Induction of apoptosis by the extract from Stephania venosa (Bl.) spreng tuber on lymphocyte

Abstract

There are thousands of herbs in Thailand used in traditional medicine that are claimed to possess anticancer activity, but most of them have no scientific evidences to support their alleged medicinal quality. One among them is Stephania venosa (Bl.) spreng or S.venosa. The tuber of S.venosa has been used in many Thai traditional medicines as remedy of various diseases including cancer. In this study, we aim to apply cell culture technique and design a scientific model for testing and proving the claimed anticancer activity of these Thai herbs. Lymphocytes obtained from healthy blood donors were cultured in RPMI medium with the density of 4x10[superscript 5] cells/ml and used in various tests throughout the study. The inhibitory concentration at 50% (IC[subscript 50]) of water extract of S.venosa was determined by exposing 4x10[superscript 5] lymphocytes to various concentrations of S.venosa for 48 hours, and trypan blue dye exclusion was the technique applied to detect the cytotoxic activity. Detection of pH change and the cytotoxic activity of aliquots of S.venosa were studied for a period of 12 weeks to monitor the stability of the water extract of the herb. Besides the study of cytotoxic activity, IC[subscript 50] of water extract at 300 microgram/ml was selected for the test of apoptotic activity, both in normal human lymphocytes and lymphocytes obtained from cervical cancer patients. Both sources of lymphocytes were exposed to various treatments including: 100 microgram/ml S.venosa, 300 microgramm/ml S.venosa, 0.5 Gy [superscript 60]Co irradiation and a combination of 300 microgram/ml S.venosa plus 0.5 Gy [superscript 60]Co irradiation. The apoptotic activity was detected at 48 hour after exposure by in situ terminal deoxynucleotidyl transferase assay (TdT assay). Furthermore, inhibitory effect of various concentrations of water extract of S.venosa was tested on PHA stimulated normal human lymphocytes. Results from all studies indicated that, water extract of S.venosa tuber possessed cytotoxicity and apoptotic activities against both normal lymphocytes and lymphocytes obtained from cervical cancer patients. A combination of 0.5 Gy [superscript 60]Co irradiation with 300 microgram/ml S.venosa exhibited additive effect when detected by TdT assay. Besides, water extract of S.venosa exhibited definite inhibitory activity on PHA stimulated normal human lymphocytes at IC[subscript 50] of 40 microgram/ml. With the above data, it can then be primarily concluded that water extract of S.venosa should be further investigated and developed in line of other anticancer herbal drugs.