The role of alveolar macrophages (AMs) in lung carcinogenesis has been extensively studied, yielding significant insights. However, the status of AMs in tumor-bearing lungs remains incompletely characterized. Using orthotopic Lewis Lung Carcinoma (LLC) mouse models, we found that tumors induced an inflammatory extra-tumoral lung microenvironment (ETLME), distinct from the immunosuppressive tumor microenvironment (TME). T cells with an exhaustion phenotype and tumor-associated macrophages (TAMs) mainly accumulated in the TME rather than the ETLME. Surprisingly, AMs were absent from the tumor lesions and remained in the lung tissues, but they displayed a more active dynamic balance between proliferation and death in ETLME. Furthermore, AMs presented an activated phenotype characterized by upregulation of CD11b and downregulation of Siglec-F, elevated expression of inflammatory genes, and enhanced phagocytic and efferocytotic activity. Notably, AMs in ETLME retained their lipid metabolism capacity and responsiveness to external stimuli. More importantly, LLC-experienced AMs display enhanced anti-tumor ability. These findings indicate that AMs maintain their tissue localization and functional integrity within the ETLME.
Copyright © 2025 Ren, Dou, Yue, Ma, Yu, Shang, Wang, Wang, Li and Li.

Following ischemic stroke, peripheral immune cell infiltration is characterized by myeloid cell predominance in the acute phase and lymphoid cell infiltration in the subacute to chronic phases. Endothelial cells, as a critical interface between the peripheral circulation and the brain, upregulate adhesion molecules to facilitate immune cell infiltration. However, it remains unclear whether endothelial cells exhibit functional differences at different stages after ischemic stroke and how these differences affect immune cell infiltration.
We performed single-cell RNA sequencing on peripheral immune and endothelial cells from Sham and middle cerebral artery occlusion (MCAO) mice at 3 and 14 days post-MCAO. Subsequent analysis of the sequencing data, combined with flow cytometry and immunofluorescence staining, was used to investigate the relationship between endothelial cell changes at different stages of stroke and immune cell infiltration.
We observed that the infiltration capacity of peripheral immune cells did not significantly increase at different stages after MCAO. However, endothelial cells underwent significant changes. By Day 3 post-MCAO, there was an increased proportion of venous endothelial cells with enhanced angiogenesis and adhesion functions. In this acute phase, newly formed venous endothelial cells with high expression of the adhesion molecule ICAM-1 were observed, promoting the infiltration of myeloid cells and NKT cells. From the acute to chronic phases, endothelial angiogenesis gradually decreased, accompanied by a marked increase in antigen presentation function. At 14 days post-MCAO, an increased proportion of VCAM-1-expressing venous endothelial cells was observed, potentially facilitating the infiltration of T cells and a subset of neutrophils. Furthermore, we discovered that the differential changes in venous endothelial cells at different stages after MCAO may be driven by distinct differentiation and proliferation patterns regulated by different signaling pathways.
Our study highlights that the differential expression of adhesion molecules and functional changes in endothelial cells at distinct stages after ischemic stroke may regulate the infiltration patterns of peripheral immune cells.
© 2025 The Author(s). CNS Neuroscience & Therapeutics published by John Wiley & Sons Ltd.

Recent studies suggest that CD4+ T cells can exert potent anti-tumor effects and improve immunotherapy efficacy by aiding CD8+ T cells. However, characterizing the mechanism of CD4+ T cells' anti-tumor activity has been challenging due to inaccurate major histocompatibility complex class II (MHC-II) peptide prediction algorithms and the lack of high-quality reagents for immune monitoring. To address this, we developed MHC2-substitution of CLIP and analytical LCMS evaluation (MHC2-SCALE), a streamlined approach combining affinity optimized class II-associated invariant chain peptide (CLIP) exchange technology, high throughput 2D-LCMS analysis, and rapid generation of peptide-bound MHC-II monomers for subsequent multimer assembly. We validated MHC-II peptide candidates predicted by the immune epitope database (IEDB) algorithm, as well as uncovered many true and immunogenic MHC-II binders that were not predicted by IEDB. Thus, MHC2-SCALE expands the opportunities for discovering, tracking, and phenotyping antigen-specific CD4+ T cells in preclinical and clinical settings, thereby improving therapies for cancer, autoimmunity, or infectious diseases.
© 2025 The Author(s).

MIWI2, a P element-induced wimpy testes (PIWI) argonaute protein known for suppressing retrotransposons during male gonadogenesis, has an unexplored role in mammalian somatic cells. We identify MIWI2 multiciliated (M2MC) cells as a rare subset of airway multiciliated cells and investigate MIWI2's function in antiviral host defense. We analyzed transcriptomes from Miwi2 heterozygous (Miwi2 +/tom) and deficient (Miwi2 tom/tom) mice following influenza A infection. During infection, Miwi2 deficiency was associated with reduced mitochondrial and ribosomal gene expression in M2MC cells, increased mitochondrial reactive oxygen species (ROS) production and ADP/ATP ratios in multiciliated cells, and enhanced viral clearance and recovery. Additionally, Miwi2-expressing cells exhibited reduced levels of small RNAs derived from nuclear mitochondrial DNA. These findings reveal a previously unrecognized role for Miwi2 in regulating small non-coding RNAs and mitochondrial oxidant production in somatic cells, indicating a function beyond its established germline activities. Our study identifies Miwi2/Piwil4 as a potential factor influencing susceptibility to severe respiratory infections.
© 2025 The Author(s).

Ischemic stroke recovery involves dynamic interactions between the central nervous system and infiltrating immune cells. Peripheral immune cells compete with resident microglia for spatial niches in the brain, but how modulating this balance affects recovery remains unclear. Here, we use PLX5622 to create spatial niches for peripheral immune cells, altering the competition between infiltrating immune cells and resident microglia in male mice following transient middle cerebral artery occlusion (tMCAO). We find that early-phase microglia attenuation promotes long-term functional recovery. This intervention amplifies a subset of monocyte-derived macrophages (RAMf) with reparative properties, characterized by high expression of GPNMB and CD63, enhanced lipid metabolism, and pro-angiogenic activity. Transplantation of RAMf into stroke-affected mice improves white matter integrity and vascular repair. We identify Mafb as the transcription factor regulating the reparative phenotype of RAMf. These findings highlight strategies to optimize immune cell dynamics for post-stroke rehabilitation.
© 2025. The Author(s).