加味四君子汤对H22荷瘤小鼠移植瘤血管生成的影响
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摘要
目的:观察加味四君子汤对H22荷瘤小鼠移植瘤血管生成的影响,并探讨其机制。方法:小鼠成瘤实验观察加味四君子汤对荷瘤小鼠的抑瘤和对癌周血管生长的影响。苏木素-伊红(HE)染色及免疫组化(IHC)检测荷瘤小鼠移植瘤中血管分布水平和血管内皮标记物(CD31)的表达水平。逆转录-聚合酶链式反应(RT-PCR)检测肿瘤组织血管内皮生长因子(VEGF),血管内皮生长因子受体2(VEGFR2),肿瘤坏死因子-α(TNF-α) mRNA表达。结果:加味四君子汤低、中、高剂量组(ig,11. 83,23. 66,47. 32 mg·kg~(-1)·d~(-1))抑瘤率分别达到29. 97%,59. 80%,82. 34%。与模型组相比,加味四君子汤中、高剂量组的平均瘤重明显降低(P <0. 05)。通过观察癌周血管数及形态发现,加味四君子汤中、高剂量组癌周血管数较模型组有所减少(P <0. 05),且血管管腔较窄。HE染色显示加味四君子汤中、高剂量组移植瘤中的血管分布均少于模型组。免疫组化显示加味四君子汤各剂量组的CD31阳性表达量均低于模型组(P <0. 01)。RT-PCR结果显示加味四君子汤中、高剂量组移植瘤VEGF,VEGFR2,TNF-αmRNA表达量均低于模型组(P <0. 01)。结论:加味四君子汤能够抑制H22荷瘤小鼠肿瘤生长,并抑制移植瘤血管新生,这可能与降低TNF-α,VEGF,VEGFR2的表达水平有关。
Objective: To investigate the effect of modified Si Junzitang on angiogenesis in transplanted tumor of H22 tumor-bearing mice. Method: The effect of modified Si Junzitang on tumor inhibition and growth of peripheral blood vessels in tumor-bearing mice was observed by tumorigenesis experiment in mice. Hematoxylineosin( HE) staining and immunohistochemistry( IHC) were used to detect the distribution of blood vessels and the expression of vascular endothelial markers( CD31) in tumor-bearing mice. Reverse transcription-polymerase chain reaction( RT-PCR) was used to detect the mRNA expression levels of vascular endothelial growth factor( VEGF),vascular endothelial growth factor receptor 2( VEGFR2) and tumor necrosis factor-α( TNF-α) in tumor tissue.Result: The inhibition rates of modified Sijunzi Tang in low-dose group( ig,11. 83 mg·kg~(-1)·d~(-1)),middle-dose group( ig,23. 66 mg·kg~(-1)·d~(-1)) and high-dose group( ig,47. 32 mg·kg~(-1)·d~(-1)) were 29. 97%,59. 80% and 82. 34%,respectively. Compared with the model group,the average tumor weight was lower in middle and highdose groups,with statistically significant differences( P < 0. 05). Through the observation of the number and morphology of pericardial vessels,the number of pericytes in middle and high-dose modified Sijunzitang groups was reduced compared with the model group( P < 0. 05),and the vascular lumen was narrow. HE staining showed that the distribution of blood vessels in middle and high-dose modified Si Junzitang groups was less than that of the model group. IHC showed that the positive expression of CD31 in each dose modified Si Junzitang group was lower than that in the model group( P < 0. 01). RT-PCR showed that the mRNA expressions of VEGF,VEGFR2 and TNF-α in middle and high-dose modified Si Junzitang groups were lower than those in the model group( P <0. 01). Conclusion: Modified Si Junzitang can inhibit the tumor growth of H22 tumor-bearing mice and the angiogenesis of transplanted tumors,which may be related to the reduction of TNF-α,VEGF and VEGFR2 expression levels.
