Assistant Professor University of Colorado Anschutz Medical Campus Aurora, Colorado, United States
Disclosure(s):
Shaikh Atif, PhD: No financial relationships to disclose
Introduction/Rationale: In chronic beryllium disease (CBD), elevated levels of the inflammatory chemokines CCL3 and CCL4 in the lungs coincide with expanded populations of CD4+ T cells specific to beryllium (Be)-modified peptides derived from these chemokines. Here, we generated HLA-DP2 transgenic (Tg) CCL3-deficient mice (CCL3-/-) that also lack CCL4 to investigate their role in disease development.
Methods: Mice strains: HLA-DP2 Tg WT, FVB-N, CCL3, and TNF-α. Mice were intratracheally administered with BeO (100µg) on days 0,1, and 2 and euthanized at day 12 or boosted at days,14,15,18,19, and euthanized on day 21. BAL fluid (BALF) was obtained from the instillation of 1 ml PBS. BeO-exposed WT HLA-DP2 Tg (CD45.2) mice were injected intravenously with DR-a or CCL4-CAR- T cells on day 8. Flow cytometry, q-PCR, IFN-γ and IL-2 ELISPOT, Histology (H&E), Tetramer staining, RNA-sequencing, and CAR-T cell design and preparation.
Results: Be-exposed CCL3-/- mice maintained normal numbers of lung macrophages and dendritic cells (DCs) but exhibited significantly reduced total and HLA-DP2-CCL/Be tetramer-specific CD4+ T cells, IFN-γ-producing CD4+ T cells, and peribronchovascular aggregates, consistent with attenuated inflammation. CCL3 was predominantly expressed in macrophages and DCs, and bone marrow chimera studies confirmed that hematopoietic-derived DCs are the key regulators of CCL/Be-specific CD4+ T cell responses. RNA sequencing of lung-resident CCL4/Be tetramer-positive CD4+ T cells revealed a transcriptional profile enriched for inflammatory and cholesterol metabolism pathways, with elevated IFN-γ, TNF-α, and IL-17a gene expression. Moreover, Be-exposed HLA-DP2 Tg mice lacking TNF-α or treated with peptide-MHC II CAR-T cells targeting CCL4/Be-specific CD4+ T cells showed reduced T cell responses.
Conclusion: These findings demonstrate that CCL3 and CCL4 promote CCL/Be-specific CD4+ T cell responses and highlight peptide-MHC II CAR-T cells as a novel strategy for depleting self-peptide/Be-specific CD4+ T cells in CBD.
