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Blood, Vol. 113, Issue 5, 1129-1138, January 29, 2009
Placenta growth factor induces 5-lipoxygenase–activating protein to increase leukotriene formation in sickle cell disease
Blood Patel et al.
113: 1129
Supplemental materials for: Patel et al
Files in this Data Supplement:
- Figure S1. Dose dependent effect of PlGF on FLAP mRNA expression in THP-1 cells (JPG, 24.6 KB)
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THP-1 cells (3 × 106 cells/2ml) were incubated in serum free RPMI-1640 medium overnight followed by treatment with indicated concentration of recombinant PlGF (25–250 ng/mL) for 24h. Total RNA (5 µg) was subjected to RPA analysis, as described in Materials and Methods section. GAPDH shows equal loading. Results: As shown in Fig. S1, PlGF showed increase in FLAP mRNA expression in a dose dependent manner (25–500 ng/mL). The optimal increase in FLAP expression occurred at 250 ng/mL of PlGF (3.3-fold), though the increase can be seen at concentration as low as 25 (1.6-fold) and 50 ng/mL (2.2-fold).
- Figure S2. Time course of PlGF mediated FLAP mRNA expression in THP-1 cells (JPG, 21.9 KB)
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THP-1 cells (3 × 106 cells/2ml) were incubated in serum free RPMI-1640 overnight followed by treatment with PlGF (250 ng/mL) for the indicated time points (2–24 hr). Total RNA (5 µg) was subjected to RPA analysis, as described in Materials and Methods section. GAPDH shows equal loading. Results: Fig. S2 shows that there was a time dependent increase in the FLAP mRNA expression in response to PlGF treatment. FLAP mRNA did not change at 2 and 4 hr. However, FLAP mRNA expression showed an increase at 8 hr (1.5-fold) with maximal increase at 24 hr (3.6-fold).
- Figure S3. Effect of PlGF on LTE4 release at early time points in THP-1 cells (JPG, 28.9 KB)
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THP-1 cells (1.5 × 106 cells/ml) were incubated in serum free RPMI-1640 medium overnight followed by treatment with PlGF (250 ng/mL) for indicated time periods (5–120 min). The culture supernatants were collected and 100 µL was used to analyze LTE4 release by ELISA. The control values were considered 100%. Results: As shown in Fig. S3, PlGF stimulation of THP-1 cells at early time points (5–120 min) did not show any significant change in LTE4 release when compared to untreated control cells.
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