Anticoagulant Treatment in Severe ARDS COVID-19 Patients

Patients with COVID-19 may complicate their evolution with thromboembolic events. Incidence of thromboembolic complications are high and also, patients with the critically-ill disease showed evidence of microthrombi and microangiopathy in the lung probably due to endothelial damage by directly and indirectly injured endothelial and epithelial cells. Pulmonary embolism, deep venous thrombosis and arterial embolism were reported in patients with COVID-19, and several analytical abnormal coagulation parameters have been described as well. D-dimer, longer coagulation times and lower platelet counts have been associated with poor outcomes. The use of anticoagulation or high doses of prophylactic heparin is controversial. Despite the use of anticoagulation or high prophylactic dose of heparin have been associated with better outcomes in observational studies, only in patients with non-critically ill disease benefits for anticoagulation was observed. In critically-ill patient, anticoagulation was not associated with better outcomes. Other measures such as antiplatelet therapy, fibrinolytic therapy or nebulized anticoagulants are being studied in ongoing clinical trials

Nebulized Heparin Attenuates Pulmonary Coagulopathy and Inflammation through Alveolar Macrophages in a Rat Model of Acute Lung Injury

Objective Alveolar macrophages play a key role in the development and resolution of acute respiratory distress syndrome (ARDS), modulating the inflammatory response and the coagulation cascade in lungs. Anti-coagulants may be helpful in the treatment of ARDS. This study investigated the effects of nebulized heparin on the role of alveolar macrophages in limiting lung coagulation and inflammatory response in an animal model of acute lung injury (ALI). Methods Rats were randomized to four experimental groups. In three groups, ALI was induced by intratracheal instillation of lipopolysaccharide (LPS) and heparin was nebulized at constant oxygen flow: the LPS/Hep group received nebulized heparin 4 and 8 hours after injury; the Hep/LPS/Hep group received nebulized heparin 30 minutes before and 4 and 8 hours after LPS-induced injury; the LPS/Sal group received nebulized saline 4 and 8 hours after injury. The control group received only saline. Animals were exsanguinated 24 hours after LPS instillation. Lung tissue, bronchoalveolar lavage fluid (BALF) and alveolar macrophages isolated from BALF were analysed. Results LPS increased protein concentration, oedema and neutrophils in BALF as well as procoagulant and proinflammatory mediators in lung tissue and alveolar macrophages. In lung tissue, nebulized heparin attenuated ALI through decreasing procoagulant (tissue factor, thrombin-anti-thrombin complexes, fibrin degradation products) and proinflammatory (interleukin 6, tumour necrosis factor alpha) pathways. In alveolar macrophages, nebulized heparin reduced expression of procoagulant genes and the effectors of transforming growth factor beta (Smad 2, Smad 3) and nuclear factor kappa B (p-selectin, CCL-2). Pre-treatment resulted in more pronounced attenuation. Conclusion Nebulized heparin reduced pulmonary coagulopathy and inflammation without producing systemic bleeding, partly by modulating alveolar macrophages

Lung Extracellular Matrix Hydrogels Enhance Preservation of Type II Phenotype in Primary Alveolar Epithelial Cells

One of the main limitations of in vitro studies on lung diseases is the difficulty of maintaining the type II phenotype of alveolar epithelial cells in culture. This fact has previously been related to the translocation of the mechanosensing Yes-associated protein (YAP) to the nuclei and Rho signaling pathway. In this work, we aimed to culture and subculture primary alveolar type II cells on extracellular matrix lung-derived hydrogels to assess their suitability for phenotype maintenance. Cells cultured on lung hydrogels formed monolayers and maintained type II phenotype for a longer time as compared with those conventionally cultured. Interestingly, cells successfully grew when they were subsequently cultured on a dish. Moreover, cells cultured on a plate showed the active form of the YAP protein and the formation of stress fibers and focal adhesions. The results of chemically inhibiting the Rho pathway strongly suggest that this is one of the mechanisms by which the hydrogel promotes type II phenotype maintenance. These results regarding protein expression strongly suggest that the chemical and biophysical properties of the hydrogel have a considerable impact on the transition from ATII to ATI phenotypes. In conclusion, culturing primary alveolar epithelial cells on lung ECM-derived hydrogels may facilitate the prolonged culturing of these cells, and thus help in the research on lung diseases

Evaluation of the Kinetics of Pancreatic Stone Protein as a Predictor of Ventilator-Associated Pneumonia

