De-Epithelialization Protocol with Tapered Sodium Dodecyl Sulfate Concentrations Enhances Short-Term Chondrocyte Survival in Porcine Chimeric Tracheal Allografts

Background: Tracheal transplantation is indicated in cases where injury exceeds 50% of the organ in adults and 30% in children. However, transplantation is not yet considered a viable treatment option partly due to high morbidity and mortality associated with graft rejection. Recently, decellularization (decell) has been explored as a technique for creating bioengineered tracheal grafts. However, risk of post-operative stenosis increases due to the death of chondrocytes, which are critical to maintain the biochemical and mechanical integrity of tracheal cartilage. In this project, we propose a new de-epithelialization protocol that adequately removes epithelial, mucosal, and submucosal cells while maintaining a greater proportion of viable chondrocytes. Methods: The trachea of adult male outbred Yorkshire pigs were extracted, decontaminated, and decellularized according to the original and new protocols before incubation at 37 °C in DMEM for 10 days. Chondrocyte viability was quantified immediately following post-decellularization and on days 1, 4, 7, and 10. Histology was performed pre-decellularization, post-decellularization, and post-incubation. Results: The new protocol showed a significant (p < 0.05) increase in chondrocyte viability up to four days after de-ep when compared to the original protocol. We also found that the new protocol preserves ECM composition to a similar degree as the original protocol. When scaffolds created using the new protocol were re-epithelialized, cell growth curves were near identical to published data from the original protocol. Conclusion: While not without limitations, our new protocol may be used to engineer chimeric tracheal allografts without the need for cartilage regeneration

Mathematical models applied in the assessment of respiratory mechanics in mice with smooth muscle´s agonist challenges.

Modelos matemáticos são utilizados como ferramentas na avaliação da mecânica respiratória para a compreensão da fisiologia e patologias do sistema respiratório. A presente pesquisa visou avaliar, através da aplicação de modelos matemáticos, a mecânica respiratória em camundongos submetidos à metacolina. Deu-se ênfase no modelo linear de compartimento único e suas variantes não lineares. Camundongos C57BL/6 (n = 8) foram traqueostomizados, ventilados mecanicamente (flexiVent, SCIREQ, Canadá) e perturbações em volume foram aplicadas para a modelagem do sistema respiratório. O protocolo experimental foi elaborado de forma a se analisar a variação dos parâmetros respiratórios durante a aplicação do agente broncoativo e também se verificou a divisão do sinal quasi-senoidal em expirações e inspirações durante a técnica de oscilação forçada (FOT) com perturbação de frequência 2,5 Hz. Com base nisso, uma rotina computacional própria foi desenvolvida para a análise dos experimentos realizados no ventilador mecânico e foram pesquisadas as vantagens e desvantagens dos modelos matemáticos aplicados. Os resultados demonstraram um aumento no desvio padrão dos parâmetros do modelo linear unicompartimental e suas variantes não lineares após a aplicação do broncoconstritor. Acredita-se que esta grande variação nos parâmetros esteja relacionada com o enrijecimento do parênquima e da heterogeneidade da ventilação pulmonar após a utilização da droga. Devido à correlação dos parâmetros do modelo com a fisiologia ocorrer somente no modelo linear, acredita-se que este ainda é o mais indicado na avaliação da mecânica respiratória e as variantes não lineares seriam indicadas como opção em casos onde o modelo linear é incapaz de realizar ajustes adequados ou para informações complementares.Mathematical models are used as tools in the assessment of respiratory mechanics for the understanding of the physiology and pathologies of the respiratory system. This study aimed to assess the respiratory mechanics by applying mathematical models in mice subjected to challenges with methacholine. Emphasis was placed on linear single-compartment model and its nonlinear variants. C57BL/6 mice (n = 8) were tracheostomized, mechanically ventilated (flexiVent, SCIREQ, Canada) and disturbances in volume were applied to the modeling of the respiratory system. The experimental protocol was developed in order to analyze the variation of respiratory parameters during the application of the bronchoactive agent. The division of quasisinusoidal signal in expirations and inspirations during the forced oscillation technique (FOT) with frequency perturbation of 2.5 Hz was also observed. Based on that, a proper computational routine was developed in order to analyze the experiments in the mechanical ventilator and the advantages and disadvantages of the applied mathematical models. The results demonstrated an increase in the standard deviation of the linear single-compartment model and its nonlinear variants parameters after the application of bronchoconstrictor. It is believed that this large variation in the parameters relates to the parenchyma stiffening and to the heterogeneity of pulmonary ventilation after the use of the drug. Due to the fact that the correlation between the model parameters and the physiology occurred only in the linear model, it is believed that this is still the most suitable model in the assessment of respiratory mechanics. Nonlinear variations of the single-compartment model would be indicated only as an option, for example, in cases where the linear model is incapable of performing appropriate fits or when additional information about the respiratory system is required

