwent EUS evaluation were enrolled. We recorded demographic and clinical characteristics, location and size of the mass, type of needle (FNA/FNB), and complications related to the procedure. Results: A total of 43 patients were included. The median age was 50.5 (range: 23-83) years, and 22 (51.2%) were female. The initial symptom was abdominal pain in 23 (52.3%) cases and weight loss in 11 (25%). Initial imaging was by computed tomography in 33 (75%) patients. Diagnosis with EUS-GTA was reached in 67.5% (29/43) cases. The most frequent histological diagnosis was carcinoma, in 25.5% (11/43). A malignant etiology was found in 31 (72%): 20 were primary tumors from the retroperitoneum, and 11 were metastases. In patients with metastasis, surgery was avoided and medical treatment was indicated. No adverse events were reported. Conclusion: EUS and EUS-GTA can frequently provide accurate tissue diagnosis and significantly impact the subsequent management.

Control de síntomas al final de la vida con Dexmedetomidina: Estudio observacional de centro único / End-of-life symptom control with dexmedetomidine: A single-center observational study

Introducción: El manejo de los síntomas refractarios con dexmetomidina (DXM) es una alternativa en pacientes oncológicos en la atención al final de la vida (AFV) lo que puede convertirse en un gran desafío, ante la necesidad de iniciar sedación paliativa. El objetivo del presente estudio es determinar el efecto de la DXM comparado con el midazolam (MDZ) en el control de síntomas refractarios en pacientes con cáncer avanzado AFV, como parte de una estrategia asistencial de sedación paliativa. Metodología: Estudio observacional, longitudinal, se revisaron los informes médicos en las historias clínicas de pacientes hospitalizados con diagnóstico de cáncer avanzado terminal y síntomas refractarios (dolor, disnea y delirio), que recibieron sedación paliativa con DXM o MDZ. Se utilizaron los registros de Escala de Evaluación de Síntomas de Edmonton - revisada (ESAS-r) para intensidad de síntomas y la Escala de Sedación/Agitación de Richmond (RASS) para respuesta a la sedación. Resultados: Un total de 35 pacientes recibieron DXM y 18 MDZ. El dolor (32 %) y delirio (30 %) fueron los síntomas refractarios más identificados. Dolor intenso en 89.3 % antes del inicio de DXM. A las 72 horas, el grupo DXM presentó disminución de la intensidad del delirio a leve y moderado, mientras que la mayoría de los pacientes del grupo MDZ fallecieron antes de las 72 horas. Conclusiones: El uso clínico de la DXM permite un enfoque multimodal, ampliando su utilidad en la atención al final de la vida para el manejo del dolor y delirio, ya que permite un estado de respuesta para la interacción con los miembros de la familia, función relevante en cuidados paliativos.

Introduction: Managing refractory symptoms with dexmedetomidine (DXM) is an alternative for cancer patients in end-of-life care (EOLC), which can become a great challenge, given the need to start palliative sedation. This study aims to determine the effect of DXM compared to midazolam (MDZ) in controlling refractory symptoms in patients with advanced EOLC cancer as part of a palliative sedation care strategy. Methodology: In this observational, longitudinal study, the medical reports in the medical records of hospitalized patients diagnosed with terminal advanced cancer and refractory symptoms (pain, dyspnea, and delirium) who received palliative sedation with DXM or MDZ were reviewed. The Edmonton Symptom Assessment Scale-revised (ESAS-r) scores for symptom intensity and the Richmond Sedation/Agitation Scale (RASS) for response to sedation were used. Results: A total of 35 patients received DXM, and 18 received MDZ. Pain (32%) and delirium (30%) were the most identified refractory symptoms. Intense pain in 89.3% before DXM started; at 72 hours in 64.3%, the pain was reported as mild. Delirium was severe in 77.7%, DXM was used in 59.3%, and MDZ was used in 40.7%. At 72 hours, the DXM group presented a decrease in delirium intensity to mild and moderate, while most of the patients in the MDZ group died before 72 hours. Conclusions: The clinical use of DXM allows a multimodal approach, expanding its usefulness in end-of-life care for managing pain and delirium since it provides a state of response for interaction with family members, a function relevant in palliative care.