Ventilator-associated pneumonia (VAP) is a severe condition. Early and adequate antibiotic treatment is the most important strategy for improving prognosis. Pancreatic Stone Protein (PSP) has been described as a biomarker that increases values 3-4 days before the clinical diagnosis of nosocomial sepsis in different clinical settings. We hypothesized that serial measures of PSP and its kinetics allow for an early diagnosis of VAP. METHODS: The BioVAP study was a prospective observational study designed to evaluate the role of biomarker dynamics in the diagnosis of VAP. To determine the association between repeatedly measured PSP and the risk of VAP, we used joint models for longitudinal and time-to-event data. RESULTS: Of 209 patients, 43 (20.6%) patients developed VAP, with a median time of 4 days. Multivariate joint models with PSP, CRP, and PCT did not show an association between biomarkers and VAP for the daily absolute value, with a hazard ratio (HR) for PSP of 1.01 (95% credible interval: 0.97 to 1.05), for CRP of 1.00 (0.83 to 1.22), and for PCT of 0.95 (0.82 to 1.08). The daily change of biomarkers provided similar results, with an HR for PSP of 1.15 (0.94 to 1.41), for CRP of 0.76 (0.35 to 1.58), and for PCT of 0.77 (0.40 to 1.45). CONCLUSION: Neither absolute PSP values nor PSP kinetics alone nor in combination with other biomarkers were useful in improving the prediction diagnosis accuracy in patients with VAP. Clinical Trial Registration: Registered retrospectively on August 3rd, 2012. NCT02078999

Evaluation of the Kinetics of Pancreatic Stone Protein as a Predictor of Ventilator-Associated Pneumonia

Funding Information: This study was partially supported by Abionic, CIBERES, and Research and Innovation Institute Parc Tauli (I3PT). A.C. acknowledges receiving financial support from Instituto de Salud Carlos III (ISCIII; Sara Borrell 2021: CD21/00087). The funding sources had no role in the design and conduct of the study; the collection, management, analysis, and interpretation of the data; the preparation, review, or approval of the manuscript; or in the decision to submit the manuscript for publication. Publisher Copyright: © 2023 by the authors.BACKGROUND: Ventilator-associated pneumonia (VAP) is a severe condition. Early and adequate antibiotic treatment is the most important strategy for improving prognosis. Pancreatic Stone Protein (PSP) has been described as a biomarker that increases values 3–4 days before the clinical diagnosis of nosocomial sepsis in different clinical settings. We hypothesized that serial measures of PSP and its kinetics allow for an early diagnosis of VAP. METHODS: The BioVAP study was a prospective observational study designed to evaluate the role of biomarker dynamics in the diagnosis of VAP. To determine the association between repeatedly measured PSP and the risk of VAP, we used joint models for longitudinal and time-to-event data. RESULTS: Of 209 patients, 43 (20.6%) patients developed VAP, with a median time of 4 days. Multivariate joint models with PSP, CRP, and PCT did not show an association between biomarkers and VAP for the daily absolute value, with a hazard ratio (HR) for PSP of 1.01 (95% credible interval: 0.97 to 1.05), for CRP of 1.00 (0.83 to 1.22), and for PCT of 0.95 (0.82 to 1.08). The daily change of biomarkers provided similar results, with an HR for PSP of 1.15 (0.94 to 1.41), for CRP of 0.76 (0.35 to 1.58), and for PCT of 0.77 (0.40 to 1.45). CONCLUSION: Neither absolute PSP values nor PSP kinetics alone nor in combination with other biomarkers were useful in improving the prediction diagnosis accuracy in patients with VAP. Clinical Trial Registration: Registered retrospectively on August 3rd, 2012. NCT02078999.publishersversionpublishe

A long-lasting porcine model of ARDS caused by pneumonia and ventilator-induced lung injury

