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. 2014 Apr 23;3(6):745–759. doi: 10.5966/sctm.2013-0200

Figure 1.

Figure 1.

EPI induces TLR2, MyD88, and IL-6 expression in BM-MSCs. (A): BM-MSCs (passages 3–5) were exposed to EPI (50–1,000 nM), and secreted IL-6 levels in the cell culture supernatants were determined by enzyme-linked immunosorbent assay, as described in Materials and Methods. Values are expressed as pg/μg protein (mean ± SD). ∗∗, p < .05 versus 4-hour control (C); ∗∗, p < .05 versus 24-hour control (n = 4). (B): TLR2 mRNA expression was measured in EPI (50 nM)-treated BM-MSCs using reverse transcription-polymerase chain reaction (RT-PCR), as described in Materials and Methods. Values are expressed as fold change versus control. MALP2 (100 ng/ml) was used as a positive control, as described in Materials and Methods. ∗, p < .05 versus control (n = 4). (C): TLR2 and MyD88 protein levels were measured in EPI (50 nM)-treated BM-MSC lysates using Western blot assay. GAPDH was used as the loading control, and MALP2 (100 ng/ml) as positive control. (D): Densitometric analysis of the Western blots. Protein/GAPDH ratio values are expressed as fold change versus control (mean ± SD). ∗, p < .05 versus control (n = 4). MALP2 induces β2-AR mRNA and protein expression in BM-MSCs. (E): β2-AR (ADRB2) mRNA expression was measured in MALP2 (100 ng/ml)-treated cells using RT-PCR. Values are expressed as fold change (in mRNA/housekeeping gene ratio) versus control (mean ± SD), as described in Materials and Methods. ∗, p < .001 versus control (n = 3). EPI (50 nM) was used as a positive control. (F): β2-AR protein expression was measured in MALP2 (100 ng/ml)-treated cells by Western blot. α-Tubulin was used as the loading control, and EPI (50 nM) as positive control. (G): Densitometric analysis of the Western blots. β2-AR/α-tubulin ratio values are expressed as fold change versus control (mean ± SD). ∗, p < .05 versus control (n = 4). Abbreviations: ADRB2, β2-adrenergic receptor; β2-ARs, β2-adrenergic receptors; C, control; EPI, epinephrine; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; IL-6, interleukin-6; MALP2, macrophage-activating lipopeptide-2; MyD88, myeloid differentiation factor 88; TLRs, Toll-like receptors.