Triple primario oncológico y su impacto en el dolor. Reporte de caso / Triple primary malignancy and its impact on pain. A case report

Introducción: Los pacientes con cáncer presentan dolor en un 40% y el 38% de ellos lo califica como moderado a intenso. Casi el 85% de los pacientes con dolor por cáncer pueden lograr un buen control con medicamentos orales convencionales. Caso clínico: masculino de 57 años con triple neoplasia maligna primaria metacrónica asociada a tres tipos de dolor con incremento progresivo de la intensidad y requerimiento de varias líneas de tratamiento analgésico opioide. Conclusión: Las neoplasias malignas primarias triples son infrecuentes y están asociadas a dolor complejo, siendo los opioides la opción terapéutica más adecuada.

Introduction: Pain is an unpleasant sensory and emotional experience associated with real or potential tissue damage, which treatment is more difficult in patients with multiple and advanced cancer, influencing their functionality, mood, sleep, and quality of life. Clinical case: 57-year-old male with metachronous triple primary malignancy associated with three types of pain with a progressive increase in intensity and requiring several lines of opioid analgesic treatment. Conclusion: Triple primary malignancies are uncommon and primarily associated with complex pain; opioids are the most appropriate therapeutic option.

Interplay of Oxidants and Antioxidants in Mammalian Embryo Culture System.

One principal purpose of assisted reproductive technology (ART) is to produce viable and good quality embryos. However, a variety of environmental factors may induce epigenetic changes in the embryo. Moreover, laboratory conditions including the culture media may also affect embryo development. Therefore, media change is an important factor in maintaining proper oxidant/antioxidant balance during embryo culture. Alterations in the oxidant/antioxidant balance are related to various cellular responses such as an increase in the level of reactive oxygen species (ROS) and consequent lipid peroxidation (LPO), DNA damage, and apoptosis. The current study focuses on the role of external factors on embryo culture and the ability of antioxidants to enhance in vitro fertilization (IVF) outcomes. Indeed, an optimization of media culture by the addition of enzymatic and nonenzymatic antioxidants in animal models and human embryos in ART has been updated in this study, with an emphasis on comparing the available results and their possible reasons.

Fluorescence-Linked Aptamer Assay for SARS-CoV-2 Spike-Protein: A Step-by-Step Performance Analysis in Clinical Samples.

The COVID-19 pandemic has been a main concern over the last two years and has become one of the most important crises in the history of human health. Today, there is still a need for affordable and reliable diagnostic tests for massive disease monitoring. Previously, a set of highly specific DNA-aptamers (C7/C9) binding to the SARS-CoV-2 Spike (S) protein were isolated but its performance in clinical samples remained to be tested. Here, 242 samples were collected through three different methods and subjected to florescence-linked aptamer assays (FLAA) based on C7/C9 aptamers through two readout protocols. Then, a step-by-step statistical approach which included agreement tests, proportion comparisons and binomial and multinomial logistic regressions was used to predict optimal conditions for the novel C7/C9 FLAA test. RTqPCR threshold cycles, symptoms onset and processing time were influential factors on FLAA test results. Naturally occurring mutations on S were also detected and analyzed. Aminoacidic substitutions D614G and T732A appeared relevant for aptamer recognition although further studies are necessary. The methodology presented here is the first step to determine the performance and diagnosis across a range of clinical contexts and it might serve as a base for a complete analysis applicable to other designs of new diagnostic tests.