Objective: To investigate the effect of modified Si Junzitang on angiogenesis in transplanted tumor of H22 tumor-bearing mice. Method: The effect of modified Si Junzitang on tumor inhibition and growth of peripheral blood vessels in tumor-bearing mice was observed by tumorigenesis experiment in mice. Hematoxylineosin( HE) staining and immunohistochemistry( IHC) were used to detect the distribution of blood vessels and the expression of vascular endothelial markers( CD31) in tumor-bearing mice. Reverse transcription-polymerase chain reaction( RT-PCR) was used to detect the mRNA expression levels of vascular endothelial growth factor( VEGF),vascular endothelial growth factor receptor 2( VEGFR2) and tumor necrosis factor-α( TNF-α) in tumor tissue.Result: The inhibition rates of modified Sijunzi Tang in low-dose group( ig,11. 83 mg·kg~(-1)·d~(-1)),middle-dose group( ig,23. 66 mg·kg~(-1)·d~(-1)) and high-dose group( ig,47. 32 mg·kg~(-1)·d~(-1)) were 29. 97%,59. 80% and 82. 34%,respectively. Compared with the model group,the average tumor weight was lower in middle and highdose groups,with statistically significant differences( P < 0. 05). Through the observation of the number and morphology of pericardial vessels,the number of pericytes in middle and high-dose modified Sijunzitang groups was reduced compared with the model group( P < 0. 05),and the vascular lumen was narrow. HE staining showed that the distribution of blood vessels in middle and high-dose modified Si Junzitang groups was less than that of the model group. IHC showed that the positive expression of CD31 in each dose modified Si Junzitang group was lower than that in the model group( P < 0. 01). RT-PCR showed that the mRNA expressions of VEGF,VEGFR2 and TNF-α in middle and high-dose modified Si Junzitang groups were lower than those in the model group( P <0. 01). Conclusion: Modified Si Junzitang can inhibit the tumor growth of H22 tumor-bearing mice and the angiogenesis of transplanted tumors,which may be related to the reduction of TNF-α,VEGF and VEGFR2 expression levels.
引文
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[14]禹雯琦,孙珏,周诣,等.四君子汤及加味方治疗恶性肿瘤的临床及基础研究进展[J].上海中医药大学学报,2017,31(5):95-101.
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[18] Sitohy B,Nagy J A,Dvorak H F. Anti-VEGF/VEGFR therapy for cancer:reassessing the target[J]. Cancer Res,2012,72(8):1909-1914.
[19] WANG Y,Nakayama M,Pitulescu M E,et al. EphrinB2 controls VEGF-induced angiogenesis and lymphangiogenesis[J]. Nature,2010,465(7297):483-486.
[20] Lee J K,Park S R,Jung B K,et al. Exosomes derived from mesenchymal stem cells suppress angiogenesis by down-regulating VEGF expression in breast cancer cells[J]. PLo S One,2013,8(12):e84256.
[21]王容容,谭小宁,李勇敏,等.健脾消癌方对大肠癌肝转移裸鼠模型肝组织PI3K/Akt通路相关蛋白表达的影响[J].中国实验方剂学杂志,2018,24(6):177-181.
[22] WANG F,WANG L,LI Y,et al. PAC-1 and its derivative WF-210 Inhibit Angiogenesis by inhibiting VEGF/VEGFR pathway[J]. Eur J Pharmacol,2018,821:29-38.
[23] Kim K W,Lee S J,Kim J C. TNF-alpha upregulates HIF-1 alpha expression in pterygium fibroblasts and enhances their susceptibility to VEGF independent of hypoxia[J]. Exp Eye Res,2017,164:74-81.
[2] Mcglynn K A, Petrick J L, London W T. Global epidemiology of hepatocellular carcinoma an emphasis on demographic and regional variability[J]. Clin Liver Dis,2015,19(2):223-238.
[3]邵艳,管静芝.原发性肝癌治疗进展[J].传染病信息,2016,29(4):248-252.