Assessment of the mechanics of a tissue-engineered rat trachea in an image-processing environment

OBJECTIVES: Despite the recent success regarding the transplantation of tissue-engineered airways, the mechanical properties of these grafts are not well understood. Mechanical assessment of a tissue-engineered airway graft before implantation may be used in the future as a predictor of function. The aim of this preliminary work was to develop a noninvasive image-processing environment for the assessment of airway mechanics.METHOD: Decellularized, recellularized and normal tracheas (groups DECEL, RECEL, and CONTROL, respectively) immersed in Krebs-Henseleit solution were ventilated by a small-animal ventilator connected to a Fleisch pneumotachograph and two pressure transducers (differential and gauge). A camera connected to a stereomicroscope captured images of the pulsation of the trachea before instillation of saline solution and after instillation of Krebs-Henseleit solution, followed by instillation with Krebs-Henseleit with methacholine 0.1 M (protocols A, K and KMCh, respectively). The data were post-processed with computer software and statistical comparisons between groups and protocols were performed.RESULTS: There were statistically significant variations in the image measurements of the medial region of the trachea between the groups (two-way analysis of variance [ANOVA], p<0.01) and of the proximal region between the groups and protocols (two-way ANOVA, p<0.01).CONCLUSIONS: The technique developed in this study is an innovative method for performing a mechanical assessment of engineered tracheal grafts that will enable evaluation of the viscoelastic properties of neo-tracheas prior to transplantation

Geoeconomic variations in epidemiology, ventilation management, and outcomes in invasively ventilated intensive care unit patients without acute respiratory distress syndrome: a pooled analysis of four observational studies

Background: Geoeconomic variations in epidemiology, the practice of ventilation, and outcome in invasively ventilated intensive care unit (ICU) patients without acute respiratory distress syndrome (ARDS) remain unexplored. In this analysis we aim to address these gaps using individual patient data of four large observational studies. Methods: In this pooled analysis we harmonised individual patient data from the ERICC, LUNG SAFE, PRoVENT, and PRoVENT-iMiC prospective observational studies, which were conducted from June, 2011, to December, 2018, in 534 ICUs in 54 countries. We used the 2016 World Bank classification to define two geoeconomic regions: middle-income countries (MICs) and high-income countries (HICs). ARDS was defined according to the Berlin criteria. Descriptive statistics were used to compare patients in MICs versus HICs. The primary outcome was the use of low tidal volume ventilation (LTVV) for the first 3 days of mechanical ventilation. Secondary outcomes were key ventilation parameters (tidal volume size, positive end-expiratory pressure, fraction of inspired oxygen, peak pressure, plateau pressure, driving pressure, and respiratory rate), patient characteristics, the risk for and actual development of acute respiratory distress syndrome after the first day of ventilation, duration of ventilation, ICU length of stay, and ICU mortality. Findings: Of the 7608 patients included in the original studies, this analysis included 3852 patients without ARDS, of whom 2345 were from MICs and 1507 were from HICs. Patients in MICs were younger, shorter and with a slightly lower body-mass index, more often had diabetes and active cancer, but less often chronic obstructive pulmonary disease and heart failure than patients from HICs. Sequential organ failure assessment scores were similar in MICs and HICs. Use of LTVV in MICs and HICs was comparable (42·4% vs 44·2%; absolute difference -1·69 [-9·58 to 6·11] p=0·67; data available in 3174 [82%] of 3852 patients). The median applied positive end expiratory pressure was lower in MICs than in HICs (5 [IQR 5-8] vs 6 [5-8] cm H2O; p=0·0011). ICU mortality was higher in MICs than in HICs (30·5% vs 19·9%; p=0·0004; adjusted effect 16·41% [95% CI 9·52-23·52]; p&lt;0·0001) and was inversely associated with gross domestic product (adjusted odds ratio for a US$10 000 increase per capita 0·80 [95% CI 0·75-0·86]; p&lt;0·0001). Interpretation: Despite similar disease severity and ventilation management, ICU mortality in patients without ARDS is higher in MICs than in HICs, with a strong association with country-level economic status