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Method Article
Construction of Lung Adenocarcinoma Xenograft Model in Mice and Evaluation of the Efficiency of XY Decoction through MAPK Pathway
Neste Artigo
Resumo
This study demonstrates that XY decoction exerts therapeutic effects against lung adenocarcinoma by regulating the MAPK pathway.
Resumo
Lung adenocarcinoma is increasing worldwide, prompting the exploration and development of effective treatment methods. In recent years, the therapeutic potential of traditional Chinese medicine for lung adenocarcinoma, such as the XiaoYi (XY) decoction, has been increasingly recognized. The mechanism of action of traditional Chinese medicine in treating lung adenocarcinoma continues to be elucidated. In this study, animal models were used to investigate the therapeutic effects of XY decoction in the treatment of lung adenocarcinoma. The blood-entry chemical components of XY decoction in mice were separated and analyzed in both positive and negative ion modes using UHPLC-QE-MS. The findings were then validated through in vivo experiments, which demonstrated reduced tumor size, improved pathological changes, and increased apoptosis. As a result, fifteen components absorbed into the blood following XY decoction administration were identified: Stachyose, Quercetin-3-O-beta-glucopyranosyl-7-O-alpha-rhamnopyranoside; Imperialine; Spinosin B; Peimine; Flavone base + 3O, 2MeO, O-Hex; Picropodophyllotoxin; (2S,3R,4S,5S,6R)-2-[(2R,3R,4S,5S,6R)-4,5-dihydroxy-2 [[(3S,5R,8R,10R,12R,13R,14R,17S)-12-hydroxy-17-(2-hydroxy-6-methylhept-5-en-2-yl) 4,4,8,10,14-pentamethyl-2,3,5,6,7,9,11,12,13,15,16,17-dodecahydro-1H cyclopenta[a]phenanthren-3-yl]oxy]-6-(hydroxymethyl)oxan-3-yl]oxy-6-(hydroxymethyl)oxane-3,4,5-triol; Tricin; Ginsenoside Rg1; (20R)-Ginsenoside Rh1; Jujuboside B; Aucubin; Ginsenoside Rg3; and beta-Elemonic acid. Animal experiments demonstrated that XY decoction inhibited tumor growth, improved pathological changes, and promoted tumor cell apoptosis in a dose-dependent manner. The mechanisms involved may be related to the upregulation of p-JNK and p-P38 protein expression in the MAPK pathway and the downregulation of p-ERK protein expression in the MAPK pathway. In conclusion, XY decoction has the potential to regulate the MAPK pathway and exert therapeutic effects against lung adenocarcinoma.
Introdução
Lung cancer incidence is rising worldwide. It accounts for 18% of all malignancies and is responsible for a significant number of cancer-related deaths1,2. Non-small cell lung cancer (NSCLC) is a prevalent form of lung cancer, with adenocarcinoma as the main pathological type3. Due to its insidious symptoms, NSCLC is often diagnosed at an advanced stage and has a high mortality rate4,5. Stage IV NSCLC patients have a five-year survival rate of only 5%6, while even surgically resectable early-stage NSCLC h
Protocolo
All animal procedures were approved by the Animal Ethics Committee of Changchun University of Traditional Chinese Medicine (Ethics No. 2024008). Six-week-old C57BL/6 male mice (20 ± 2 g), bred under specific pathogen-free (SPF) conditions, exhibited typical physical activity patterns in a controlled environment with constant temperature and humidity. They had ad libitum access to food and water and were maintained under a 12-h light/dark cycle. Details of the reagents and equipment used in this study are provided in the Table of Materials.
1. Preparation of XY decoction aqueous extract
- Place Pa
Resultados
XY powder solution and serum samples from the control group and the XY group were analyzed using UHPLC-QE-MS, revealing well-separated peaks. The total ionization diagrams for the positive and negative ion modes are shown in Figure 3. The composition of the XY decoction was identified based on retention time, acquisition mode, and mass spectrum fragments. Fifteen blood-entry components were identified by comparing the serum of the blank and XY decoction oral ...
