Biology Reference
In-Depth Information
supernatants of conditioned media was verified by Western blot (Figure 1B) and
the function of sFlt-1 was validated by endothelial cell growth inhibition assay in
vitro. The concentrated conditioned media obtained from BMSCs infected with
Adv-GFP-sFlt-1 or with Adv-GFP were applied to the mouse endothelial cells
grown in 24-well plates, followed by stimulation with a 10 ng/ml of VEGF 15
minutes later. Conditioned media from BMSCs infected with Adv-GFP-sFlt-1
resulted in inhibition of endothelial cells proliferation by about 50% compared
with that from BMSCs infected with Adv-GFP or uninfected BMSCs (Figure
1C, P < 0.05), indicating that the BMSCs infected with Adv-GFP-sFlt-1 could
effectively express and secret sFlt-1.
Figure 1.
BMSCs infection and verification of secreted sFlt-1. After the BMSCs were confirmed by flow
cytometric analysis, BMSCs were grown to 90% confluence and infected with adenoviruses at a multiple of
infection of 3000 for 2 hours. 24 hours later, BMSCs were harvested and ready for use. Fluorescence microscope
showed up to 100% GFP-positive cells. (Figure 1-A). Supernatants deposits from sFlt-1 bearing BMSCs could
be recognized by antibodies reactive to NH2 terminus of mouse Flt-1, but negative staining in supernatants
deposits from control BMSCs in Western blot analysis. (Figure 1-B). Conditioned media from Adv-sFlt-1
infected BMSCs was shown to inhibit the VEGF-driven mouse endothelial cells proliferation by about 50%
compared with controls (Figure 1-C, P < 0.05).
Anti-Metastasis effect
The mouse CT26 and LLC metastasis models were used to determine whether
BMSCs expressing sFlt-1 suppress the growth of metastatic neoplasm. The meta-
static nodules > 3 mm were assessed as an indicator of angiogenesis because the
growth of tumors > 3 mm is thought to be vasculature sensititive [19]. Most of the
mice systemically administered with control BMSCs or NaCl solution developed
Search WWH ::
Custom Search