Changes in the levels of free sialic acid during ex vivo lung perfusion do not correlate with pulmonary function. Experimental model.

BACKGROUND: Ex vivo lung perfusion (EVLP) constitutes a tool with great research potential due to its advantages over in vivo and in vitro models. Despite its important contribution to lung reconditioning, this technique has the disadvantage of incurring high costs and can induce pulmonary endothelial injury through perfusion and ventilation. The pulmonary endothelium is made up of endothelial glycocalyx (EG), a coating of proteoglycans (PG) on the luminal surface. PGs are glycoproteins linked to terminal sialic acids (Sia) that can affect homeostasis with responses leading to edema formation. This study evaluated the effect of two ex vivo perfusion solutions on lung function and endothelial injury. METHODS: We divided ten landrace swine into two groups and subjected them to EVLP for 120 min: Group I (n = 5) was perfused with Steen® solution, and Group II (n = 5) was perfused with low-potassium dextran-albumin solution. Ventilatory mechanics, histology, gravimetry, and sialic acid concentrations were evaluated. RESULTS: Both groups showed changes in pulmonary vascular resistance and ventilatory mechanics (p < 0.05, Student's t-test). In addition, the lung injury severity score was better in Group I than in Group II (p < 0.05, Mann-Whitney U); and both groups exhibited a significant increase in Sia concentrations in the perfusate (p < 0.05 t-Student) and Sia immunohistochemical expression. CONCLUSIONS: Sia, as a product of EG disruption during EVLP, was found in all samples obtained in the system; however, the changes in its concentration showed no apparent correlation with lung function.

Decreased expression of transmembrane TNFR2 in lung leukocytes subpopulations of patients with non-fibrotic hypersensitivity pneumonitis compared with the fibrotic disease.

Hypersensitivity pneumonitis (HP) is an interstitial lung disease, characterized by lung inflammation (non-fibrotic HP) that may often progresses to fibrosis (Fibrotic HP). The tumor necrosis factor (TNF) and its receptors (TNFR1 and TNFR2) can be found as soluble (sol) and transmembrane (tm) forms, playing pro-inflammatory functions but also has been related to immune regulatory functions. Bronchioalveolar lavage from fibrotic and non-fibrotic HP patients was obtained, and immune cells were characterized by flow cytometry, whereas soluble proteins were analyzed by ELISA. Compare to fibrotic HP patients, HP patients with non-fibrotic disease have accumulation of pro-inflammatory CD3+ myeloid cells, cell subpopulations that have decreased tmTNFR2 expression, and low frequency of regulatory-T cells. Whereas solTNF, solTNFR2, and IL-8 are increased. These findings suggest that the TNF pathway may explain, at least partially, the differences between both HP clinical forms. The evaluation of the TNF family molecules may help to develop new therapeutic approaches.

Delayed resolution of bleomycin-induced pulmonary fibrosis in absence of MMP13 (collagenase 3).

Matrix metalloprotease 13 (MMP13) deficiency in pulmonary fibrosis has described contradictory phenotypes on inflammatory and fibrotic responses after lung injury, and its role during lung fibrosis resolution is still undefined. MMP13 has been considered the main collagenase in rodents, and the remodeling of fibrillar collagen is widely attributed to the action of this enzyme. In this study we aimed to explore the role of MMP13 during lung fibrosis progression and resolution. Lung fibrosis was induced by intratracheal instillation, and inflammatory, fibrotic, and resolution stages were evaluated in Mmp13-null and wild-type (WT) mice. Bronchoalveolar lavage fluid was taken for cytokine array analysis and activity of gelatinases. Our results showed that MMP13 is upregulated mainly during two stages after lung injury, inflammation and resolution of fibrosis, and it is mainly expressed by alveolar and interstitial macrophages. Mmp13-null mice exhibited more extensive inflammation at 7 days after bleomycin treatment, and it was characterized by increased macrophage infiltration and significant alterations in proinflammatory cytokines. We also documented that Mmp13-deficient mice experienced more severe and prolonged lung fibrosis compared with WT mice. Delayed resolution in Mmp13-deficient lungs was characterized by a decreased overall collagenolytic activity and persistent fibrotic foci associated with emphysema-like areas. Together, our findings indicate that MMP13 plays an antifibrotic role and its activity is crucial in lung repair and restoration of tissue integrity during fibrosis resolution.

