Figure 1.

Overexpression of TIPE2 in AA-FLSs.

Notes: PCR and Western blot analyses of TIPE2 expression in freshly harvested RAW 264.7 cells, normal FLSs, AA-FLSs, and AA-FLSs stably transfected with the lentivirus vector MIGR1/TIPE2^+/+ plasmid and the control vector MIGR1 plasmid. TIPE2 expression in RAW 264.7 cells was used as a positive control. (A) RT-PCR analysis of TIPE2 mRNA. GAPDH was used as a loading control, whereas H₂O was used as the background control. (B) Quantification of mRNA expression by quantitative real-time PCR. Data are mean ± SD (n=3), ^***P<0.001 vs the normal FLS group. (C) Whole cell lysates were prepared. An aliquot (30 μg) of each whole cell lysate was fractionated on a 12% SDS gel, and TIPE2 was analyzed by Western blotting. β-Actin was used as a loading control. (D) Grayscale analysis of Figure 1C, ^***P<0.001 vs the normal FLS group. (E) mRNA levels of TNFAIP8 family members were determined by RT-PCR using total RNA isolated from cell preparations. GAPDH was used as a loading control, whereas H₂O was used as the background control. (F) Quantification of mRNA expression by quantitative real-time PCR. Data are mean ± SD (n=3), ^***P<0.001 vs the MIGR1-FLS group. (G) Whole cell lysates were prepared. An aliquot (30 μg) of each whole cell lysate was fractionated on a 12% SDS gel, and TIPE2 and β-actin were analyzed by Western blotting. (H) Grayscale analysis of Figure 1G, ^***P<0.001 vs the MIGR1-FLS group.

Abbreviations: AA, adjuvant arthritis; FLS, fibroblast-like synoviocyte; GAPDH, glyceraldehyde-phosphate dehydrogenase; mRNA, messenger RNA; RT-PCR, reverse transcription polymerase chain reaction; SD, standard deviation; SDS, sodium dodecyl sulfate.