Lymphatic and hematopoietic markers are up-regulated in HEL cells on TPA treatment. (A) VEGFR-3 is up-regulated in HEL cells on TPA treatment. HEL cells were treated with TPA. The cells were harvested 0, 24, 48, or 72 hours after TPA stimulus and analyzed for VEGFR-3 expression using Western blot. Membranes were probed with HPRT Abs as a loading control. (B) Regulation of different hematopoietic markers in HEL cells on TPA treatment. HEL cells were cultured for 72 hours in the presence or absence of TPA. Lysates of the untreated, adherent (ad), and suspension (s) populations occurring after TPA treatment were analyzed using Western blot. Membranes were probed with HPRT Abs as a loading control. (C) Regulation of megakaryocytic transcription factors in HEL cells on TPA treatment. HEL cells were incubated with or without TPA for 72 hours. RNA was isolated from untreated, adherent, and suspension populations of treated HEL cells and transcribed into cDNA. Expression of several transcription factors involved in megakaryopoiesis was assessed using semiquantitative PCR. HPRT served as a loading control. (D) Regulation of lymphatic markers in HEL cells on TPA treatment. HEL cells were incubated with TPA for 72 hours or were left untreated. RNA was isolated from untreated, adherent, and suspension populations of treated HEL cells and then transcribed into cDNA, which served as a template for semiquantitative PCR. Human LECs were used as a positive control. Amplification of HPRT served to demonstrate equal loading.