利用斑马鱼肝癌模型探究氨基葡萄糖盐酸盐对肝癌的影响  

Exploring the Effect of Glucosamine Hydrochloride on Liver Cancer Using Zebrafish Liver Cancer Model

作　　者：吕昊坤 杨腾辉 吴启赐 潘裕添[1] 薛钰[1] LU Haokun;YANG Tenghui;WU Qici;PAN Yutian;XUE Yu(Fungal Industry Engineering Technology Center of Minnan Normal University,Zhangzhou 363000,China)

机构地区：[1]闽南师范大学菌物产业工程技术中心,福建漳州363000

出　　处：《食品工业科技》2024年第20期332-340,共9页Science and Technology of Food Industry

基　　金：国家自然科学基金面上项目(32370888);福建省自然科学基金项目(2024J09046);闽南师范大学培育项目(MSPY202101)。

摘　　要：目的:探究双孢蘑菇来源的氨基葡萄糖盐酸盐(Glucosamine Hydrochloride,GAH)对肝癌的影响。方法:利用斑马鱼肝癌及血管双转基因模型Tg(flk:mCherry;kras^(G12V)),在60 mg/mL盐酸多西环素溶液的诱导下,通过激光共聚焦显微镜追踪观察第5 d到第10 d肝癌的发生发展;在Dox诱导的同时加入不同浓度GAH恢复,在激光共聚焦显微镜下比较分析正常组(Ctr)、模型组(Doxorubicin,Dox)、Dox+0.1%GAH组、Dox+0.3%GAH组中GAH对肝癌及肝癌血管生长的影响;通过qPCR和ELISA实验分析GAH对肝癌及肝癌血管相关调控因子mRNA和蛋白水平表达的影响(prmt5、tiam1、kat5、vegfaa、vegfr2、tgf-β1);最后通过TIMER2在线生物信息学分析验证tgf-β1与prmt5、tiam1、kat5与vegfaa、vegfr2的相关性。结果:Dox持续诱导会引起肝癌的发生发展;与Dox模型组相比,GAH处理极显著抑制肝癌的生长与侵袭(P<0.001),并能极显著改善肝癌血管紊乱(P<0.001);qPCR和ELISA实验结果表明GAH能够显著抑制肝癌及血管相关基因的mRNA和蛋白水平(P<0.05);生物信息学分析结果提示tgf-β1与肝癌及血管相关基因的表达具有明显相关性。结论:GAH可能通过调控TGF-β信号通路影响肝癌相关基因的表达抑制肝癌血管的生成,从而抑制肿瘤的生长与侵袭。Objective:To explore the effects of Agaricus bisporu-derived glucosamine hydrochloride(GAH)on hepatocellular carcinoma(HCC).Methods:A zebrafish liver cancer and vascular double transgenic model,Tg(flk:mCherry;krasG12V)was induced using a 60 mg/mL doxycycline hydrochloride solution.Larvae were traced and observed for the development of liver cancer from 5 d to 10 d using laser confocal microscopy.Different concentrations of GAH were added during Dox induction,larvae in Control group(Ctr),modeling group(Dox),Dox+0.1%GAH group and Dox+0.3%GAH group(Dox)were analyzed under laser confocal microscopy to observe the effect of GAH on liver cancer and tumor vascular growth.The effect of GAH on the mRNA and protein expression of HCC and vascular-related regulatory factors(prmt5,tiam1,kat5,vegfaa,vegfr2,tgf-β1)was analyzed via qPCR and ELISA assays.Finally,the correlations between tgf-β1,prmt5,tiam1,kat5,vegfaa and vegfr2 were validated through TIMER2 online bioinformatics analysis tool.Results:Continuous induction of Dox facilitated the emergence of HCC.Compared with the Dox model group,the growth and invasion of HCC were significantly inhibited after GAH treatment(P<0.001),and the vascular disorder of HCC was greatly improved(P<0.001).GAH intervention markedly ameliorated vascular dysregulation associated with liver cancer(P<0.001).qPCR and ELISA assays revealed that GAH could effectively suppress the mRNA and protein levels of genes associated with HCC and its vasculature(P<0.05).The significance of tgf-β1 in the expression of genes related to HCC and vascular pathways was proved using the TIMER2 database.Conclusion:GAH would potentially inhibit tumor growth and invasiveness by regulating the TGF-βsignaling pathway,thereby influencing the expression of genes related to HCC and inhibiting the development of HCC vasculature.

关 键 词：氨基葡萄糖盐酸盐 斑马鱼肝癌模型 盐酸多西环素 TGF-Β信号通路 

分 类 号：TS201.4[轻工技术与工程—食品科学]