Cuidados Paliativos una especialidad en Ecuador / Palliative Care a specialty in Ecuador

Perspectiva de los cuidados paliativos Los cuidados paliativos (CP) o medicina paliativa, especialidad nueva y creciente con un papel muy definido e importante; ha cosechado algunos éxitos, pero tiene que responder satisfactoriamente a nuevos retos para sobrevivir, prosperar y lograr convertirse en una aliada de la medicina curativa y poder interactuar al mismo tiempo para mejorar la atención a los pacientes y familia que se enfrentan a enfermedades complejas como el cáncer. Actualmente es mucho lo que se sabe sobre CP, pero este conocimiento no está beneficiando a las personas que lo necesitan debido al escás formación académica. Para que los CP se incorporen a un sistema de salud es necesario establecer cuatro componentes fundamentales (Figura 1). 1. Políticas. - Establecer políticas que integren los CP en el plan de salud nacional. 2. Disponibilidad de fármacos. - Mejorar la disponibilidad y acceso a los opioides que permita aumentar el cupo de opioides de la Junta Internacional de Fiscalización de Estupefacientes (JIFE) del país. 3. Educación. Convencer al público, comunidad, profesionales sanitarios, y a los políticos, incluir los programas de CP en pregrado y posgrado. 4. Implementación. Crear nuevos puestos de CP a dedicación completa, y dotar de la infraestructura para centros de referencia de excelencia, incluyendo cuidado domicilia-rio que se vayan replicando. Todo esto permitirá hacer una estimación de la necesidad para el adecuado desarrollo de los CP.

Palliative care (PC) or palliative medicine is a new and growing specialty with a very defined and essential role; It has had some successes, but it has to successfully respond to new challenges to survive, prosper and become an ally of curative medicine and to be able to interact at the same time to improve care for patients and families facing complex diseases such as cancer. Currently, much is known about PC, but this knowledge is not benefiting the people who need it due to little or no academic training. For PCs to be incorporated into a health system, it is necessary to establish four fundamental components: 1. Policies. - Establish policies that integrate PC into the national health plan. 2. Drug availability. - Improve the availability and access to opioids to increase the opioid quota of the country's International Narcotics Control Board (INCB). 3. Education. Convince the public, community, health professionals, and politicians to include PC programs in undergraduate and postgraduate courses. 4. Implementation. Create new full-time PC positions, and provide the infrastructure for reference centers of excellence, including home care that will be replicated. All of this will allow estimating the need for the proper development of PC. This editorial aims to publicize PC development in Ecuador from informality to postgraduate medical specialty.

Clinical, Histological, and Profibrotic Extracellular Matrix Protein Changes in a Model of Tracheal Stenosis Induced by Cervical Tracheal Autotransplantation.

BACKGROUND: Tracheal stenosis (TS) is a complication of prolonged intubation, tracheotomy, and tracheal surgery that compromises the vascular supply. Animal models are essential for studying its pathophysiology and the effect of interventions. OBJECTIVE: To establish a TS model in rats secondary to tracheal autotransplantation with a graft submerged in bleomycin (Atx-Bleo). Additionally, to evaluate the clinical and histological changes, as well as the expression of newly formed collagen (NFC), isoforms of transforming growth factor beta (TGFß), fibronectin (FN), elastin (ELN), integrin ß1 (ITGß1), and matrix metalloproteinase 1 (MMP1) in TS. METHODS: Twenty Wistar rats were divided into three groups: group I (n = 20) control; group II (n = 10) end-to-end anastomosis of the trachea (tracheoplasty); and group III (n = 10) Atx-Bleo. The animals were evaluated clinically, tomographically, macroscopically, morphometrically, and microscopically. NFC deposition, and the expression of profibrotic and antifibrotic proteins were evaluated in tracheal scars. RESULTS: All animals survived the surgical procedure and the study period. Compared with the other study groups, the Atx-Bleo group developed TS and fibrosis, exhibited higher expression of NFC, TGFß1, TGFß2, FN, ELN, and ITGß1, and mild expression of TGFß3 and MMP1 (p < 0.005; analysis of variance, Dunnett and Tukey tests). CONCLUSION: Atx-Bleo in TS model rats produces tomographic and histological changes, and induces the upregulation of profibrotic proteins (TGFß1, TGFß2, collagen, FN, ELN, ITGß1) and downregulation of antifibrotic proteins (TGFß3, MMP1). Therefore, this model may be used to test new pharmacological treatments for reversing or preventing TS, and conduct basic studies regarding its pathophysiology.

