| Considerable studies and clinical evidences prove: many Chinese medicines have direct efficacy at treating cancers. According to the theory of traditional Chinese medicine and based clinical practices, JinAn injection extracted from astragalus membranaceus, sophora flavescens and Ligusticum chuanxiong and so oa This formula displayed the great inhibition of growth of lung cancer in the previous experiment, and its constituents can induce apoptosis of cancer. Thus, we examine here whether JinAn and JinAn in combination with cisplatin can induce apoptosis in Lung cancer in vivo and in vitro, moreover, their molecular mechanisms were studied by morphological and biochemical procedures.This study consists of four parts: 1. To study of morphology of JinAn-induced apoptosis in Lewis lung cancer in mice; 2. The change of morphology of JinAn-induced apoptosis in both PC and PAa lung cancer cell lines in vitro; 3. The mechanism of JinAn-induced apoptosis in Lewis lung cancer; 4. To investigate the molecular mechanism of JinAn-induced apoptosis in both PG and PAa lung cancer cell lines in vitro.In experimental research, male CsBL/6J mice, weighted from 16 to 20 grams, were divided randomly into six groups: normal saline solution (NS), JinAn higher dose (JAH), JinAn middle dose (JAM), JinAn lower dose (JAL), cisplatin (DDP) and JinAn middle dose in combination with cisplatin (DJ). The cultured PG and PAa cell were divided into four groups: control (0, cisplatin (DDP), JinAn (JA) and JinAn in combination with cisplatin (DJ), respectively.Double fluorescence staining, TUNEL, DNA Ladder assay and laser confocal scanning microscope (LCSM) as well as transmission electron microscope (IRQ were used to determine the changes of induction of apoptosis after JA and DJ treatment on lung cancer in vivo and in vitro. Moreover, Annexin V and PI staining were applied to detect the changes of apoptotic rate at early and late stage of apoptosis process and alteration of cell cycle phase, in addition, mitochondrial membrane potential, intracellular calcium ion and pH value were also evaluated by flow cytometry. Furthermore, we examined the several key elements in apoptosis signaling cascades pathways, such as Fas/FasL, caspase-3 and bcl-2/bax as wsll as p53 by immunohistochemistry and the expression of bcl-2/bax were detected by in situ hybridization and RT-PCR.The main results displayed as follow:1. Marked inhibition of the growth of lung cancer after JinAn and JinAn in combination with cisplatin treatment, correlated with induction of apoptosis and arresting the cell cycle phase.After injection of JinAn of 150mg/kg, 100mg/kg and 50mg/kg respectively in mice, the inhibitory rate was 45.79%, 40.90% and 32.48%, these results showed that JinAn significantly inhibited the growth of lung cancer, and JinAn also increase the body weight. Marked nuclear condensation and the fragmentation of chromatin and pyknosis as well as apoptotic body were observed by microscope, TEM, fluorescent microscope and LCSM. Furthermore, DNA fragmentation was visualized in JAH group by electrophoresis. The rates of apoptosis in JinAn different dose group were 24.19%, 14.95% and 13.93% by flow cytometry, respectively. Using TUNEL assay apoptotic index (AI) were JAH: 0. 43 ± 5. 62, JAM: 4. 00±7.15, JAL; 1.14 ± 1. 90. The DNA content analysis implicated the percentage of S phase cell increased, but Gz/M phase decreased in JinAn each group.JinAn could retard the proliferation of PG and PAa cell dramatically. The typical apoptosis morphology was identified under fluorescent microscope, LCSM as well as TM Induction of a S phase arrest and decrease of Gi/M after JinAn treatment on PG while the majority of PAa cells showed a block in GQ/GI by flow cytometry. Results from Annexin V assay suggested the apoptotic rate of DJ was higher than DDP or JA at early and late stage of apoptosis process in PG cell. In contrast, lower in PAa cell of each group. The DNA ladder was seen in every group of PG, especially in DJ group but no one in PAa cell. AI of PG and PAa...
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