Effects of alpha-adrenoreceptor antagonists on apoptosis and proliferation of pancreatic cancer cells in vitro

Abstract

Aim: To discuss the expression of alpha-adrenoreceptors in pancreatic cancer cell lines PC-2 and PC-3 and the effects of alpha1- and alpha2-adrenoreceptor antagonists, yohimbine and urapidil hydrochloride, on the cell lines in vitro.

Methods: We cultured the human ductal pancreatic adenocarcinoma cell lines PC-2 and PC-3 and analyzed the mRNA expression of alpha1- and alpha2-adrenergic receptors by reverse transcription polymerase chain reaction (RT-PCR). The effects of yohimbine and urapidil hydrochloride on cell proliferation were assessed by 3-(4,5-dimethylthiasol-2-yl)-2,4,-diphenyltetrazolium bromide (MTT) assay. Apoptosis was detected using the terminal deoxyribonucleotidyl transferase (TdT)-mediated biotin-16-dUTP nick-end labeling (TUNEL) assay and flow cytometry (FCM).

Results: PC-2 expressed mRNA in alpha1- and alpha2-adrenoreceptors. MTT assays showed that urapidil hydrochloride had no effect on PC-3 cell lines. However, exposure to urapidil hydrochloride increased DNA synthesis in PC-2 cell lines as compared to the control group. PC-2 cell lines were sensitive to both drugs. The proliferation of the 2 cell lines was inhibited by yohimbine. Cell proliferation was inhibited by yohimbine via apoptosis induction.

Conclusion: The expression of alpha1- and alpha2-adrenoreceptors is different in PC-2 and PC-3 cell lines, which might be indicative of their different functions. The alpha2-adrenoceptor antagonist, yohimbine, can inhibit the proliferation of both cell lines and induce their apoptosis, suggesting that yohimbine can be used as an anticancer drug for apoptosis of PC-2 and PC-3 cells.

Figure 1

Expression of mRNA in adrenoceptors of human ductal pancreatic adenocarcinoma cell lines PC-2 and PC-3 by RT-PCR. Lanes 1-8 represent the α_1D-, α_1A-, α_2A-, α_2B-, α_2C-, α_1B-, β₁-, and β₂-adrenoceptor subtypes, respectively. Lane maker = 100 bp DNA ladder. PC-2 expressed mRNA in α_1D-, α_1A-, α_2A-, β₁-, and β₂-adrenoceptors and PC-3 expressed mRNA in α_2B-, β₁-, and β₂-adrenoceptors.

Figure 2

MTT assay shows that PC-2 cells treated with urapidil hydrochloride for 24 h could increase cellular proliferation in a concentration-dependent manner. Yohimbine inhibited the proliferation of PC-2 cells (A) and urapidil hydrochloride had no effect on PC-3 cell lines while the proliferation of PC-3 cell lines was inhibited by yohimbine (B).

Figure 3

TUNEL assay showing apoptotic PC-2 and PC-3 cells after treatment with 100 μmol/L urapidil hydrochloride and yohimbine for PC-2 cells and 100 μmol/L yohimibine and 100 μmol/L urapidil for PC-3 cells compared with control group.

Figure 4

Apoptotic PC-2 and PC-3 cells after treatment with 200 μmol/L urapidil hydrochloride (A) and 200 μmol/L yohimbine (B) for PC-2 cells and 200 μmol/L yohimibine for PC-3 cells (C) detected by FCM. Significant apoptosis was induced by yohimbine on both PC-2 and PC-3 cells, whereas urapidil hydrochloride had no apoptotic effect on PC-2 cell.