加味四君子汤对H22肝癌小鼠的抑瘤作用和免疫功能的影响
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摘要
目的探讨加味四君子汤的体内抑瘤活性及对肝肾毒性、免疫系统的影响。方法建立H22荷瘤小鼠模型,将荷瘤小鼠分为空白组、模型组、环磷酰胺组、加味四君子汤低、中、高剂量组。通过计算抑瘤率,观察肿瘤组织HE染色病理切片,免疫组化法检测Bax、Bcl-2、Bax/Bcl-2及caspase-3水平研究加味四君子汤的抑瘤活性及相关机制;通过肝肾生化指标观察加味四君子汤的肝肾毒性;通过计算荷瘤小鼠免疫器官指数、外周血检测血常规和酶联免疫法检测小鼠血清的IL-2、TNF-α的水平探讨加味四君子汤对H22荷瘤小鼠免疫功能的影响。结果环磷酰胺组、加味四君子汤中剂量、高剂量组的肿瘤质量均低于模型组(P<0.05),抑瘤活性差异均有统计学意义;其肿瘤细胞有不同程度的破裂坏死,免疫组化各实验组能有效上调Bax和caspase-3蛋白表达,下调Bcl-2蛋白表达,提高Bax/Bcl-2比值。同时,加味四君子汤降低了荷瘤小鼠的Cr、BUN、AST与ALT水平,提高了免疫器官指数和外周血白细胞、红细胞和血红蛋白含量,上调了TNF-α、IL-2水平,其中,高剂量组与模型组差异具有统计学意义(P<0.01)。结论加味四君子汤具有较好的体内抑瘤活性,能改善荷瘤小鼠的肝肾功能,其抑瘤作用可能与上调Bax、caspase-3表达、TNF-α和IL-2水平,下调Bcl-2蛋白表达以及增强小鼠的非特异性免疫功能有关。
Objective To investigate the antitumor activity of Jiawei Sijunzi decoction and study its liver and kidney toxicity and its effect on the immune system in a tumor-bearing mouse model.Methods Hepatoma H22 tumor-bearing mouse models were randomized into model group,cyclophosphamide(CTX) group,and low-,moderate-,and high-dose Jiawei Sijunzi decoction groups(JW-L,JW-M,and JW-H groups,respectively).The antitumor activity of Jiawei Sijunzi decoction was assessed by calculating the tumor inhibition rate and pathological observation of the tumor tissues.Immunohistochemistry was used to detect the expressions of Bax,Bcl-2,Bax/Bcl-2 and caspase-3 in the tumors.The liver and kidney toxicity of Jiawei Sijunzi decoction was analyzed by evaluating the biochemical indicators of liver and kidney functions.The immune function of the tumor-bearing mice were assessed by calculating the immune organ index,testing peripheral blood routines,and detection of serum IL-2 and TNF-α levels using enzyme-linked immunosorbent assay.Results Compared with that in the model group,the tumor mass in CTX,JW-M and JW-H groups were all significantly reduced(P<0.05) with cell rupture and necrosis in the tumors.Immunohistochemistry revealed obviously up-regulated expressions of Bax and caspase-3 and down-regulated expression of Bcl-2 protein with an increased Bax/Bcl-2 ratio in CTX,JW-M and JW-H groups.Treatment with Jiawei Sijunzi decoction significantly reduced Cr,BUN,AST and ALT levels,improved the immune organ index,increased peripheral blood leukocytes,erythrocytes and hemoglobin levels,and up-regulated the levels of TNF-α and IL-2 in the tumor-bearing mice.These changes were especially significant in JW-H group when compared with the parameters in the model group(P<0.01).Conclusion Jiawei Sijunzi decoction has a strong anti-tumor activity and can improve the liver and kidney functions of tumor-bearing mice.Its anti-tumor effect may be attributed to the up-regulation of Bax,caspase-3,TNF-α and IL-2 levels and the down-regulation of Bcl-2 expression as well as the enhancement of the non-specific immune function.