[4]张甘霖,杨国旺,于明薇,等.自噬——中医药抗肿瘤研究的新方向[J].中华中医药杂志,2014,29(2):507-510.
[5]奚胜艳,高学敏,张建军,等.从中医络病与病络理论认识肿瘤血管生成[J].北京中医药大学学报,2008,31(12):804-807,822.
[6]张广唱,武哲丽.四君子汤治疗肝癌实验研究概况[J].山东中医杂志,2015,34(8):643-645.
[7]许婉琦,王奕博,孙志蓉.石斛属植物抗肿瘤研究情况分析[J].中国现代应用药学,2017,34(1):130-134.
[8]林敬明,刘煜,罗荣城.半枝莲提取物抗人肝癌Hep-G2细胞增殖及其机制研究[J].南方医科大学学报,2006,26(7):975-977.
[9]林敬明,刘煜,罗荣城.半枝莲抑制人肝癌QGY-7701细胞增殖研究[J].南方医科大学学报,2006,26(5):591-593.
[10]初海姣,韩增祥,梁芳.中药复方抗肿瘤血管生成研究简况[J].实用中医内科杂志,2018,32(4):74-77.
[11]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.
[12]何小喜,莫书荣.红景天苷对腹水型H_(22)肝癌抑制作用及其机制的初步研究[J].中国药理学通报,2014,30(5):688-691.
[13]陈燕,胡浩,张诗军,等.四君子汤对原发性肝癌术后患者T淋巴细胞和NK细胞的影响及意义[J].中西医结合肝病杂志,2017,27(1):8-10.
[14]禹雯琦,孙珏,周诣,等.四君子汤及加味方治疗恶性肿瘤的临床及基础研究进展[J].上海中医药大学学报,2017,31(5):95-101.
[15] WANG T S,CHEN L J,WANG Z Y,et al. Purified alkaloid extract of Scutellaria barbata inhibits proliferation of hepatoma Hep G-2 cells by inducing apoptosis and cell cycle arrest at G(2)/M phase[J].Afr J Pharm Pharmacol,2011,5(8):1046-1053.
[16] Majchrzak K,Kaspera W,Szymas J,et al. Markers of angiogenesis(CD31, CD34, r CBV)and their prognostic value in low-grade gliomas[J]. Neurol Neurochir Pol,2013,47(4):325-331.
[17]郭晓强,隋爱霞.癌症抗血管生成疗法——“饿死肿瘤,挽救患者”治癌策略[J].生命世界,2016(8):76-81.
[18] Sitohy B,Nagy J A,Dvorak H F. Anti-VEGF/VEGFR therapy for cancer:reassessing the target[J]. Cancer Res,2012,72(8):1909-1914.
[19] WANG Y,Nakayama M,Pitulescu M E,et al. EphrinB2 controls VEGF-induced angiogenesis and lymphangiogenesis[J]. Nature,2010,465(7297):483-486.
[20] Lee J K,Park S R,Jung B K,et al. Exosomes derived from mesenchymal stem cells suppress angiogenesis by down-regulating VEGF expression in breast cancer cells[J]. PLo S One,2013,8(12):e84256.
[21]王容容,谭小宁,李勇敏,等.健脾消癌方对大肠癌肝转移裸鼠模型肝组织PI3K/Akt通路相关蛋白表达的影响[J].中国实验方剂学杂志,2018,24(6):177-181.
[22] WANG F,WANG L,LI Y,et al. PAC-1 and its derivative WF-210 Inhibit Angiogenesis by inhibiting VEGF/VEGFR pathway[J]. Eur J Pharmacol,2018,821:29-38.
[23] Kim K W,Lee S J,Kim J C. TNF-alpha upregulates HIF-1 alpha expression in pterygium fibroblasts and enhances their susceptibility to VEGF independent of hypoxia[J]. Exp Eye Res,2017,164:74-81.