Increased ER Stress and Unfolded Protein Response Activation in Epithelial and Inflammatory Cells in Hypersensitivity Pneumonitis.

Several types of cytotoxic insults disrupt endoplasmic reticulum (ER) homeostasis, cause ER stress, and activate the unfolded protein response (UPR). The role of ER stress and UPR activation in hypersensitivity pneumonitis (HP) has not been described. HP is an immune-mediated interstitial lung disease that develops following repeated inhalation of various antigens in susceptible and sensitized individuals. The aim of this study was to investigate the lung expression and localization of the key effectors of the UPR, BiP/GRP78, CHOP, and sXBP1 in HP patients compared with control subjects. Furthermore, we developed a mouse model of HP to determine whether ER stress and UPR pathway are induced during this pathogenesis. In human control lungs, we observed weak positive staining for BiP in some epithelial cells and macrophages, while sXBP1 and CHOP were negative. Conversely, strong BiP, sXBP1- and CHOP-positive alveolar and bronchial epithelial, and inflammatory cells were identified in HP lungs. We also found apoptosis and autophagy markers colocalization with UPR proteins in HP lungs. Similar results were obtained in lungs from an HP mouse model. Our findings suggest that the UPR pathway is associated with the pathogenesis of HP.

Role of Sonic Hedgehog in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive and lethal disease of unknown etiology and uncertain pathogenic mechanisms. Recent studies indicate that the pathogenesis of the disease may involve the abnormal expression of certain developmental pathways. Here we evaluated the expression of Sonic Hedgehog (SHH), Patched-1, Smoothened, and transcription factors glioma-associated oncogene homolog (GLI)1 and GLI2 by RT-PCR, as well as their localization in IPF and normal lungs by immunohistochemistry. The effects of SHH on fibroblast proliferation, migration, collagen and fibronectin production, and apoptosis were analyzed by WST-1, Boyden chamber chemotaxis, RT-PCR, Sircol, and annexin V-propidium iodide binding assays, respectively. Our results showed that all the main components of the Sonic signaling pathway were overexpressed in IPF lungs. With the exception of Smoothened, they were also upregulated in IPF fibroblasts. SHH and GLI2 localized to epithelial cells, whereas Patched-1, Smoothened, and GLI1 were observed mainly in fibroblasts and inflammatory cells. No staining was detected in normal lungs. Recombinant SHH increased fibroblast proliferation (P < 0.05), collagen synthesis, (2.5 ± 0.2 vs. 4.5 ± 1.0 µg of collagen/ml; P < 0.05), fibronectin expression (2-3-fold over control), and migration (190.3 ± 12.4% over control, P < 0.05). No effect was observed on α-smooth muscle actin expression. SHH protected lung fibroblasts from TNF-α/IFN-γ/Fas-induced apoptosis (14.5 ± 3.2% vs. 37.3 ± 7.2%, P < 0.0001). This protection was accompanied by modifications in several apoptosis-related proteins, including increased expression of X-linked inhibitor of apoptosis. These findings indicate that the SHH pathway is activated in IPF lungs and that SHH may contribute to IPF pathogenesis by increasing the proliferation, migration, extracellular matrix production, and survival of fibroblasts.

[Effects of propofol pretreatment on endothelin in oleic acid-induced acute lung injury]. / Efectos del pretratamiento con propofol sobre la endotelina en daño pulmonar agudo inducido por ácido oleico.

