Macrophages were incubated with PMA-stimulated neutrophils for 4 hours. For the last 30 min, DNase I (10 U/ml; M0303S, New England Biolabs) was added. Subsequently, the cells were thoroughly washed with PBS, resuspended in 0.25 M sucrose, and scraped. The cell membranes were disintegrated by passing the samples 15 times through a 20-gauge needle. The cell lysates were centrifuged at 1000g for 5 min at 4°C to remove cellular debris. The DNA was precipitated with sodium acetate and resuspended in H2O. Isolated DNA was digested with 5 U of DNAse 1 for 30 min at 37°C and separated on a 1% agarose gel. Genomic RSP18 DNA was quantified by qPCR using the following primers: hRSP18, 5′-TCCATTTAGAGCTGCCCTACTCA-3′ (forward) and 5′-CTTAACTGCAACTGGCTAGGCT-3′ (reverse). The ratio of mitochondrial to genomic DNA primers for S16 [5′-CGCATAAGCCTGCGTCAGATCAA-3′ (forward) and 5′-TGTGTTGGGTTGACAGTGAGGG-3′ (reverse)] and S18 [5′-GTAACCCGTTGAACCCCATT-3′ (forward) and 5′-CCATCCAATCGGTAGTAGCG-3′ (reverse)] was determined as described previously (37).

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