| 赖允丽张学荣△ 张钦乐胡仁统罗小玲廖明班建东.眼镜蛇毒神经生长因子抑制肝星状细胞增殖并诱导其凋亡[J].,2012,12(6):1034-4039 |
| 眼镜蛇毒神经生长因子抑制肝星状细胞增殖并诱导其凋亡 |
| Cobra Venom NGF Inhibits Proliferation and Induces Apoptosis ofHepatic Stellate Cell |
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| DOI： |
| 中文关键词: 眼镜蛇毒 NGF 凋亡 肝纤维化 |
| 英文关键词: Cobra venom NGF Liver fibrosis Apoptosis |
| 基金项目:广西自然科学基金资助项目：(2011GXNSFA018268)，(KFJJ2010-43) |
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| 中文摘要: |
| 目的：从眼镜蛇毒中分离纯化神经生长因子(Nerve Growth Factor, NGF)，观察眼镜NGF 对肝星状细胞HSC-T6 增殖、凋亡活
性的影响，进一步为蛇毒NGF 在抗肝纤维化治疗提供依据。方法：采用shephadex G-75 和CM Sepharose CL-6B 二步柱色谱对眼
镜蛇毒NGF 进行纯化分离；PC12 细胞测定各洗脱峰的活性，再用SDS-PAGE 鉴定具有NGF 活性洗脱峰的纯度和相对分子质
量。实验设立空白对照和NGF 处理组，分别作用于HSC-T6，培育相应时间，MTT 检测眼镜蛇毒NGF 对HSC-T6 细胞活力影响；
HE 染色、紫外激光显微镜与透射电镜观察HSC-T6 细胞的形态学变化；TUNEL、流式细胞技术检测眼镜蛇毒NGF 对HSC-T6 细
胞凋亡的影响。结果：眼镜蛇毒经PC-12 细胞鉴定第Ⅵ峰具有NGF 活性；SDS-PAGE 检测为电泳纯，相对分子质量为22.3KD；
NGF 对HSC-T6 细胞增殖具有明显抑制作用（2 μg/ml NGF 的抑制率为49.66% ±6.50%，P<0.05；6.25 μg/ml NGF 的抑制率为
71.33%±1.53%，P<0.05）；TUNEL 法检测发现NGF 干预组的凋亡率28.71% ±1.59%（2ug/ml NGF） 和34.4% ±2.49%（5 μg/ml
NGF）明显高于对照组的15.85% ±1.58%（P<0.05）;
流式细胞仪也有同样的发现，NGF 干预组的凋亡率16.12% ±3.02%（2 ug/ml
NGF） 和21.15% ±3.31%（5 μg/ml NGF） 明显高于对照组的2.7% ±1.55%（P<0.05）。结论：眼镜蛇毒NGF 能抑制肝星状细胞
HSC-T6 增殖并诱导其凋亡。 |
| 英文摘要: |
| Objective: Cobra venom NGF canbe isolate and purfity with a rapid method,and observe the effect of the proliferation
and apoptosis after NGF add to the HSC-T6 cells. This provide the basis for the treatment of anti-liver fibrosis in further. Methods: NGF
is separated from guangxi cobra venom through shephadex G-75 and CM Sepharose CL-6B column chromatography; PC12 cell assay the
activity of the NGF elution peak, then identified NGF purity and relative molecular mass with SDS-PAGE; we establishment groups of
control and NGF respectively, acting on the HSC-T6, nurturing the corresponding time points, MTT test HSC-T6 cells viability effected
on cobra venom NGF; hematoxylin and eosin stain、PALM MicroBeam and transmission electron microscope Morphology observed the
changes in HSC-T6 cells; Tunel、and flow cytometry detected HSC-T6 cell apoptosis after adding the cobra venom NGF; Results: The
Cobra venom elution peak Ⅵ identified by PC-12 cells having NGF active;SDS-PAGE electrophoresis show the elution peak Ⅵ pure and
the relatived molecular mass is 22.3KD; HSC-T6 cell proliferation is inhibited evidently (2 μg/mlNGF inhibition ratio is 49.66% ±
6.50%; 6.25 μg/mlNGF inhibition ratio is 71.33% ±1.53%, P<0.05); TUNEL shows apoptosis ratio of HSC-T6 is 28.71%±1.59%(2
μg/ml NGF) and 34.4% ±2.49 % (5 μg/mlNGF) that is obviously higer than control 15.85% ±1.58%, P<0.05; It is also founded in flow
cytometry, apoptosis ratio of HSC-T6 is 16.12%±3.02% (2 μg/ml NGF) and 21.15%±3.31%(5?g/mlNGF) that is obviously higer than
control 2.7%±1.55%, P<0.05. Conclusion: The cobra venom NGF inhibits HSC-T6 proliferation and induces apoptosis. |
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