INTRODUCTION: Acute lung injury (ALI) is a pathological condition characterized by injury in the alveolar-capillary membrane that triggers local and systemic inflammation. Endothelin (ET) is a protein that regulates immune response and constricts blood vessels; when it is over-expressed, it may contribute to high blood pressure and lung injury. This work tries to determine if propofol may decrease hemodynamic, gasometric, microscopic, ET-1 plasmatic concentration, and immuno-histochemical alterations in an experimental model of oleic acid-induced acute lung injury. MATERIALS AND METHODS. Animals were classified into three groups (n = 6): group I was the control group; in group II, there was oleic acid-induced ALI with no treatment, and group III with propofol pre-treatment and oleic acid-induced ALI. RESULTS: All animals survived until the end of the study, and 100% of group II and group III developed ALI, with hemodynamic, gasometric and gravimetric alterations. However, group III showed less inflammatory infiltration and lower ET-1 expression in lung tissue. CONCLUSIONS: Pretreatment with propofol in a canine model of OA-induced ALI indicates that the drug has anti-inflammatory action, with a potential therapeutic role against progression of anti-inflammation and lung damage.

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.

Pooled molecular occurrence of Mycoplasma gallisepticum and Mycoplasma synoviae in poultry: A systematic review and meta-analysis.

Worldwide, Mycoplasma gallisepticum (MG) and M. synoviae (MS) are the main agents responsible for chronic respiratory disease in poultry. Therefore, we conducted a systematic review and meta-analysis to estimate their occurrence. We searched electronic databases to find peer-reviewed publications reporting the molecular detection of MG and MS in poultry and used meta-analysis to estimate their pooled global occurrence (combined flock and individual), aggregating results at the regional and national levels. We performed a subgroup meta-analysis for subpopulations (broilers, layers, breeders and diverse poultry including turkeys, ducks and ostriches) and used meta-regression with categorical modifiers. We retrieved 2294 publications from six electronic databases and included 85 publications from 33 countries that reported 62 studies with 22,162 samples for MG and 48 studies with 26,413 samples for MS. The pooled global occurrence was 38.4% (95% CI: 23.5-54.5) for MS and 27.0% (20.4-34.2) for MG. Among regions, Europe and Central Asia had the lowest occurrence for both pathogens, while MG and MS were highly prevalent in South Asia and sub-Saharan Africa, respectively. At the national level, MG occurrence was higher in Algeria, Saudi Arabia and Sudan, whereas China, Egypt and Ethiopia reported higher values of MS. Among the poultry subpopulations, MS and MG were more prevalent in the breeders and layers (62.6% and 31.2%, respectively) than in diverse poultry. The year of publication, the sample size and the level of ambient air pollution (measured indirectly by PM2.5) were associated with the occurrence of both mycoplasmas. Our study revealed high and heterogeneous occurrence values of MG and MS and justifies the need for early detection and improved control measures to reduce the spread of these pathogens.

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.

Comparing Blend of Essential Oils Plus 25-Hydroxy-Vit-D3 Versus Monensin Plus Virginiamycin Combination in Finishing Feedlot Cattle: Growth Performance, Dietary Energetics, and Carcass Traits.

Ninety crossbreed bulls (349.5 ± 8.25 kg initial weight) were used in an 87day trial to compare the effects of a blend of essential oils plus 25-hydroxy-Vit-D3 (EO + HyD) versus the combination of monensin with virginiamycin (MON + VM) on feedlot growth performance and carcass characteristics. Dietary treatments (nine replicates/treatment) were supplemented with 40 mg/kg diet dry matter of MON + VM (equal parts) or with 120.12 mg/kg diet dry matter of a combination of standardized mixture of essential oils (120 mg) plus 0.12 mg of 25-hydroxy-vitamin-D3 (EO + HyD). There were no treatment effects on dry matter intake (DMI, p = 0.63). However, the coefficient of variation in day-to-day DMI was greater for EO + HyD than for MON + VM (11.4% vs. 3.88%, p = 0.04). There were no treatment effects (p ≥ 0.17) on daily weight gain, gain-to-feed ratio, and estimated dietary net energy. Cattle supplemented with EO + HyD had greater Longissimus muscle area (7.9%, p < 0.01) and estimated retail yield (1.6%, p = 0.03), and tended to have heavier (1.7%, p = 0.10) carcass weight. Differences among treatments in dressing percentage, fat thickness, kidney−pelvic−heart fat, and marbling score were not appreciable (p > 0.10). It is concluded that growth performance response and dietary energetic are similar for finishing cattle supplemented with EO + HyD vs. MON + VM. However, compared with MON + VM, supplementation with EO + HyD during the finishing phase may improve carcass Longissimus area and carcass yield.