Background: Animal models of acute respiratory distress syndrome (ARDS) do not completely resemble human ARDS, struggling translational research. We aimed to characterize a porcine model of ARDS induced by pneumonia—the most common risk factor in humans—and analyze the additional effect of ventilator-induced lung injury (VILI). Methods: Bronchoscopy-guided instillation of a multidrug-resistant Pseudomonas aeruginosa strain was performed in ten healthy pigs. In six animals (pneumonia-with-VILI group), pulmonary damage was further increased by VILI applied 3 h before instillation and until ARDS was diagnosed by PaO2/FiO2 &lt; 150 mmHg. Four animals (pneumonia-without-VILI group) were protectively ventilated 3 h before inoculum and thereafter. Gas exchange, respiratory mechanics, hemodynamics, microbiological studies and inflammatory markers were analyzed during the 96-h experiment. During necropsy, lobar samples were also analyzed. Results: All animals from pneumonia-with-VILI group reached Berlin criteria for ARDS diagnosis until the end of experiment. The mean duration under ARDS diagnosis was 46.8 ± 7.7 h; the lowest PaO2/FiO2 was 83 ± 5.45 mmHg. The group of pigs that were not subjected to VILI did not meet ARDS criteria, even when presenting with bilateral pneumonia. Animals developing ARDS presented hemodynamic instability as well as severe hypercapnia despite high-minute ventilation. Unlike the pneumonia-without-VILI group, the ARDS animals presented lower static compliance (p = 0.011) and increased pulmonary permeability (p = 0.013). The highest burden of P. aeruginosa was found at pneumonia diagnosis in all animals, as well as a high inflammatory response shown by a release of interleukin (IL)-6 and IL-8. At histological examination, only animals comprising the pneumonia-with-VILI group presented signs consistent with diffuse alveolar damage. Conclusions: In conclusion, we established an accurate pulmonary sepsis-induced ARDS model.</p

A long-lasting porcine model of ARDS caused by pneumonia and ventilator-induced lung injury

Animal models of acute respiratory distress syndrome (ARDS) do not completely resemble human ARDS, struggling translational research. We aimed to characterize a porcine model of ARDS induced by pneumonia-the most common risk factor in humans-and analyze the additional effect of ventilator-induced lung injury (VILI). Bronchoscopy-guided instillation of a multidrug-resistant Pseudomonas aeruginosa strain was performed in ten healthy pigs. In six animals (pneumonia-with-VILI group), pulmonary damage was further increased by VILI applied 3 h before instillation and until ARDS was diagnosed by PaO/FiO < 150 mmHg. Four animals (pneumonia-without-VILI group) were protectively ventilated 3 h before inoculum and thereafter. Gas exchange, respiratory mechanics, hemodynamics, microbiological studies and inflammatory markers were analyzed during the 96-h experiment. During necropsy, lobar samples were also analyzed. All animals from pneumonia-with-VILI group reached Berlin criteria for ARDS diagnosis until the end of experiment. The mean duration under ARDS diagnosis was 46.8 ± 7.7 h; the lowest PaO/FiO was 83 ± 5.45 mmHg. The group of pigs that were not subjected to VILI did not meet ARDS criteria, even when presenting with bilateral pneumonia. Animals developing ARDS presented hemodynamic instability as well as severe hypercapnia despite high-minute ventilation. Unlike the pneumonia-without-VILI group, the ARDS animals presented lower static compliance (p = 0.011) and increased pulmonary permeability (p = 0.013). The highest burden of P. aeruginosa was found at pneumonia diagnosis in all animals, as well as a high inflammatory response shown by a release of interleukin (IL)-6 and IL-8. At histological examination, only animals comprising the pneumonia-with-VILI group presented signs consistent with diffuse alveolar damage. In conclusion, we established an accurate pulmonary sepsis-induced ARDS model. The online version contains supplementary material available at 10.1186/s13054-023-04512-8

Nebulised heparin as a treatment for COVID-19: scientific rationale and a call for randomised evidence

Nebulised unfractionated heparin (UFH) has a strong scientific and biological rationale and warrants urgent investigation of its therapeutic potential, for COVID-19-induced acute respiratory distress syndrome (ARDS). COVID-19 ARDS displays the typical features of diffuse alveolar damage with extensive pulmonary coagulation activation resulting in fibrin deposition in the microvasculature and formation of hyaline membranes in the air sacs. Patients infected with SARS-CoV-2 who manifest severe disease have high levels of inflammatory cytokines in plasma and bronchoalveolar lavage fluid and significant coagulopathy. There is a strong association between the extent of the coagulopathy and poor clinical outcomes. The anti-coagulant actions of nebulised UFH limit fibrin deposition and microvascular thrombosis. Trials in patients with acute lung injury and related conditions found inhaled UFH reduced pulmonary dead space, coagulation activation, microvascular thrombosis and clinical deterioration, resulting in increased time free of ventilatory support. In addition, UFH has anti-inflammatory, mucolytic and anti-viral properties and, specifically, has been shown to inactivate the SARS-CoV-2 virus and prevent its entry into mammalian cells, thereby inhibiting pulmonary infection by SARS-CoV-2. Furthermore, clinical studies have shown that inhaled UFH safely improves outcomes in other inflammatory respiratory diseases and also acts as an effective mucolytic in sputum-producing respiratory patients. UFH is widely available and inexpensive, which may make this treatment also accessible for low- and middle-income countries. These potentially important therapeutic properties of nebulised UFH underline the need for expedited large-scale clinical trials to test its potential to reduce mortality in COVID-19 patients

