养营活血汤干预肝癌皮下瘤的实验研究

Experimental Study of Yangying Huoxue Decoction(养营活血汤)on Subcutaneous Tumor of Liver Cancer

目的 中药复方药干预肝癌皮下瘤模型在中医药研究肝癌体内实验中越来越多,是中医科研工作者的研究肝癌的重要手段。适宜的细胞悬液注射量、细胞悬液注射浓度、参药介质、给药的中药复方浓度等细节贯穿整个造模和干预的过程,直接影响中药复方的干预效果,是中药复方体内实验研究取得成功的细节和必备点。基于课题组在进行养营活血汤干预肝癌皮下瘤模型的过程中查阅相关文献及制备方法,对肝癌皮下瘤造模及养营活血汤干预制备模型进行详细的梳理,并提出一些见解,以飨同道。方法 40只裸鼠BALB/c随机分成4组,每组10只。第1组10只裸小鼠注射密度为1×10^(7)个/mL 0.2 mL混合基质胶的细胞悬液;第2组10只裸小鼠注射密度为1×10^(7)个/mL 0.1 mL混合基质胶的细胞悬液;第3组10只裸小鼠注射细胞密度为1×10^(6)/mL 0.1 mL氯化钠溶液混悬的细胞悬液;第4组10只裸小鼠注射细胞密度为1×10^(6)mL 0.1混合基质胶注的细胞悬液。其中第1组与第2组注射量不同做对比;第2组与第3组不同浓度做对比;第3组与第4组不同介质做对比;细胞悬液分别皮下注射于裸鼠右腋,期间观察肿瘤大体特征、成瘤时间、成瘤率、成瘤大小、成瘤大小差异等,病理切片及电镜下观察肿瘤病理学特征。造模成功后使用中药复方养营活血汤干预皮下瘤模型7 d,期间观察小鼠肿瘤增大速度趋势、小鼠状态等。结果 第2组中当每个注射点细胞悬液浓度为1×10^(7)时混合基质胶皮下注射0.1 mL时,成瘤时间相对较短,8 d成瘤率90%,成瘤差异较小,其8d成瘤最大直径平均值长度为(0.98±0.88)cm。养营活血汤干预7 d后,小鼠状态较第1组好,其肿物增长趋势在合适区间内。结论 本实验4种方法均成功建立了H22肝癌裸鼠皮下移植瘤模型,当每个注射点H22悬液浓度为1×10^(7)时混合基质胶皮下注射0.1 mL时(第2组),该组模型成瘤率高,瘤体形状较规则、实验鼠皮下瘤大小较一致,更适合笔者一周左右的中药复方干预。第1组造模后皮下瘤肿物平均最长直径过大超过合适区间内,在干预期间7 d内小鼠病死率高,第3、4组皮下瘤模型成瘤速度慢,可能相对于第2组而言更适合较长期的中药复方干预。此次造模探究为研究短期中医干预措施提供了较为理想的动物模型。

Objective TCM compound drugs intervene in subcutaneous tumor models of liver cancer in more and more in vivo experiments of TCM research on liver cancer,and it is an important method for TCM researchers to study liver cancer.The de-tails of the appropriate cell suspension injection volume,cell suspension injection concentration,ginseng medium,and the con-centration of the traditional Chinese medicine compound to be administered run through the entire modeling and intervention process,and directly ffect the intervention effect of the traditional Chinese medicine compound,which are the details and must haves for success.Based on the reviewr of relevant literature and preparation methods in the process of Yangying Huoxue Decoc-tion(养营活血汤)intervening liver cancer subcutaneous tumor model,the model of liver cancer subcutaneous tumor and Yangy-ing Huoxue Decoction intervention preparation model were sorted out in detail and some insights were put forward to enjoy fellow.Methods Forty nude mice BALB/c were randomly divided into 4 groups with 10 mice in each group.10 nude mice in group 1 were injected with cell suspension with a density of 1×10^(7)clls/mL in 0.2 mL of mixed matrigel;10 nude mice in group 2 were injected with a density of1×10^(7)cells/mL with 0.1 mL of mixed matrigel cell suspension;10 nude mice in group 3 were injected with cell suspension with a cell density of1×10^(6)/mL and 0.1 mL of sodium chloride solution;10 nude mice in group 4 were injected with a cell density of 1×10^(6) 0.1 mL of mixed matrigel-injected cell suspension.Among them,the first and second groups were compared with different injection doses;the second and third groups were compared with different concentrations;the third and fourth groups were compared with different media;the cell suspensions were subcutaneously injected into the right axilla of nude mice,respectively.During the period,the general characteristics of the tumor,the time of tumor formation,the tumor formation rate,the size of the tumor,and the difference in the size of the tumor were observed.After successful modeling,the subcutaneous tumor model was intervened with the traditional Chinese medicine compound Yangying Huoxue Decoction for 7 days,during which the trend of tumor growth rate and the state of the mice were observed.Results In group 2,when the concentration of cell suspension at each injection point was 1×107,when 0.1 mL of mixed matrigel was injected subcutaneously,the tumor formation time was relatively short,the 8-day tumor formation rate was 90%,and the difference in tumor formation was small.The average length of the largest diameter of the tumor was(0.98±0.88)cm.After 7 days of intervention with Yangying Huoxue Decoction,the state of mice was better than that of the first group,and the tumor growth trend was within an appropriate range.Conclusion The four methods in this experiment had successfully established the H22 liver cancer subcutaneous transplanted tumor model in nude mice.When the concentration of H22 suspension at each injection point was 1×10^(7),0.1 mL of matrigel is injected subcutaneously(group 2).The tumor formation rate was high,the shape of the tumor body was relatively regular,and the size of the subcutaneous tumor in the experimental mice was relatively consistent,which was more suitable for the author s intervention of traditional Chinese medicine for about a week.The average longest diameter of the subcutaneous tumor in the first group was too large and exceeded the appropriate range,and the mortality rate of the mice was high within 7 days during the intervention period.For the group,it is more suitable for longer-term traditional Chinese medicine compound intervention.This modeling exploration provides an ideal animal model for the study of short-term TCM interventions.