Intensity distribution segmentation in ultrafast Doppler combined with scanning laser confocal microscopy for assessing vascular changes associated with ageing in murine hippocampi.

The hippocampus plays an important role in learning and memory, requiring high-neuronal oxygenation. Understanding the relationship between blood flow and vascular structure-and how it changes with ageing-is physiologically and anatomically relevant. Ultrafast Doppler ([Formula: see text]Doppler) and scanning laser confocal microscopy (SLCM) are powerful imaging modalities that can measure in vivo cerebral blood volume (CBV) and post mortem vascular structure, respectively. Here, we apply both imaging modalities to a cross-sectional and longitudinal study of hippocampi vasculature in wild-type mice brains. We introduce a segmentation of CBV distribution obtained from [Formula: see text]Doppler and show that this mice-independent and mesoscopic measurement is correlated with vessel volume fraction (VVF) distribution obtained from SLCM-e.g., high CBV relates to specific vessel locations with large VVF. Moreover, we find significant changes in CBV distribution and vasculature due to ageing (5 vs. 21 month-old mice), highlighting the sensitivity of our approach. Overall, we are able to associate CBV with vascular structure-and track its longitudinal changes-at the artery-vein, venules, arteriole, and capillary levels. We believe that this combined approach can be a powerful tool for studying other acute (e.g., brain injuries), progressive (e.g., neurodegeneration) or induced pathological changes.

Endothelin-Converting Enzyme 1 and Vascular Endothelial Growth Factor as Potential Biomarkers during Ex Vivo Lung Perfusion with Prolonged Hypothermic Lung-Sparing.

Lung transplantation requires optimization of donor's organ use through ex vivo lung perfusion (EVLP) to avoid primary graft dysfunction. Biomarkers can aid in organ selection by providing early evidence of suboptimal lungs during EVLP and thus avoid high-risk transplantations. However, predictive biomarkers of pulmonary graft function such as endothelin-converting enzyme (ECE-1) and vascular endothelial growth factor (VEGF) have not been described under EVLP with standard prolonged hypothermic preservation, which are relevant in situations where lung procurement is difficult or far from the transplantation site. Therefore, this study is aimed at quantifying ECE-1 and VEGF, as well as determining their association with hemodynamic, gasometric, and mechanical ventilatory parameters in a swine model of EVLP with standard prolonged hypothermic preservation. Using a protocol with either immediate (I-) or delayed (D-) initiation of EVLP, ECE-1 levels over time were found to remain constant in both study groups (p > 0.05 RM-ANOVA), while the VEGF protein was higher after prolonged preservation, but it decreased throughout EVLP (p > 0.05 RM-ANOVA). Likewise, hemodynamic, gasometric, mechanical ventilatory, and histological parameters had a tendency to better results after 12 hours of hypothermic preservation in the delayed infusion group.

National survey regarding the timing of endoscopic procedures during the COVID-19 pandemic.

BACKGROUND: During the COVID-19 pandemic, several questions have arisen about which endoscopic procedures (EPs) must be performed and which ones can be postponed. The aim of this study was to conduct a nationwide survey regarding the appropriate timing of EPs during the COVID-19 pandemic. METHODS: This prospective study was performed through a nationwide electronic survey. The survey consisted of 15 questions divided into three sections. The first evaluated the agreement for EPs classified as "time sensitive" and "not time sensitive". Two other sections assessed "high-priority" and "low-priority" scenarios. Agreement was considered when > 75% of respondents answered a question in the same direction. RESULTS: The response rate was 27.2% (214/784). Among the respondents, agreement for the need to perform EP in < 72 h was only reached for variceal bleeding (93.4%). Dysphagia with alarm symptoms was the scenario in which the highest percentage of physicians (95.9%) agreed that an EP needed to be performed within a month. Less than 30% of endoscopists would perform an EP within the first 72 h for patients with mild cholangitis, non-variceal upper gastrointestinal bleeding without hemodynamic instability, or severe anaemia without overt bleeding. In time-sensitive clinical scenarios suggestive of benign disease, none of the scenarios reached agreement in any sense. Among the time-sensitive clinical scenarios suggestive of malignancy, > 90% of the surveyed respondents considered that EP could not be postponed for > 8 weeks. CONCLUSIONS: There was no consensus among endoscopists about the timing of EPs in patients with pathologies considered time sensitive or in those with high-priority pathologies. Agreement was only reached in five (17%) of the evaluated clinical scenarios.