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.

The effect of CTLA-4Ig, a CD28/B7 antagonist, on the lung inflammation and T cell subset profile during murine hypersensitivity pneumonitis.

Hypersensitivity pneumonitis (HP) is an inflammatory lung disease characterized by an influx of activated T cells to the lung, in which the CD28/B7 costimulatory signals are essential for the T cell activation and the outcome of the inflammatory response. In this study, we investigated the effect of the CD28/B7 antagonist, CTLA-4Ig, on the lung inflammation and the T cell subset profile in experimental Saccharopolyspora recivirgula (SR)-induced HP. C57BL/6 mice were treated with SR or saline during two and three weeks and in addition of CTLA-4Ig was administrated after either the second or third week and mice were sacrificed seven days later. The extent of the lung inflammation was quantified by histopathology and the lung T cell subsets (Treg, Th17, γδT and NKT) were analyzed by flow cytometry. Mice treated with CTLA-4Ig showed a significant decrease in the extent of lung damage (p<0.05), and exhibited a decreased number of inflammatory cells in the bronchoalveolar lavage (BAL) with diminished CD4/CD8 T cell ratio. Also, a significant increase in the percentage of lung γδT (p<0.01) and NKT (p<0.05) cells was observed in two weeks SR-treated mice with the administration of CTLA-4Ig/SR. At 3 weeks, SR-treated mice showed an increased percentage of regulatory T cells but no significantly differences were found in the percentage of Th17 cells when compared with CTLA-4Ig/SR-treated mice. Our findings suggest that the treatment with CTLA-4Ig affects the HP progression and the lung T cell subset kinetics in mice.

Canine thoracic radiographic images as an educational dataset for distance learning and research on vertebral heart score.

Distance learning is a rapidly spreading form of education worldwide and it plays a crucial role to provide access to millions of people in developing countries. However, the benefits of online learning extend far beyond and became increasingly popular also in medical schools, including veterinary medicine [1,2]. In these new teaching settings, high-quality, validated and easily accessible multimedia materials are of great importance, especially in specific fields, such as radiology, where graphic resources can greatly support learning [3]. The dataset presented along with this article reviews examples of 153 canine latero-lateral thoracic radiographic medical images taken and collected at the Small Animal Veterinary Teaching Hospital, Veterinary Sciences Research Institute, Autonomous University of Baja California. serves as a basis for teaching VHS calculation. Images on this dataset contain all relevant anatomical structures in the determination of VHS, which makes them optimal images for practice in calculating VHS and teaching this procedure. The number of cardiothoracic radiographic images presented here can be a great support in learning the calculation of VHS, especially when combined with distance competency-based educators' support. Acquired heart diseases in the dog are very common, clinical evidence of degenerative valvular disease is detected in approximately 30% of dogs aged 13 years and older [4]. The prevalence of Dilated Cardiomyopathy (DCM) is remarkably high in certain breeds, approximately 25% of Irish Wolfhounds, 33% of female Doberman Pinschers, and 50% of male Doberman Pinschers are diagnosed with DCM [5]. Thoracic radiography is a key component in cardiovascular evaluation, which is used to calculate the vertebral heart score (VHS). VHS measurement has been described as one of the most objective methods for assessing cardiomegaly in dogs. VHS, in addition to thorough patient history and physical examination, can be very efficient to raise clinical suspicion of underlying heart disease. The measurement is based on cardiac height and width and is normalized to overall body size by comparison to vertebral body length. ln addition to the initial assessment of heart size, the VHS method is also useful for monitoring changes in heart size in response to treatment or to assess the progression of cardiomegaly over time in canine patients [6]Fig. 1. Importantly, considerable breed and individual variation exist with VHS and dogs can have values that fall outside of the normal range even without any cardiac disease. Thus, while the VHS method is a very informative tool, it should not be used as the only means of diagnosing cardiac disease in any given patient [7].