Discussão
The incidence of cancer is rising annually, attracting significant clinical and research interest22,23. Recent studies have demonstrated the effectiveness of traditional Chinese medicine (TCM) in treating lung cancer24,25,26. For instance, Maimendong decoction has been shown to enhance the number and activity of NK cells, thereby inhibiting lung cancer metastasis
Divulgações
The authors have nothing to disclose.
Agradecimentos
This work was supported by the Jilin Province Science and Technology Development Plan Item (YDZJ202301ZYTS459).
Materiais
| Name | Company | Catalog Number | Comments |
| Acetonitrile | CNW Technologies | 5/8/1975 | |
| 2-Chloro-L phenylalanine | Shanghai Hengbo Biotechnology Co., Ltd. | 103616-89-3 | |
| Ammonium Hydroxide Solution | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | #2028 | |
| Antibody-β-actin | Abcam | ab8228 | |
| Anti-Erk1 (pT202/pY204) + Erk2 (pT185/pY187) Antibody | Abcam | ab4819 | |
| Anti-ERK1/ERK2 Antibody | Abcam | ab184699 | |
| Anti-JNK1 (pY185) + JNK2 (pY185) + JNK3 (pY223) Antibody | Abcam | ab76572 | |
| Anti-JNK1/JNK2/JNK3 Antibody | Abcam | ab208035 | |
| Anti-Ki67 Antibody | Cell Signaling Technology | GB121141 | |
| Anti-p38 (phospho T180) Antibody | Abcam | ab178867 | |
| Anti-p38 alpha/MAPK14 Antibody | Abcam | ab170099 | |
| BCA protein concentration assay kit | Boster Biological Technology | 17E17B46 | |
| Bovine Serum Albumin (BSA) | DAKO | BIOFROXX | |
| Broad-spectrum phosphatase inhibitor (100x) | Boster Biological Technology | AR1183 | |
| C57bl/6 mice | Liaoning Changsheng Biotechnology Co., Ltd. | SCXK2021-0006 | |
| Chromatographic column | Waters | ACQUITY UPLC BEH C18 1.7 μm2.1*100 mm | |
| Cisplatin | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | 232120 | |
| Citrate (pH 6.0) Antigen Retrieval Solution | Wuhan Baiqiandu Biotechnology Co., Ltd. | B2010 | |
| Cover Slip | Jiangsu Shitai Laboratory Equipment Co., Ltd. | 10212432C | |
| Cryogenic Stage | Wuhan Junjie Electronics Co., Ltd. | JB-L5 | |
| DAB Chromogenic Kit | Jiangsu Shitai Laboratory Equipment Co., Ltd. | 2005289 | |
| Decolorization shaker | Beijing Liu Yi Instrument Factory | WD-9405A | |
| Dehydrator | Wuhan Junjie Electronics Co., Ltd. | JJ-12J | |
| Drying Oven | Shanghai Huitai Instrument Manufacturing Co., Ltd. | DHG-9140A | |
| Dulbecco's modified eagle medium | Gibco | 8121587 | |
| EDTA (pH 8.0) Antigen Retrieval Solution | Wuhan Baiqiandu Biotechnology Co., Ltd. | B2001 | |
| EDTA (pH 9.0) Antigen Retrieval Solution | Wuhan Baiqiandu Biotechnology Co., Ltd. | B2002 | |
| Electrophoresis buffer | NCM Biotech | 20230801 | |
| Embedding machine | Wuhan Junjie Electronics Co., Ltd. | JB-P5 | |
| Eosin | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | 212954 | |
| Ethanol | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | 493511 | |
| Fetal bovine serum | Gibco | 2166090RP | |
| Formic acid | SIGMA | 64-18-6 | |
| GraphPad Prism software | GraphPad software, LLC | Version 9.0.0 | |
| Hematoxylin | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | H3136 | |
| High performance liquid chromatography | Agilent | 1260 Infinity II Prime | |
| High speed refrigerated centrifuge | Thermo Fisher Scientific | Legend Micro 17R | |
| High-Resolution Mass Spectrometry | Thermo Fisher Scientific | Q Exactive Focus | |