The impact of the COVID-19 pandemic on depression and stress levels in pregnant women: a national survey during the COVID-19 pandemic in Mexico.

BACKGROUND: COVID-19 outbreak has been associated with a wide variety of psychiatric manifestations such as panic, anxiety, and depression. We aim to assess the impact of the COVID - 19 pandemic on the levels of stress and depression of pregnant women in Mexico. METHODS: A cross-sectional web survey was carried out in pregnant women in 10 states of the Mexican Republic during the COVID-19 pandemic among public and private hospitals. The perception of stress was assessed using the Perceived Stress Scale, while depressive symptoms were evaluated using the Edinburgh Postnatal Depression Scale. RESULTS: A total of 549 surveys were applied, of which 96.1% (n = 503) were included in the data analysis. The mean participant's age was 28.1 years old. The mean perceived stress scale score was 24. 33.2% (n = 167) of participants had a score equal to 27 points or more and were considered highly stressed. The mean depression score was 9. A total of 17.5% (n = 88) participants had more than 14 points on the Edinburgh's depression scale, and were considered depressed. Stress levels were higher at later gestational ages (p = .008). CONCLUSIONS: COVID-19 pandemic has caused mental health issues in pregnant women reflected by high perceived stress levels and depression.

Expression of Claudin-4 in Lung Ischemia-Reperfusion Injury in Experimental Lung Transplantation.

OBJECTIVE: To assess the presence of CLDN4 in bronchoalveolar lavage fluid (BALF) and pulmonary tissue as an early indicator of LIRI and its relationship with changes in pulmonary physiology, edema formation and histology in an experimental porcine model of LTx with CIT of 50 min or 6 h. METHODS: In 12 pigs, LIRI was produced by: group I (n = 6) LTx with 50 min of CIT (LTx-50 min-CIT); and group II (n = 6) LTx with 6 h of CIT (LTx-6h-CIT). The lung function, edema formation, macroscopic and microscopic changes were assessed. CLDN4 expression in BALF and pulmonary tissue were determined. RESULTS: Both groups presented similar clinical, edema, and histological damage, as well as similar expression of CLDN4 in BALF and tissue (p > 0.05, RM-ANOVA). CONCLUSION: CLDN4 expressed in BALF and the pulmonary tissue during the first 5 h within 72 h of the PGD window are not associated by the deterioration of lung function, edema and lung histological injury, in LTx with CIT 50 min or 6 h, CLDN4 does not seem to be a valuable indicator of LIRI.

Quantifying the environmental impact of clustering strategies in waste management: A case study for plastic recycling from large household appliances.

The complex composition of waste electrical and electronic equipment (WEEE) plastics represents a challenge during post-consumption plastic recycling. A single WEEE category, e.g. large household appliances (LHA), can contain several different plastic types with overlapping material properties, making the sorting of individual plastics a challenge. Significant increases in plastic recovery rates can be expected by clustering product categories, as clustering can avoid mixing of non-compatible plastics with overlapping material properties. For this purpose, a life cycle assessment (LCA) is conducted to investigate the influence of different clustering strategies on the environmental performance of waste treatment and the production of recycled plastic from LHA waste stream. To assure comparability between waste treatment scenarios a system expansion approach is applied, and to allocate the burden of shared processes over the first and second use cycle of the material partitioning is applied. Results show that an increased separation of product clusters by plastic type can improve the plastic recovery rate from 5.8% to 47.1% and reduce the overall environmental impact, quantified with the ReCiPe (2016) method, by up to 23%. The environmental impacts of using recycled plastics from LHA waste can be reduced by 27 to 38% compared to single-use plastic. The holistic approach used in this study demonstrates (1) the potential benefits of implementing product clustering strategies for LHA plastic recycling, (2) the relevance of different allocation procedures when integrating recycling into an LCA, (3) the importance of using less virgin material and avoiding final waste treatment, and (4) the limitation of the recycling system to reduce the environmental burden associated with products.

