TLR signaling in PBMCs acquired from BALB/c mice (10 days post-BMT) after being transplanted with T-cell depleted bone marrow alone (BMTCD), or in combination with TLR2,4,7-tolerant splenocytes, over their PBS-pretreated counterparts.

Murine splenocytes were isolated from the spleens of C57BL/6J mice and were pretreated in vitro for three days in the presence of PBS, Pam3CSK4 (1 μg/ml), high pure LPS from E.coli O111:B4 (100 ng/ml) and R848 (5 μg/ml). BALB/c irradiated recipients, were transplanted with T-cell depleted bone marrow alone, or with the aforementioned pretreated splenocytes, and 10 days after, PBMCs from all experimental groups were collected for TLR signaling pathway analysis.We used Qiagen Toll-like Receptor RT2 Profiler PCR Array kit to quantitate gene expression profiling of the TLR signaling pathway. qPCR gene expression profiling. PBMCs from 5 mice were used and treated separately as indicated in the summary. Equal amount of total cDNA from each mouse (reverse-transcribed from 0.5 μg RNA) was used prior to gene expression analysis.