The effect of short term peripheral parenteral nutrition on treatment outcomes and mortality in critically ill pediatric canine patients.

BACKGROUND: Peripheral parenteral nutrition (PPN) is increasingly considered as an alternative to central parenteral nutrition (CPN) given the higher cost and more frequent clinical complications associated with the latter. However, the assessment of potential risks and benefits of PPN in critically ill pediatric canine patients has not been extensively performed. In this study, we aimed to explore the effect of short-term, hypocaloric PPN on weight loss, length of hospital stay, the incidence of complications, adverse effects, and mortality in critically ill pediatric canine patients. RESULTS: Between August 2015 and August 2018, a total of 59 critically ill pediatric canine patients aged from 1 to 6 months admitted at the Veterinary Sciences Research Institute of the Autonomous University of Baja California were included in this non-randomized clinical trial. Canine pediatric patients were initially allocated to 3 groups: 11 in group 1 receiving parenteral nutrition (PN) supplementation equivalent to 40% of the resting energy requirement (RER), 12 in group 2 receiving supplementation of 50% of the RER, and 36 in group 3 receiving no PN supplementation. After establishing that there was no significant difference between 40 and 50% of PN supplementation, these groups were not separated for downstream analysis. Similar lengths of hospital stays were noted among study subjects who received PN supplementation and those who did not (4.3 ± 1.5 vs. 5.0 ± 1.5, days, p = 0.097). No metabolic-, sepsis- or phlebitis-related complications were observed in any animal in the PPN supplemented group. Higher mortality (19.4% vs. 0%, p = 0.036), and a greater percentage of weight loss (9.24% vs. 0%, p <  0.001) were observed in patients who received no supplementation. CONCLUSION: Even though short-term, hypocaloric PPN did not reduce the length of hospital stay, it was associated with lower mortality and resulted in mitigation of weight loss. In contrast to previous studies evaluating central and peripheral parenteral nutrition protocols, we observed a lower frequency of metabolic, septic, and phlebitis complications using a 40-50% parenteral nutrition treatment. The parenteral nutrition therapeutic intervention used in our study may reduce PN-related adverse effects and promote a favorable disease outcome in critically ill canine patients. Larger studies will be needed to confirm these observations.

Experimental Tracheal Replacement: Angiogenesis and Null Apoptosis Promote Stenosis.

BACKGROUND: Tracheal replacement is a challenge for thoracic surgeons due to stenosis in the trachea-prosthesis anastomosis. We propose that stenosis occurs due to fibrosis as a result of an abnormal healing process, characterized by an increased expression of wound healing growth factors (vascular endothelial growth factor [VEGF], survivin, and CD31), which promote angiogenesis and decrease apoptosis. We analyzed the immunoreactivity of VEGF, survivin, CD31, and caspase-3 in the development of fibrotic stenosis in prosthetic tracheal replacement. METHODS: Fourteen dogs were operated on: group I (n=7) received a 6-ring cervical tracheal segment autograft, while in group II (n=7), a 6-ring segment of the cervical trachea was resected and tracheal continuity was restored with a Dacron prosthesis. The follow-up was 3 months. Immunoreactivity studies for VEGF, survivin, CD31, and caspase-3 were performed. A statistical analysis was done using the Wilcoxon signed rank test. RESULTS: Four animals in group I were euthanized on the 10th postoperative day due to autograft necrosis. Three animals completed the study without anastomotic stenosis. Moderate expression of VEGF (p=0.038), survivin (p=0.038), and CD31 (p=0.038) was found. All group II animals developed stenosis in the trachea-prosthesis anastomotic sites. Microscopy showed abundant collagen and neovascularization vessels. Statistically significant immunoreactive expression of VEGF (p=0.015), survivin (p=0.017), and CD31 (p=0.011) was observed. No expression of caspase-3 was found. CONCLUSION: We found a strong correlation between fibrosis in trachea-prosthesis anastomoses and excessive angiogenesis, moderate to intense VEGF, CD31, and survivin expression, and null apoptotic activity. These factors led to uncontrolled collagen production.

