399 Establishing an in Vivo Model to Study Pulmonary Neutrophil Extracellular Trap (NET) Formation After Burn Injury
Bibliographic record
Abstract
It is well known that burn patients suffer from serious lung complications such as acute respiratory distress syndrome, acute lung injury and pneumonia. However, the reasons for the development of these conditions after burn injury (BI) are not clearly understood. Several studies examined the importance of macrophages after burn injury. However, the involvement of neutrophils in lung complications after BI has not been studied in detail. Particularly, the importance of recently identified neutrophil extracellular traps (NETs) in the lungs after BI is unknown. NETs are considered to help trap infectious agents to protect the host; however, excess NETs could damage and destroy the airways and cause lung dysfunction. There are no good animal models available to study the pulmonary NETs during BI. Therefore, we established an in vivo model to study NETosis in the lungs after BI using an LPS model. We have first created a 15% body burn, and instilled various amounts of LPS (0–50 mg/kg) into the airways of C57B/6 mice. After various time points (3 h, 16 h, 24 h, 48 h, 72 h) we have collected bronchoalveolar lavage (BAL) fluid and blood samples. Immune cells present in the BAL fluid were deposited on slides by Cytospin preparations, stained and quantified by microscopy. Cell and platelet counts in the blood samples were determined by an automated cell counter, and confirmed by blood smears, H and E staining and microscopy. DNA-protein complexes present in the BAL supernatant were analyzed by agarose gel electrophoresis and pocigreen assays. Presence of a NET marker, citrullinated histone, was analyzed by Western blots. The data obtained from these studies show that neutrophils are not detectable in the airways under baseline or after BI; however, different numbers of neutrophils and amounts of NETs were present under various experimental conditions and time points-post BI. These data show that we could measure NET components in the airways of mice instilled with LPS after BI. Blood analyses show that cells concentrations also differ among various experimental conditions, indicating the importance of neutrophil and NET-promoting components (e.g., platelets) in the blood after BI and/or LPS instillation. We have successfully established a mouse model to study pulmonary NETosis in BI, and optimized the range of LPS concentrations and time points necessary to observe differences in NETosis under various experimental conditions. This model should help to understand the roles of NETs in pulmonary dysfunction after BI, and for testing potential drugs for correcting NET-mediated lung complications.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 0.001 |
| Bibliometrics | 0.001 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.001 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.001 | 0.002 |
| Insufficient payload (model declined to judge) | 0.003 | 0.002 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".