Objective To investigate the antitumor activity of Jiawei Sijunzi decoction and study its liver and kidney toxicity and its effect on the immune system in a tumor-bearing mouse model.Methods Hepatoma H22 tumor-bearing mouse models were randomized into model group,cyclophosphamide(CTX) group,and low-,moderate-,and high-dose Jiawei Sijunzi decoction groups(JW-L,JW-M,and JW-H groups,respectively).The antitumor activity of Jiawei Sijunzi decoction was assessed by calculating the tumor inhibition rate and pathological observation of the tumor tissues.Immunohistochemistry was used to detect the expressions of Bax,Bcl-2,Bax/Bcl-2 and caspase-3 in the tumors.The liver and kidney toxicity of Jiawei Sijunzi decoction was analyzed by evaluating the biochemical indicators of liver and kidney functions.The immune function of the tumor-bearing mice were assessed by calculating the immune organ index,testing peripheral blood routines,and detection of serum IL-2 and TNF-α levels using enzyme-linked immunosorbent assay.Results Compared with that in the model group,the tumor mass in CTX,JW-M and JW-H groups were all significantly reduced(P<0.05) with cell rupture and necrosis in the tumors.Immunohistochemistry revealed obviously up-regulated expressions of Bax and caspase-3 and down-regulated expression of Bcl-2 protein with an increased Bax/Bcl-2 ratio in CTX,JW-M and JW-H groups.Treatment with Jiawei Sijunzi decoction significantly reduced Cr,BUN,AST and ALT levels,improved the immune organ index,increased peripheral blood leukocytes,erythrocytes and hemoglobin levels,and up-regulated the levels of TNF-α and IL-2 in the tumor-bearing mice.These changes were especially significant in JW-H group when compared with the parameters in the model group(P<0.01).Conclusion Jiawei Sijunzi decoction has a strong anti-tumor activity and can improve the liver and kidney functions of tumor-bearing mice.Its anti-tumor effect may be attributed to the up-regulation of Bax,caspase-3,TNF-α and IL-2 levels and the down-regulation of Bcl-2 expression as well as the enhancement of the non-specific immune function.
引文
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[2]Schmid I,von Schweinitz D.Pediatric hepatocellular carcinoma:challenges and solutions[J].J Hepatocellul Carcinom,2017,4(10):15-21.
[3]Samonakis DN,Kouroumalis EA.Systemic treatment for hepatocellular carcinoma:Still unmet expectations[J].World JHepatol,2017,9(2):80-90.
[4]Jaferian S,Soleymaninejad M,Negahdari BA.Stem cell,biomaterials and growth factors therapy for hepatocellular carcinoma[J].Biomed Pharmacoth,2017,88(9):1046-53.
[5]Gong YL.Therapy response of Chinese herbal medicine in primary liver cancer[J].J Clin Oncol,2015,33(3,S):327-35.
[6]Qian J,Xie H,Guo CX,et al.Sijunzi Decoction Demolition Parties Inhibit Proliferation and Iinduce Apoptosis Human Gastric Cancer Bgc823 Side Population[J].African J Tradit Complement Alternat Med,2015,12(6):77-89.
[7]Jie Y,He W,Yang X,et al.Kruppel-like factor 4 acts as a potential therapeutic target of Sijunzi decoction for treatment of colorectal cancer[J].Cancer Gene Ther,2017,24(9):361-6.
[8]施胜英,林海桢,周溦,等.加味四君子汤含药血清对肝癌Hep-G2细胞的影响[J].中国实验方剂学杂志,2016,22(18):88-93.
[9]Liu XN,Zhu XX.Stellera chamaejasme L.extract induces apoptosis of human lung cancer cells via activation of the death receptor-dependent pathway[J].Exp Ther Med,2012,4(4):605-10.
[10]Gan D,Xu A,Du H,et al.Chinese classical formula sijunzi decoction and chronic atrophic gastritis:evidence for treatment approach[J].Evidence-Based Complement Alternat Med,2017,92(9):1022-34.
[11]Jia J,Qin Y,Zhang L,et al.Sijunzi decoction-treated rat serum induces apoptosis of side population cells in gastric carcinoma[J].Exp Ther Med,2018,15(2):1718-27.
[12]Qian J,Li J,Jia J,et al.Diffrent concentrations of sijunzi decoction inhibit proliferation and induce apoptosis of human gastric cancer SGC-7901 side population[J].African J Tradit Complement Alternat Med,2016,13(4):145-56.
[13]Shim JH,Gim H,Lee S,et al.Inductions of caspase-,MAPK-and ROS-dependent apoptosis and chemotherapeutic effects caused by an ethanol extract of scutellaria barbata D.don in human gastric adenocarcinom cells[J].J Pharmacopunct,2016,19(2):129-36.