Central Alteration in Peripheral Neuropathy of Trembler-J Mice: Hippocampal pmp22 Expression and Behavioral Profile in Anxiety Tests.

Charcot-Marie-Tooth (CMT) type 1 disease is the most common human hereditary demyelinating neuropathy. Mutations in pmp22 cause about 70% of all CMT1. Trembler-J (TrJ/+) mice are an animal model of CMT1E, having the same spontaneous pmp22 mutation that is found in humans. We compared the behavior profile of TrJ/+ and +/+ (wild-type) in open-field and elevated-plus-maze anxiety tests. In these tests, TrJ/+ showed an exclusive head shake movement, a lower frequency of rearing, but a greater frequency of grooming. In elevated-plus-maze, TrJ/+ defecate more frequently, performed fewer total entries, and have fewer entries to closed arms. These hippocampus-associated behaviors in TrJ/+ are consistent with increased anxiety levels. The expression of pmp22 and soluble PMP22 were evaluated in E17-hippocampal neurons and adult hippocampus by in situ hybridization and successive immunohistochemistry. Likewise, the expression of pmp22 was confirmed by RT-qPCR in the entire isolated hippocampi of both genotypes. Moreover, the presence of aggregated PMP22 was evidenced in unmasked granular hippocampal adult neurons and shows genotypic differences. We showed for the first time a behavior profile trait associated with anxiety and a differential expression of pmp22/PMP22 in hippocampal neurons of TrJ/+ and +/+ mice, demonstrating the involvement at the central level in an animal model of peripheral neuropathy (CMT1E).

Corneal perforation associated with isolated ocular lichen planus: a case report.

INTRODUCTION: To report a case of unilateral corneal perforation due to isolated ocular lichen planus. METHODS: Interventional case report. Informed consent by the patient was obtained to publish clinical images. A 64-year-old male presented with severe vision loss and a 2-week history of corneal perforation treated with penetrating keratoplasty in the left eye. He had a longstanding diagnosis of severe chronic dry eye disease. On the initial assessment a visual acuity of 20/50 in the RE and HM perception in the left eye were documented. Biomicroscopy revealed subepithelial fibrosis on the tarsal conjunctiva and clinical signs of severe dry eye disease in both eyes. A clear corneal button and a white cataract were observed in the left eye. No other skin or mucosal lesions were observed. RESULTS: An excisional biopsy of the bulbar conjunctiva was performed under topical anesthesia. Direct immunofluorescence analysis revealed a linear deposit of fibrinogen in the basement membrane consistent with ocular lichen planus. Clinical improvement was achieved using aggressive topical lubrication, corneal epithelial regenerators, topical tacrolimus, and immunosuppressive therapy with systemic corticosteroids and cyclophosphamide. CONCLUSION: Isolated ocular lichen planus is an extremely infrequent presentation of lichen planus often indistinguishable from other cicatricial conjunctivitis. Corneal perforation is a severe complication associated with severe dry eye, not previously reported with ocular lichen planus. An adequate clinical assessment and histopathologic diagnosis are crucial to lead prompt treatment and prevent sight-threatening complications.

Experimental design of a culture approach for corneal endothelial cells of New Zealand white rabbit.

