Background: Senolytic agents (dasatinib + quercetin, navitoclax) selectively eliminate senescent cells, reducing SASP burden. However, apoptotic senescent cells release large quantities of hypomethylated mitochondrial DNA (mtDNA) and oxidized nuclear DNA fragments into the extracellular space. These cell-free DNA (cfDNA) fragments structurally resemble bacterial CpG motifs.

Hypothesis: Senolytic-induced mass apoptosis of senescent cells generates a transient surge of hypomethylated cfDNA that activates TLR9 on tissue-resident macrophages, triggering a paradoxical pro-inflammatory wave (IL-6, TNF-α, IFN-α) that accelerates epigenetic aging (GrimAge2, DunedinPACE) in the 2-4 weeks post-treatment. Co-administration of recombinant DNase I (dornase alfa) during the senolytic window will neutralize cfDNA, prevent TLR9 activation, and convert the net effect from transient age acceleration to sustained deceleration.

Testable predictions:

1. Senolytic-treated aged mice (24mo C57BL/6) show elevated plasma cfDNA (>3-fold) and TLR9-dependent IL-6/TNF-α spikes at days 3-7, with GrimAge2 acceleration at day 14.
2. TLR9-knockout mice on identical senolytic regimen show NO transient inflammatory spike or epigenetic age acceleration.
3. Wild-type mice co-treated with senolytics + dornase alfa show cfDNA clearance, blunted TLR9 signaling, and net GrimAge2 deceleration at day 28.
4. In human PBMC ex vivo: conditioned media from senolytic-treated senescent fibroblasts activates TLR9 reporter cells; DNase I pre-treatment abolishes activation.

Clinical relevance: This may explain variable senolytic responses in SLE/RA where TLR9-driven IFN pathways are already dysregulated. Patients with high baseline IFN signatures may experience senolytic-induced flares unless cfDNA is cleared - predicting a pharmacogenomic biomarker (TLR9 polymorphisms, baseline IFN score) for senolytic response stratification.