In vivo performance of decellularized tracheal grafts in the reconstruction of long length tracheal defects: Experimental study.

BACKGROUND: The repair of long-segment tracheal lesions remains an important challenge. Nowdays no predictable and dependable substitute has been found. Decellularized tracheal scaffolds have shown to be a promising graft for tracheal transplantation, since it is non-immunogenic. OBJECTIVE: Evaluate in vivo decellularized tracheal allografts performance to replace long tracheal segment. METHODS: Forty-five swines underwent surgery as follows: Fifteen trachea donors and 30 receptors of decellularized trachea allografts. The receptors were randomly divided in five groups (n = 6). In groups I and II, donor trachea segment was decellularized by 15 cycles with sodium deoxycholate and deoxyribonuclease, in group II, the allograft was reinforced with external surgical steel wire. Groups, III, IV, and V decellularization was reduced to seven cycles, supplemented with cryopreservation in group IV and with glutaraldehyde in group V. A 10 rings segment was excised from the receptor swine and the decellularized trachea graft was implanted to re-establish trachea continuity. RESULTS: Both decellularization cycles caused decreased stiffness. All trachea receptors underwent euthanasia before the third post-implant week due to severe dyspnea and trachea graft stenosis, necrosis, edema, inflammation, hemorrhage, and granulation tissue formation in anastomotic sites. Histologically all showed total loss of epithelium, separation of collagen fibers, and alterations in staining. CONCLUSIONS: Both decellularization techniques severely damaged the structure of the trachea and the extracellular matrix of the cartilage, resulting in a no functional graft, in spite of the use of surgical wire, cryopreservation or glutaraldehyde treatment. An important drawback was the formation of fibrotic stenosis in both anastomosis.

Lyophilized glutaraldehyde-preserved bovine pericardium for experimental atrial septal defect closure.

A variety of patch materials has been used to close large atrial septal defects (ASD). Autologous pericardium and glutaraldehyde-preserved bovine pericardium are the most used. Lyophilized bovine pericardium has not been tested inside the cardiovascular system. The aim of this work was to study the behaviour and effectiveness of lyophilized glutaraldehyde-preserved bovine pericardium in ASD closure. Sixteen mongrel dogs were operated on. A 3 cm diameter atrial septal defect was created, and closed with: Group I (n=8): Lyophilized glutaraldehyde preserved bovine pericardium (LGPBP). Group II (n=8): Vascular Dacron patch. The animals were evaluated clinically, by echocardiography, macroscopically, and microscopically. Statistical analysis was done with analysis of variance (ANOVA) and Student's t-test. All the animals survived the surgical procedure and study time (6 months). Clinically all the animals displayed normal physical activity, with normal cardiac sounds. Echocardiography showed that both groups had a normal heart without intracardiac shunts, no thrombus formation, and no vegetations. Macroscopically all the animals showed good integration of the lyophilized bioprosthesis and Dacron patch. All group I animals presented a decrease of the area of the ASD in the left atrium (p<0.001 by ANOVA and Student's t-test). Microscopically, group I presented dense and well-organized collagenous tissue, areas of cartilaginous metaplasia and remnants of the lyophilized bioprosthesis (p<0.001 by ANOVA and Student's t-test). Group II showed encapsulated Dacron patch covered with collagenous tissue and cartilaginous metaplasia. In conclusion, the new lyophilized bioprosthesis is well integrated into the atrial septum, without complications and is effective for ASD closure.

Inflammatory response and dynamics of lung T cell subsets in Th1, Th2 biased and Th2 deficient mice during the development of hypersensitivity pneumonitis.

