蝎毒多肽干预K562/BALB/c-nu小鼠Hedgehog通路传导的机制研究

Mechanism of polypeptide extract from scorpion venom inhibiting Hedgehog signaling transduction in K562/BALB/c-nu leukemia mice

目的探究蝎毒多肽干预K562/BALB/c-nu白血病荷瘤小鼠Hedgehog通路信号传导的机制,从基因、蛋白水平解析蝎毒多肽抑制慢性粒细胞白血病发展的分子机制和作用靶点。方法将K562/BALB/c-nu白血病荷瘤小鼠分为对照组、模型组、伊马替尼(50 mg/kg)组和蝎毒多肽高、中、低剂量(0.3、0.6、1.2 mg/kg)组,药物干预14 d后,实时荧光定量PCR(q RT-PCR)法检测小鼠瘤组织Hedgehog信号通路上游因子Shh、Ptch、Smo m RNA表达水平,Western blotting法检测Shh、Ptch、Smo蛋白表达水平,ELISA法检测小鼠瘤组织Hedgehog信号通路下游因子Gli1蛋白表达水平。结果与模型组比较,蝎毒多肽干预组小鼠瘤组织Shh m RNA和蛋白表达水平均升高;蝎毒多肽低、中剂量组小鼠瘤组织Ptch、Smo m RNA及蛋白表达水平均降低;伊马替尼组各上游因子与模型组比较差异不显著;蝎毒多肽低、中剂量组小鼠瘤组织Gli1蛋白表达水平降低,蝎毒多肽高剂量组、伊马替尼组Gli1蛋白表达水平无显著差异。结论蝎毒多肽能够抑制Hedgehog信号通路上游因子Ptch、Smo以及下游因子Gli1的表达,而伊马替尼对Hedgehog信号通路无明显抑制作用。

Objective To study the mechanism of the Hedgehog signaling transduction intervened by polypeptide extract from scorpion venom(PESV)on K562/BALB/c-nu leukemia mice.Trying to analyze the molecular mechanisms and targets of the inhibited effect of PESV on chronic myeloid leukemia(CML)in vivo.Methods After establishing the K562/BALB/c-nu leukemia mice successfully,the model mice were divided into six groups which were the blank group,the PESV high,medium,and low doses(0.3,0.6,1.2 mg/kg)group,the Imatinib(50 mg/kg)group,and the model group.After 14 d drug intervention,the levels of gene and protein expression of Hedgehog signaling pathway upstream factors Shh,Ptch,and Smo were detected by q RT-PCR and Western blotting,and the protein expression of downstream factor Gli1 was determined by ELISA test.Results Compared to the model group,the genetic and protein expression of Shh which was an upstream factor were increased in the PESV groups.The m RNA and protein expression of Ptch and Smo in PESV low-dose and medium-dose groups were decreased.There were no significant differences of upstream factors between Imatinib group and model group.The concentration of downstream Gli1 protein significantly decreased within low-dose and medium-dose PESV groups,while there was no significant difference between high-dose PESV group and Imatinib group.Conclusion PESV can inhibit the expression of Hedgehog signaling pathway upstream factor Ptch,Smo and downstream factor Gli1 on the m RNA and protein level,while Imatinib has no obvious inhibitory effect on the Hedgehog signaling pathway.