| HRP-conjugated Goat Anti-Mouse | SeraCare | 5220-0341 | |
| HRP-conjugated Goat Anti-Rabbit | SeraCare | 5220-0336 | |
| HRP-conjugated Goat Anti-Rabbit/Mouse Universal Secondary Antibody | DAKO | K5007 | |
| HRP-conjugated Goat Anti-Rat | SeraCare | 5220-0364 | |
| HRP-conjugated Rabbit Anti-Goat | SeraCare | 5220-0362 | |
| Hydrochloric acid | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | H9892 | |
| Hydrogen Peroxide | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | B12555 | |
| Hypersensitive ECL chemiluminescence kit | NCM Biotech | P10300 | |
| ImageJ software | National Institutes of Health | v1.8.0 | |
| Imaging System | Nikon (Japan) | NIKON DS-U3 | |
| Lewis lung carcinoma cells | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | 90020104 | |
| Mesh cloth | Baijie | 100110779650 | |
| Metal heating temperature control instrument | Baiwan Electronictechnology Co., Ltd. | HG221-X3 | |
| Methanol | Aladdin | M116118 | |
| Micropipette | Dlab Scientific Co., Ltd. | KE0003087/KA0056573 | |
| Microscope Slide | Jiangsu Shitai Laboratory Equipment Co., Ltd. | 10212432C | |
| Microtome | Shanghai Leica Instruments Co., Ltd. | RM2016 | |
| Microwave Oven | Galanz Microwave Electrical Appliances Co., Ltd. | P70D20TL-P4 | |
| Neutral Balsam | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | 10004160 | |
| Optical microscope | Nikon (Japan) | Nikon Eclipse CI | |
| Oven | Shanghai Huitai Instrument Manufacturing Co., Ltd. | DHG-9140A | |
| PAGE gel rapid preparation kit | Biosharp | PL566B-5 | |
| Paraformaldehyde | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | 8.18715 | |
| Phosphate buffered saline (1x) | Gibco | 8120485 | |
| Pre-stained color protein marker (10-180 kDa) | Cywin Innovation (Beijing) Biotechnology Co., Ltd. | SW176-02 | |
| Protein loading buffer (5X) | Boster Biological Technology | AR1112 | |
| pulverizer | Shangzhiqiao Co., Ltd. | DFT-100A | |
| PVDF (0.45 μm) | Cywin Innovation (Beijing) Biotechnology Co., Ltd. | SW120-01 | |
| Rapid membrane transfer solution | Cywin Innovation (Beijing) Biotechnology Co., Ltd. | SW171-02 | |
| RIPA lysate | Boster Biological Technology Co., Ltd. | AR0105 | |
| TBS buffer | NCM Biotech | 23HA0102 | |
| Tissue Flotation Workstation | Jinhua Kedi Instrument Equipment Co., Ltd., Zhejiang Province. | KD-P | |
| Transmembrane buffer | NCM Biotech | 23CA2000 | |
| Trypsin (0.25%, 1x) | HyClone | J210045 | |
| Ultrasonic cell disruption apparatus | Ningbo Xinyi ultrasonic equipment Co., Ltd. | JY92-IIDN | |
| Upright Optical Microscope | Nikon (Japan) | Nikon Eclipse CI | |
| vacuum lyophilizer | Lanjing Co., Ltd. | HD-LG20D | |
| Vortex mixer | Kylin-Bell | XW-80A | |
| Xylene | China National Pharmaceutical Group Chemical Reagent Co., Ltd. | 214736 |
Referências
- Ferlay, J. et al. Cancer statistics for the year 2020: An overview. Int J Cancer. 149 (4), 778-789 (2021).
- Sung, H. et al. Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 71 (3), 209-249 (2021).
- Ahn, M. J. et al. Asian Thoracic Oncology Research Group (ATORG) expert consensus statement on MET alterations in NSCLC: Diagnostic and therapeutic considerations. Clin Lung Cancer. 23 (8), 670-685 (2022).
- Seguin, L., Durandy, M., Feral, C. C. Lung adenocarcinoma tumor origin: A guide for persona
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