Journal:

Article Title: In Vivo Modulation of FGF Biological Activity Alters Cranial Suture Fate

doi:

Figure Lengend Snippet: A: Representative Northern blot analysis demonstrating an increase in TGF-β1 mRNA expression in RNA isolated from vehicle and AdCALacZ-infected NRCs in response to a 6-hour stimulation with FGF-2 (10 ng/ml). This increase in TGF-β1 mRNA in response to FGF-2 was not seen in NRCs infected with AdCAFGF-TR. A similar pattern of TGF-β1 mRNA expression was seen in PFDCs after infection and stimulation (data not shown). 50 = 50 plaque forming units/cell; 100 = 100 plaque forming units/cell. B: Representative Northern blot analysis demonstrating an increase in collagen type I mRNA expression in RNA isolated from vehicle and AdCALacZ-infected PFDCs in response to 48-hour stimulation with FGF-2 (10 ng/ml). PFDCs infected with AdCAFGF-TR failed to demonstrate an increase in collagen type I mRNA. Interestingly, infected NRCs did not demonstrate differences in collagen type I gene expression in response to FGF-2 (data not shown). C: Cells infected with AdCAFGF-TR proliferated significantly slower than cells infected with vehicle and AdCALacZ. Graphic representation of BrdU incorporation by NRCs infected with vehicle, AdCALacZ, or AdCAFGF-TR. Note statistically significant decreased proliferation in NRCs infected with AdCAFGF-TR (*, P < 0.001). All cells were stimulated with rhFGF-2. D: Results of proliferation assay. Note significantly decreased proliferation of NRCs infected with AdCAFGF-TR (*, P < 0.001). All cells were stimulated with rhFGF-2. E: Representative Northern blot analysis demonstrating an increase in baseline TGF-β1 mRNA expression in RNA isolated from AdCAsFGF-2-infected NRCs. Similar pattern of TGF-β1 mRNA expression was seen in PFDCs after AdCAsFGF-2 infection (data not shown). F: Cells infected with AdCAsFGF-2 proliferated significantly faster than cells infected with vehicle and AdCALacZ. Graphic representation of BrdU incorporation by NRCs infected with vehicle, AdCALacZ, or AdCAsFGF-2. Note statistically significantly increased proliferation in NRCs infected with AdCAsFGF-2. (*, P < 0.001). G: Results of proliferation assay. Note significantly increased proliferation of NRCs infected with AdCAsFGF-2 (*, P < 0.001).

Article Snippet: After 24 hours (ie, 48 hours after infection), 10 ng/ml of recombinant human basic FGF (rhFGF-2; R&D Systems, Minneapolis, MN) was added to the serum-free media.

Techniques: Northern Blot, Expressing, Isolation, Infection, Gene Expression, BrdU Incorporation Assay, Proliferation Assay

Journal:

Article Title: In Vivo Modulation of FGF Biological Activity Alters Cranial Suture Fate

doi:

Figure Lengend Snippet: A: Northern (left) and Western blot (right) analysis demonstrating high-level expression of FGF-TR. Note increase in FGF-TR mRNA and protein with increasing plaque-forming units. Compare AdCAFGF-TR 50 with AdCAFGF-TR 100. 50 = 50 plaque-forming units/cell; 100 = 100 plaque-forming units/cell. B: Western blot analysis demonstrating increased phosphorylation of ERK-1 and -2 after FGF-2 stimulation (+) in vehicle and AdCALacZ-infected NRCs (top immunoblot). In contrast, AdCAFGF-TR-infected NRCs demonstrate no increase in ERK-1 and -2 phosphorylation in response to rhFGF-2 stimulation. Total ERK-2 immunoblot of same blot (bottom immunoblot) demonstrating presence of similar levels of unphosphorylated protein in all lanes. C: Western blot analysis for FGF-2 demonstrates increased expression of FGF-2 protein in NRCs infected with AdCsFGF-2. D: Western blot analysis demonstrating increased phosphorylation of ERK-1 and -2 after AdCAsFGF-2 infection of NRCs (top immunoblot). Total ERK-1 and -2 immunoblot of same membrane (bottom immunoblot) demonstrating presence of similar levels of unphosphorylated protein in all lanes. In this experiment, cells were maintained in serum-free media without the addition of rhFGF-2.

Article Snippet: After 24 hours (ie, 48 hours after infection), 10 ng/ml of recombinant human basic FGF (rhFGF-2; R&D Systems, Minneapolis, MN) was added to the serum-free media.

Techniques: Northern Blot, Western Blot, Expressing, Phospho-proteomics, Infection, Membrane