Clustering COVID-19 ARDS patients through the first days of ICU admission. An analysis of the CIBERESUCICOVID Cohort

Background Acute respiratory distress syndrome (ARDS) can be classified into sub-phenotypes according to different inflammatory/clinical status. Prognostic enrichment was achieved by grouping patients into hypoinflammatory or hyperinflammatory sub-phenotypes, even though the time of analysis may change the classification according to treatment response or disease evolution. We aimed to evaluate when patients can be clustered in more than 1 group, and how they may change the clustering of patients using data of baseline or day 3, and the prognosis of patients according to their evolution by changing or not the cluster.Methods Multicenter, observational prospective, and retrospective study of patients admitted due to ARDS related to COVID-19 infection in Spain. Patients were grouped according to a clustering mixed-type data algorithm (k-prototypes) using continuous and categorical readily available variables at baseline and day 3.Results Of 6205 patients, 3743 (60%) were included in the study. According to silhouette analysis, patients were grouped in two clusters. At baseline, 1402 (37%) patients were included in cluster 1 and 2341(63%) in cluster 2. On day 3, 1557(42%) patients were included in cluster 1 and 2086 (57%) in cluster 2. The patients included in cluster 2 were older and more frequently hypertensive and had a higher prevalence of shock, organ dysfunction, inflammatory biomarkers, and worst respiratory indexes at both time points. The 90-day mortality was higher in cluster 2 at both clustering processes (43.8% [n = 1025] versus 27.3% [n = 383] at baseline, and 49% [n = 1023] versus 20.6% [n = 321] on day 3). Four hundred and fifty-eight (33%) patients clustered in the first group were clustered in the second group on day 3. In contrast, 638 (27%) patients clustered in the second group were clustered in the first group on day 3.Conclusions During the first days, patients can be clustered into two groups and the process of clustering patients may change as they continue to evolve. This means that despite a vast majority of patients remaining in the same cluster, a minority reaching 33% of patients analyzed may be re-categorized into different clusters based on their progress. Such changes can significantly impact their prognosis

Nebulized anti-coagulants as a therapy for acute lung injury and acute respiratory distress syndrome

