Figure 1
Silencing growth hormone receptor (GHR) inhibited viability and anchorage-independent growth of pancreatic ductal adenocarcinoma (PDAC) cell lines. (a) GHR expression levels from PANC-1 HPAC and MIAPaCa-2 were compared with normal pancreatic cells using western blot. (b) Representative immunohistochemical analysis of GHR by tumor stage in pancreatic adenocarcinoma tissues and normal pancreas tissue. (c) Three predesigned GHR small interfering RNAs (siRNAs) (a–c) were transfected in PANC-1 and HPAC cells at 10 nM along with scrambled control siRNA (labeled as ‘S’), untreated control PANC-1 and HPAC cells (labeled as ‘C’) and mock control transfection reagent alone (labeled as ‘M’). GHR siRNA silencing efficacy was examined using western blot in PANC-1 and HPAC cells. (d) Effect of GHR knockdown on cell viability of PANC-1 and HPAC. PANC-1 and HPAC cells were transfected with 10 nM of GHR siRNA and scrambled siRNA. MTS assay kit was used to determine the cell viability of post transfected cells after 48 h. Error bars denote the s.d. between triplicates. (e) Loss of anchorage-independent growth in GHR silenced PDAC cells, PANC-1 and HPAC. Colony-forming abilities of GHR transfected PANC-1 and HPAC were determined using soft agar colony formation assay along with scrambled control. Transfected PANC-1 and HPAC cells were allowed to grow in 0.7% agarose in growth media supplemented 10% fetal bovine serum (FBS) for 16 and 28 days, respectively. Representative images of colony formation were shown from two independent experiments done in triplicate. (f) Percentage of colonies in transfected PANC-1 and HPAC cells were compared with scrambled control (SCR) serving as the baseline. Data presented as the mean±s.d. (n=3). *P<0.05.
