Effects of Andrographolide on Intracellular pH Regulation, Cellular Migration, and Apoptosis in Human Cervical Cancer Cells (Running Tittle: Effects of Andrographolide on pH Regulators and Apoptosis in Cervical Cancer)

Cancer cells have been characterized with alkaline intracellular pH (pH(i)) values (≥7.2) to enable cancer proliferation, migration, and progression. The aim of the present study was to explore the concentration-dependent effects of Andrographolide, an active diterpenoid compound of herb Andrographis paniculata, on Na(+)/H(+) exchanger isoform 1 (NHE1), cellular migration and apoptosis in human cervical cancer cells (HeLa). The pH(i) was detected by microspectrofluorometry method, and intracellular acidification was induced by NH(4)Cl prepulse technique. Viability and protein expression were determined by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and Western blot, respectively. Human normal endocervical cells (End1), ectocervical cells (Ect1), and HeLa were bought commercially. The resting pH(i) value of HeLa (≈7.47) was significantly higher than that of End1 and Ect1 (≈7.30), and shifted from alkaline to acidic following acid/base impacts. In HEPES (4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic acid | N-(2-Hydroxyethyl)piperazine-N′-(2-ethanesulfonic acid) -buffered superfusate, NHE1 and V-ATPase co-existed functionally for acid extrusion in HeLa, while only NHE1 existed functionally in End/Ect1. Andrographolide (3–1000 μM) concentration-dependently inhibited NHE1 activity. Cell-migration and expressions of NHE1, V-ATPase, PARP (poly-ADP-ribose-polymerase), pro-Caspase-3, and Bcl-2 were significantly reduced by pretreating with Andrographolide (≥100 μM) for 24–48 h in HeLa. Andrographolide inhibited cell viability of End1-cells/Ect1 and HeLa (≥100 and ≥30 μM, respectively). The present findings implicate the promising clinical applications of Andrographolide on cervical cancer treatment.