Abstract 4137414: Uncovering MLKL as a Novel Regulator of Endothelial Cells in the Splenic Niche to Repress Hematopoiesis During Atherosclerosis
Bibliographic record
Abstract
Background: Atherosclerosis occurs as lipid and immune cell rich plaques deposit within the arterial wall of the heart. These immune cells are produced from hematopoietic stem and progenitor cells (HSPCs) in the bone marrow (BM) and spleen, a process known as hematopoiesis that is dictated by their microenvironment, including endothelial cells (ECs). Objective: While others have shown adverse remodeling of BM ECs during atherogenesis, whether splenic ECs show a similar dysfunction leading to exacerbated hematopoiesis has not been described. Methods: We analyzed RNA sequencing data from chow fed C57Bl/6 mice and high cholesterol diet (HCD) fed Apoe -/- mouse BM and splenic ECs obtained from published studies, as well as our own. In vivo studies were performed in mice on an atherogenic background ( Apoe -/-& Ldlr -/-) fed a HCD for 4 to 16 weeks. Knockdown of the mixed lineage kinase domain-like protein ( Mlkl KD) was achieved by weekly administration of antisense oligonucleotides (ASOs) or scrambled control (50mg/kg sc.). In vitro studies were performed in primary mouse splenic ECs transfected with siRNA. Results: Pathway analysis revealed remarkable tissue-specific responses to an atherogenic milieu, including dysregulation of Lipid Metabolism, Endocytosis and Cell Death pathways in splenic ECs. Interestingly, we previously showed that the mixed lineage kinase domain-like protein (MLKL) regulates the endocytic trafficking of lipids and cell death in macrophages. Indeed using Mlkl KD Apoe -/- mice, transplantation of Mlkl -/- BM into Ldlr -/- mice or EC-specific Mlkl -/- Apoe -/- mice, we show that loss of MLKL drives myelopoiesis and plaque development through regulation of splenic ECs. Both in vivo and in vitro , silencing Mlkl in splenic but not BM ECs potently increased lipid content and disrupted endocytosis by an accumulation of the multivesicular body marker CHMP4B, which mirror the dysregulated pathways we identified above. Furthermore, co-culture with Mlkl KD splenic ECs increased HSPC activation (pStat5) and myeloid colony formation, an effect that was lost when ECs were pre-treated with the upstream endocytosis inhibitor Dynasore. Conclusions: In conclusion, we establish a novel role for MLKL in regulating the balance of HSPCs, specifically through preservation of splenic, but not BM, ECs that repress hematopoiesis, highlighting the importance of splenic lipid metabolism and endocytosis and its impact on atherogenesis.
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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) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.003 | 0.001 |
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".