The harvesting of corneal endothelial cells (CEC) has received special attention due to its potential as a therapy for corneal blindness. The main challenges are related to the culture media formulation, cellular density at the primary isolation, and the number of passages in which CEC can retain their functional characteristics. To alternate different media formulations to harvest CEC has an impact on the cellular yield and morphology. Therefore, we analyzed four different sequences of growth factor-supplemented Stimulatory (S) and non-supplemented Quiescent (Q) media, upon passages to find the optimal S-Q culture sequence. We assessed cell yield, morphology, procollagen I production, Na+/K+-ATPase function, and the expression of ZO-1 and Na+/K+-ATPase. Our results show SQSQ and SQQQ sequences with a balance between an improved cell yield and hexagonal morphology rate. CEC cultured in the SQQQ sequence produced procollagen I, showed Na+/K+-ATPase function, and expression of ZO-1 and Na+/K+-ATPase. Our study sets a culture approach to guarantee CEC expansion, as well as functionality for their potential use in tissue engineering and in vivo analyses. Thus, the alternation of S and Q media improves CEC culture. SQQQ sequence demonstrated CEC proliferation and lower the cost implied in SQSQ sequences. We discarded the use of pituitary extract and ROCK inhibitors as essential for CEC proliferation.

Decoding Aging: Understanding the Complex Relationship among Aging, Free Radicals, and GSH.

N-aryl maleimides can undergo a 1,4-Michael-type addition reaction with reduced glutathione (GSH), leading to a decreased concentration of GSH and an increased concentration of free radicals (FRs) in cells. GSH is a critical scavenging molecule responsible for protecting cells from oxidation and for maintaining redox homeostasis. N-aryl maleimides disturb redox homeostasis in cells because they scavenge thiol-containing molecules, especially GSH. This study aimed at measuring the concentrations of GSH and FRs by electronic paramagnetic resonance (EPR), in the brain and liver tissue of male Wistar rats (ex vivo) at different ages and after treatment with 3,5-dimaleimylbenzoic acid (3,5-DMB). Our results showed a relationship between age and the concentrations of GSH and FRs in cells. In young rats, the concentration of GSH was higher than in old rats, while the concentration of FRs was higher in adult rats than in young rats, suggesting an inverse relationship between GSH and FRs. On the other hand, the reaction of 3,5-DMB (an electrophilic maleimide) with cellular GSH increased the FR content. The results of this study contribute to the awareness that the process of aging implies not only a loss of tissue function but also essential changes in the molecular contents of cells, especially the concentrations of FRs and GSH.

Treatment with Hyaluronic Acid and Collagen-Polyvinylpyrrolidone Improves Extracellular Matrix Assembly for Scarring after Tracheal Resection.

Treatment of tracheal stenosis is occasionally performed in combination with wound healing modulators to manipulate new extracellular matrix (ECM) formation and prevent fibrosis. Hyaluronic acid (HA) and collagen-polyvinylpyrrolidone (collagen-PVP) decrease fibrosis in experimental tracheal healing. However, they have not been used clinically as their effect on ECM components, which modify tracheal scarring, has not been described. Objective. To evaluate the effect of the application of HA, collagen-PVP, a mixture of HA and collagen-PVP (HA+collagen-PVP), and mitomycin C on the expression of decorin, matrix metalloproteinase 1 (MMP1), and MMP9, as well as the type of collagen and deposits formed in the scar after resection and end-to-end anastomosis (REEA) of the cervical trachea using an experimental model. Materials and Methods. Thirty dogs underwent REEA of the cervical trachea and were treated with different wound healing modulators: group I (n = 6), control; group II (n = 6), HA; group III (n = 6), collagen-PVP; group IV (n = 6), HA+collagen-PVP; and group V (n = 6), mitomycin C. The dogs were evaluated clinically and endoscopically for 4 weeks. Subsequently, macroscopic and microscopic changes, expression of ECM proteins, and collagen deposition in tracheal scars were analysed. Results. Groups II, III, and IV showed reduced endoscopic, macroscopic, and microscopic inflammation, improved neovascularization, high decorin expression (p < 0.01, analysis of variance (ANOVA)), and moderate expression of MMP1 (p < 0.003, ANOVA) and type I and III collagen (p < 0.05, Kruskal-Wallis). Groups IV and V developed fewer collagen deposits (p < 0.001, ANOVA). Conclusion. Treatment with HA and collagen-PVP improved post-REEA healing by increasing neovascularization, stimulating the expression of decorin, and regulating the expression of MMP1, as well as type I and III collagen and their deposition.