ABSTRACT
<p><b>OBJECTIVE</b>To explore the apoptosis mechanism of Wenxia Changfu Formula (, WCF) in reversing drug resistance of lung cancer in vivo.</p><p><b>METHODS</b>Thirty model mice were randomly assigned to three groups: control group, cisplatin (CDDP) group, and WCF group. A transplanted tumor model of lung adenocarcinoma was established in all groups. Mice in the WCF group received intragastric administration of WCF (0.2 mL/10 g body weight) everyday in addition to CDDP intraperitoneally (5 mg/kg body weight) twice a week. The mice in the CDDP group received CDDP intraperitoneally (5 mg/kg body weight) twice a week, while the control group received normal saline intraperitoneally (0.2 mL/10 g body weight) everyday. The weight of the nude mice and respective tumors, tumor volume and tumor-inhibiting rate were measured. Electron microscopy was used to observe the existence of apoptosis body. Apoptosis index (AI) was detected by TdT-mediated dUTP nick end labeling staining. The expression of Fas and FasL mRNA was investigated by reverse transcription polymerase chain reaction, while immunohistochemistry was applied to detect the protein expression of Fas and FasL, caspase-3 and caspase-activated DNase (CAD), respectively.</p><p><b>RESULTS</b>Compared with CDDP group and control group, WCF could significantly reduce the tumor volume from the 19th day and alleviate the tumor weight (P <0.05), and the apoptosis body was found in tumor cells in the WCF group. WCF could also enhance the level of AI, up-regulate the expression of caspase apoptosis pathway related protein caspase-3 and CAD, as well as the expression of Fas, FasL mRNA and protein (P <0.05).</p><p><b>CONCLUSION</b>WCF could improve the sensitivity of tumor cells to CDDP and reverse the drug resistance by inducing the apoptosis.</p>
Subject(s)
Animals , Female , Humans , Adenocarcinoma , Drug Therapy , Pathology , Apoptosis , Caspase 3 , Metabolism , Cell Line, Tumor , Chromatography, High Pressure Liquid , Cisplatin , Pharmacology , Drug Resistance, Neoplasm , Drugs, Chinese Herbal , Pharmacology , Therapeutic Uses , Fas Ligand Protein , Genetics , Metabolism , Fluorescent Antibody Technique , In Situ Nick-End Labeling , Lung Neoplasms , Drug Therapy , Pathology , Mice, Nude , RNA, Messenger , Genetics , Metabolism , Tumor Burden , Xenograft Model Antitumor Assays , fas Receptor , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To observe the effect of the combination of Wenxia Changfu Formula ([see text], WCF) with cisplatin (CDDP) on inhibiting non-small cell lung cancer (NSCLC) in vitro and In Vivo and explore its mechanism from its effect on cell cycle.</p><p><b>METHODS</b>In vitro, WCF-containing serum was prepared and the rhubarb b1, emodin, and aconitine were detected qualitatively by high-performance liquid chromatogram (HPLC). A549 cell lines were treated with blank control (dimethyl sulfoxide), normal serum, normal serum with CDDP (1.25, 2.5, and 5.0 μg/mL, respectively), WCF-containing serum plus different doses of CDDP (1.25, 2.5, and 5.0 μg/mL, respectively). The inhibitory effect was detected by 3-(4,5)-dimethylthiazo(-zy1)-3,5-diphenylterazolium bromide (MTT). The cell cycle was detected by flow cytometry. The protein and mRNA expressions of cyclin D1, proliferating cell nuclear antigen (PCNA), retinoblastoma (Rb), and p16 were observed with immunofluorescence and RT-PCR, respectively. In Vivo, nude mice xenograft model was established and grouped into the control, CDDP, WCF, and combination groups. The combination's inhibition of tumor growth and influence on the weight, spleen, and thymus gland were observed.</p><p><b>RESULTS</b>The inhibitory rate of the combination against A549 cell lines excelled the CDDP alone significantly (P <0.05); the combination showed a synergism inhibitory effect (Q=1.19). Compared with the monotherapy, the combination increased the cell percentage in G(0)/G(1) phase and decreased the cell percentage in S phase significantly (P <0.05); the protein and mRNA expressions of cyclin D1, PCNA, and Rb were significantly reduced; the protein and mRNA expressions of p16 were significantly enhanced. Compared with the monotherapy, the combination inhibited the tumor growth significantly In Vivo and reduced the weight of tumor (P <0.05); compared with the CDDP group, the spleen and thymus gland index of the combination group were enhanced significantly (P <0.05).</p><p><b>CONCLUSIONS</b>The combination of WCF with CDDP significantly inhibited the A549 cell lines proliferation in vitro and the growth of the tumor In Vivo; it inhibited effectively the atrophy of the immune organ caused by chemotherapy. The combination inhibited overproliferation of A549 cell lines by arresting the G(0) /G(1) phase of cell cycle and affecting the protein and mRNA expressions of cell cycle-related proteins, cyclin D1, etc.</p>