GM-CSF increases TLR2 and CD14 mRNA levels in neutrophils.

Neutrophils were incubated with GM-CSF (10-100 U/mL) or medium alone for 2 hours, and mRNA was extracted for Northern blot analysis. Positive control mRNA was extracted from Chinese hamster ovary (CHO) cells stably transfected with human CD14 alone or CD14 in combination with human TLR2. Samples, each containing 10 μg RNA, were electrophoresed and blotted on nylon-nitrocellulose membranes. (Equal loading was confirmed by ethidium-bromide staining and hybridization of the blots with an 18S ribosomal cDNA probe.) Northern blots were probed with labeled human TLR2 (upper blot), CD14 (lower blot), and TLR4 (not shown) cDNAs.