It is considered that hypersensitivity pneumonitis (HP) occurs with a Th1 cell dominance; however, the role of Th1/Th2 balance is still unclear. C57BL/6 (Th1-biased), BALB/c wt (Th2-biased) and BALB/c Stat6-/- (Th2 deficient) mice were treated with Saccharopolyspora rectivirgula (SR) or saline during 3 weeks, and sacrificed 1 and 4 days (early and late response) after the last administration. Lung isolated T cell subpopulations were analyzed and lung damage extent was quantified. C57BL/6 wt mice exhibited a significant increase in the extent of lung damage when sacrificed at 4 days compared with those sacrificed 1 day after the last SR administration. In contrast, BALB/c wt mice showed a progressive decrease in the extent of lung damage. A significant increase of NKT CD4+ subset was found in C57BL/6 mice while NKT DN cells were increased in BALBc wt mice. Also, NK and gammadelta T cells were increased in BALB/c mice at 1 and 4 days. Stat6-/- mice behave similar to the C57BL/6 mice, showing a progressive increase in the extent of lung damage. A significant increase in the levels of Th1 and Th2 cytokines was observed in bronchoalveolar lavage from the SR-treated mice. These results confirm a predominant role of the Th1 response in HP and suggest that the control of inflammation by Th2 biased mice may be related with the increase of NKT DN cells and regulatory NK and gammadelta T cells.

[Experimental model of canine oleic acid-induced acute lung injury: establishment of a working model]. / Modelo experimental canino de daño pulmonar agudo producido con ácido oleico: estableciendo un modelo de trabajo.

INTRODUCTION: There are several experimental model of acute lung injury induced by oleic acid (OA); however, there are few studies that show how this injury develops. OBJECTIVE: This study seeks to detail the x-ray, hemodynamic, gasometrical, gravimetrical, macroscopic and microscopic alterations developed in an experimental model of canine OA-induced acute lung injury (ALI). MATERIAL AND METHODS: Twelve dogs were divided in 2 study groups: Group I (n=6): Control group without ALI. Group II (n=6); OA-induced ALI. All dogs were submitted to X-ray, hemodynamic and gasometric evaluation before ALI induction, and later every 15 minutes during 150 minutes. At the end of the study, the animals were euthanatized and were evaluated the changes gravimetric, macroscopic and microscopic in injured lungs. RESULTS: All the animals survived through the study. In group II, 100% of the animals developed x-ray (p < 0.003 Wilcoxon), hemodynamic, gasometrical and gravimetric (p < 0.5 ANOVA, Tukey), macroscopically and microscopically (p < 0.001 Wilcoxon) ALI. CONCLUSIONS: The OA-induced ALI is a model in which dogs develop X-ray, hemodynamic, gasometrical, gravimetrical, macroscopically and microscopically injuries of the exudative phase that lung with ALI injury presents.

Identification of Autophagy-related Proteins in Lungs From Hypersensitivity Pneumonitis Patients.

Autophagy has been involved in the pathogenesis of various lung diseases. However, it is not yet known whether autophagy plays a role in hypersensitivity pneumonitis (HP). HP is an interstitial lung disease resulting from exposure to a wide variety of antigens that provoke an exaggerated immune response in susceptible individuals. The aim of this study was to explore the localization of autophagy key proteins in lungs from HP patients and controls by immunohistochemistry and analyze their expression levels by immunoblot. Macrophages and epithelial cells were strongly positive for the autophagosome biomarker LC3B (microtubule-associated protein light chain 3 beta) in HP lungs compared with controls. A similar pattern was found for the autophagy receptor p62 and the enzyme ATG4B. Unexpectedly, nuclear p62 signal was also noticed in macrophages from HP lungs. Regarding ATG5 and ATG7 localization, we observed positive staining in neutrophils, vascular smooth muscle cells, and endothelial cells. Our findings provide for the first time evidence that proteins from the autophagy machinery are highly expressed in the lungs of HP patients and describe the specific cellular and subcellular localization of LC3B, p62, ATG4B, ATG5, and ATG7 in HP lungs.