Figure 4.

Slit2 alters HAS1 and HAS2 expression levels and HA synthesis in fibrocytes and GD-OF. (A) Fibrocytes were incubated in the absence or presence of rhSlit2 (50 ng/mL) for 7 to 9 days and then were untreated or received bTSH (5 mIU/mL) for 6 hours. (B) GD-OF were transfected with scrambled (control) siRNA (3 µg) or Slit2-targeting siRNA (3 µg) and then re-cultured for 3 days. They were then incubated in the absence or presence of bTSH for 6 hours. Cell layers were harvested, RNA was extracted, reverse-transcribed, and subjected to RT-PCR for the targets indicated. Values were normalized to their respective GAPDH levels. (C) Fibrocytes were untreated or received rhSlit2 as in panel A while in (D) GD-OF were transfected with control or Slit2-targeting siRNA and then in the absence or presence of bTSH (5 mIU/mL) for 72 hours. Media were harvested and subjected to HA ELISA. Values were normalized to cell layer protein content. Data are expressed as the mean ± SD; * P < 0.05; ** P < 0.01; *** P < 0.001 of 3 independent determinations. Experiments were performed 3 times.