La síndrome de distrés respiratori agut (ARDS) és una insuficiència respiratòria aguda amb una incidència global a Europa de 17,9 per cada 100.000 persones-any. Tot i els avenços en el tractament de suport dels pacients amb ARDS, la mortalitat continua sent alta (40%) i els pacients que sobreviuen presenten seqüeles persistents. Actualment no existeix un tractament efectiu. La fisiopatologia de l’ARDS es caracteritza per l’activació de la coagulació i la inflamació a nivell pulmonar, juntament amb el trencament de la barrera alveolar-capil·lar. Això comporta la formació d’edema proteic, la infiltració dels neutròfils cap al compartiment alveolar i l'activació dels macròfags cap a un fenotip pro-inflamatori. Estudis previs en models pre-clínics de lesió pulmonar aguda (ALI) i en pacients amb ARDS han demostrat els efectes beneficiosos del anti-coagulants, tot i que aquests efectes positius es veuen contrarestats pel risc d’hemorràgia sistèmica. Els anti-coagulants podrien ser efectius gràcies a la seva activitat anti-inflamatòria a més de les seves propietats anti-coagulants. Atesa l’estreta interacció entre aquestes vies i la seva influència en la permeabilitat, els anti-coagulants també podrien restaurar la barrera alveolar-capil·lar. La nebulització dels anti-coagulants directament al compartiment alveolar podria augmentar l'eficàcia local i disminuir el risc d'hemorràgia sistèmica. La hipòtesi d'aquesta tesi és que l'heparina nebulitzada i/o antitrombina (ATIII) limitaran la resposta pro-inflamatòria i pro-coagulant pulmonar després de la LPA, promovent, també, la restauració de la barrera alveolar-capil·lar. La co-administració dels anti-coagulants directament als pulmons mitjançant nebulització produirà un efecte sinèrgic que potenciarà les propietats de l'heparina i l’ATIII, reduint la lesió pulmonar i evitant el risc d'hemorràgia sistèmica. Com a part d’aquesta tesi es mostren els resultats de l'acció de l'heparina o l’ATIII específicament en poblacions pulmonars primàries de cèl·lules humanes lesionades i l'administració directa d'heparina i/o ATIII als pulmons per nebulització en un model de rata d’ALI. La nebulització d'heparina i/o d’ATIII atenuen la inflamació i coagulació pulmonar sense produir hemorràgia sistèmica en el model d’ALI. El tractament amb heparina nebulitzada modula els macròfags alveolars mitjançant la reducció dels efectors de TGF-β i NF-κB i la via de coagulació i disminueix el reclutament de neutròfils a l'espai alveolar. L'administració local d'ATIII augmenta els efectes beneficiosos en la coagulació, mentre que la combinació d'ATIII i heparina tenen un major impacte en la reducció de la permeabilitat i la disminució de la infiltració de macròfags en el compartiment alveolar. En estudiar l'acció translacional en humans d'ambdós anti-coagulants en poblacions cel·lulars humanes lesionades aïllades de biòpsies pulmonars, l'heparina disminueix l'expressió de marcadors proinflamatoris en els macròfags alveolars i desactiva la via NF-κB en cèl·lules alveolars tipus II; disminuint l'expressió dels seus mediadors i efectors. D’altra banda, l'ATIII redueix els nivells de mediadors proinflamatoris i augmenta les unions estretes en les cèl·lules alveolars tipus II lesionades. Els estudis actuals demostren que l'heparina nebulitzada i l'ATIII poden ser un tractament potencial per a la ARDS, ja que actuen en diferents vies i processos de la fisiopatologia d’aquesta síndrome. L'administració local d'anti-coagulants atenua la lesió pulmonar disminuint la inflamació, la coagulació i proveeix millores en la permeabilitat sense causar hemorràgia sistèmica.Acute respiratory distress syndrome (ARDS) is an acute respiratory failure with a global incidence in Europe of 17.9 per 100,000 person-year. Although significant advances have been performed in supportive care of patients with ARDS, mortality remains high (40%) and survivors present persistent sequelae. An effective pharmacological therapy for this syndrome is not available yet. ARDS pathophysiology involves pulmonary activated coagulation and inflammation together with the breakdown of the alveolar-capillary barrier. This leads to proteinaceous edema, neutrophils infiltration into the alveolar compartment and the activation of macrophages towards a pro-inflammatory phenotype. Beneficial effects of anti-coagulants have been proved in pre-clinical models of acute lung injury (ALI) and in ARDS patients, although systemic bleeding offset its positive effects. Anti-coagulants could be effective for their anti-inflammatory activity in addition to their anti-coagulant properties. Moreover, given the cross talk of these pathways and their influence on permeability, anti-coagulants could also restore the alveolar-capillary barrier. Nebulization of anti-coagulants directly into the alveolar compartment might increase local efficacy and decrease the risk of systemic bleeding. The hypothesis of this thesis is that nebulized heparin and/or antithrombin (ATIII) limit the pro-inflammatory and pro-coagulant response in the lungs after ALI, also promoting the restoration of the alveolar-capillary barrier. The co-administration of both anti-coagulants directly into the lungs via nebulization produces a synergistic effect enhancing the properties of heparin and ATIII, reducing lung injury and avoiding the risk of systemic bleeding. As part of this thesis we are showing the results of the action of heparin or ATIII in specific primary human injured cell lung populations and the direct administration of heparin and/or ATIII into the lungs by nebulization in a rat model of ALI. Nebulized heparin and/or ATIII attenuated pulmonary inflammation and coagulation and did not produce systemic bleeding in the model of ALI. Treatment with nebulized heparin modulated alveolar macrophages through reducing TGF-β and NF-κB effectors and the coagulation pathway and decreased the recruitment of neutrophils into the alveolar space. Local administration of ATIII alone increased beneficial effects in coagulation, while combined ATIII and heparin had a higher impact reducing permeability and decreasing the infiltration of macrophages into the alveolar compartment. The translational action into humans of both anti-coagulants was also studied. In injured human cell lung populations isolated from lung biopsies, heparin diminished the expression of pro-inflammatory markers in alveolar macrophages and deactivated the NF-κB pathway in alveolar type II cells; decreasing the expression of its mediators and effectors. Also, ATIII decreased levels of pro-inflammatory mediators and increased levels of tight junctions in injured alveolar type II cells. The current studies prove that nebulized heparin and ATIII might be a potential treatment for ARDS, as they act in different pathways and processes of the pathophysiology of this syndrome. Local administration of anti-coagulants attenuates lung injury decreasing inflammation, coagulation and proving ameliorations on permeability without causing systemic bleeding