[14]Wang TS,Chen LJ,Wang ZY,et al.Purified alkaloid extract of Scutellaria barbata inhibits proliferation of hepatoma HepG-2 cells by inducing apoptosis and cell cycle arrest at G(2)/M phase[J].Afican J Pharm Pharmacol,2011,5(8):1046-53.
[15]Kan XE,Zhang WL,You RX,et al.Scutellaria barbata D,don extract inhibits the tumor growth through down-regulating of Treg cells and manipulating Th1/Th17 immune response in hepatoma H22-bearing mice[J].BMC Complement Altern Med,2017,17(41):1330-43.
[16]Jiang QQ,Li QY,Chen HW,et al.Scutellaria barbata D.Don inhibits growth and induces apoptosis by suppressing IL-6-inducible STAT3 pathway activation in human colorectal cancer cells[J].Exp Ther Med,2015,10(4):1602-8.
[17]Zheng QP,Qiu DS,Liu XJ,et al.Antiproliferative effect of Dendrobium catenatum Lindley polypeptides against human liver,gastric and breast cancer cell lines[J].Food Funct,2015,6(5):1489-95.
[18]Prasad R,Koch B.Antitumor activity of ethanolic extract of dendrobium formosum in T-Cell lymphoma:an in vitro and in vivo study[J].Biomed Res Int,2014,7534(51):6102-14.
[19]Meier P,Finch A,Evan G.Apoptosis in development[J].Nature,2000,407(6805):796-801.
[20]Oberst A,Bender C,Green DR.Living with death:the evolution of the mitochondrial pathway of apoptosis in animals[J].Cell Death Differ,2008,15(7):1139-46.
[21]Peng C,Rao W,Zhang L,et al.Mitofusin 2 ameliorates hypoxiainduced apoptosis via mitochondrial function and signaling pathways title of article(vol 69,pg 29,2015)[J].Intern J Biochem Cell Biol,2016,73(2):137-48.
[22]Andreu-Fernandez V,Sancho M,Genoves A,et al.Bax transmembrane domain interacts with prosurvival Bcl-2 proteins in biological membranes(vol 114,pg 310,2016)[J].Proc Natl Acad Sci USA,2017,114(8):E1574-85.
[23]Reyna DE,Gavathiotis E.Self-regulation of BAX-induced cell death[J].Oncotarget,2016,7(41):66326-7.
[24]Zhao H,Yenari MA,Cheng D,et al.Bcl-2 overexpression protects against neuron loss within the ischemic margin following experimental stroke and inhibits cytochrome c translocation and caspase-3 activity[J].J Neurochem,2003,85(4):1026-36.
[25]Haddadi R,Nayebi AM,Brooshghalan SE.Silymarin prevents apoptosis through inhibiting the Bax/caspase-3 expression and suppresses toll like receptor-4 pathway in the SNc of 6-OHDAintoxicated rats[J].Biomed Pharmacoth,2018,104(2):127-36.
[26]Garg AD,More S,Rufo N,et al.Trial watch:Immunogenic cell death induction by anticancer chemotherapeutics[J].Oncoimmunol,2017,6(12):139-44.
[27]Josephs SF,Ichim TE,Prince SM,et al.Unleashing endogenous TNF-alpha as a cancer immunotherapeutic[J].J Transl Med,2018,16(242):497-506.
[28]Coffin CS,Fraser HF,Panaccione R,et al.Liver diseases associated with Anti-Tumor necrosis factor-alpha(TNF-alpha)use for inflammatory bowel disease[J].Inflamm Bowel Dis,2011,17(1):479-84.
[29]Zhang HR,Park Y,Wu JX,et al.Role of TNF-alpha in vascular dysfunction[J].Clin Sci,2009,116(3/4):219-30.
[30]Ito S,Bollard CM,Carlsten MA,et al.Ultra-low dose interleukin-2promotes immune-m(o)dulating function of regulatory T cells and natural killer cells in healthy volunteers[J].Molecul Therapy,2014,22(7):1388-95.
[31]Sockolosky JT,Trotta E,Parisi G,et al.Selective targeting of engineered T cells using orthogonal IL-2 cytokine-receptor complexes[J].Science,2018,359(6379):1037-57.