Phlegm Side Containing Serum Human Lung Cancer Cells To Apoptosis Signaling

Objective:Hua Tan San lie Fang is composed of semen strychni, panax notoginseng saponins,rhizoma pinelliae,rhizoma arisaematis,e/ al.lt is brought forth by my tutor -professor Yang Qinjian, instructed by the theory of Bian Zheng Lun Zhi, according to the main pathogentic and pathological nature of bronchogenic carcinoma-"mutual massing of phlegm and blood stasis" and long observation of experiment and clinic on bronchogenic carcinoma. It has shown good effects on the treatment of bronchogenic carcinoma. It has proved that the pharmaceutical serum containing Hua Tan San lie Fang has function of inducing the apoptosis of lung cancer cells. In order to clarify the antineoplastic mechanism of Hua Tan San lie Fang, we applies the experimental method of "serological pharmacology" combined with the theory and laboratory technique of modern cytobiology and molecular biology in studying comprehensively the apoptosis induced by pharmaceutical serum containing Hua Tan San lie Fang on the cell molecular level. Methods:(1)Ten New Zealand rabbits were divided into two groups (drug group and control group) at random . Hua Tan San lie Fang were administered po to rabbits in drug group at the dose of 7g/kg every day. Saline water were administered po to rabbits in control group at the dose of 3.5ml/kg. Ten days later , collecting blood through carotid artery, taking serum apart , freezing and storing in low temperature. (2)The rabbit serum of drug group was tested by HPLC through representative ingredient horseflesh(It contains strychnin). (3)SPC-A1 cells were inbucated with the pharmaceutical serum containing Hua Tan San lie Fang and rabbit serum not containing drug separately for 12h,24h,36h,48h,then collecting cells .suspending cells in acetic acid buffer, freezing and then melting for three times. (4)PC-A1 cells were inbucated with the pharmaceutical serum containing Hua Tan San Jie Fang and rabbit serum not containing drug separately for 12h,24h,36h,48h, then the expression of Fas and FasL were tested by immunechemical method. (5)SPC-A1 cells were inbucated with the pharmaceutical serum containing Hua Tan San Jie Fang and rabbit serum not containing drug separately for 12h,24h,36h,48h,then the expression of TNFR1 mRNA was examined by in situ hybridzation method. Results:(1)By HPLC, the retention time of strychnin standard and the pharmaceutical serum containing Hua Tan San Jie Fang was similar. (2)The cAMP concentration of 12h and 24h were lower than that of 36h and 48h,but there were no significantdifference between the cGMP concentration of 12h,24h and that of 36h,48h. So the ratio of cAMP/cGMP in SPC-A1 cells increased.(3)The expression of Fas in group of pharmaceutical serum containing Hua Tan San lie Fang was higher than the one of control group(P < 0.05), but the expression of FasL in group of pharmaceutical serum containing Hua Tan San lie Fang was lower than everyone of control group(P < 0.01). (4)The expression of TNFR1 mRNA in group of pharmaceutical serum containing Hua Tan San Jie Fang was higher than the one of control group( P < 0.05). Conclusions:The rabbit serum of drug group contains some component of Hua Tan San Jie Fang .The pharmaceutical serum containing Hua Tan San Jie Fang has the function of inducing the apoptosis of lung cancer cells. Its possible mechanism is : (1)the increasing of cAMP concentration and the ratio of cAMP/cGMP in SPC-A1 cells; (2)the up-regulating of expression of Fas and the down-regulating of expression of FasL in SPC-A1 cells; (3)the up-regulating of expression of TNFR1 SPC-A1 cells.