重组人内抑素对兔耳增生性瘢痕成纤维细胞内钙离子浓度的影响

黄天宇; 龚永芳; 王晶; 李洋; 杨鑫雨; 项平

doi:10.13898/j.cnki.issn.1000-2200.2020.02.010

重组人内抑素对兔耳增生性瘢痕成纤维细胞内钙离子浓度的影响

Effect of recombinant human endostatin on the intracellular Ca²⁺ concentration in hypertrophic scar fibroblasts in a rabbit ear model

摘要

摘要:
目的通过探讨重组人内抑素（rhEndostatin）促进兔耳增生性瘢痕成纤维细胞（HSFs）凋亡的部分机制，为临床上药物治疗增生性瘢痕（HS）提供实验依据。
方法取新西兰大耳兔6只制备瘢痕模型。以HSFs为研究对象，应用激光共聚焦显微镜（CLSM）结合Fluo-4/AM（Ca²⁺荧光指示剂）检测在Hanks与D-Hanks液中（有、无细胞外钙）rhEndostatin（100 μg/mL）对HSFs胞内Ca²⁺浓度（Ca²⁺i）的影响。
结果当细胞外液为Hanks液时，rhEndostatin（100 μg/mL）可使HSF胞质内Ca²⁺i水平持续增加，当持续10 s给予rhEndostatin处理后，HSFs胞内Ca²⁺i可迅速增加达峰顶再缓慢下降，在停止药物处理后，HSFs胞内Ca²⁺i继续下降，停止处理约50 s后，胞内Ca²⁺i尚未至基线水平；而当细胞处于D-Hanks液中时，rhEndostatin对HSFs胞质内Ca²⁺i无明显影响。
结论rhEndostatin可通过干扰HSFs胞内钙离子稳态，推动胞外Ca²⁺内流导致胞内钙超载来诱导HSFs凋亡。

Abstract:
ObjectiveTo investigate the mechanism of recombinant human endostatin(rhEndostatin) promoting the apoptosis of hypertrophic scar fibroblast(HSFs) in rabbit ear, and provide the experimental basis for clinical drug treatment of hypertrophic scar(HS).
MethodsThe HS model was prepared in 6 healthy New Zealand white albino rabbits.The effects of rhEndostatin(100 μg/mL) on the intracellular Ca²⁺ concentration(Ca²⁺ i) of HSFs in Hanks and D-Hanks fluids(with and without extracellular calcium) were detected using laser confocal microscopy(CLSM) combined with Fluo-4 /AM(Ca²⁺ fluorescence indicator).
ResultsWhen the extracellular fluid was the Hanks fluid, the rhEndostatin could cause the HSFs Ca²⁺i increasing, and the levels of Ca²⁺i reached to peak after 10 s of treatment, then slowly decreased with time.After stopping the drug treatment, the levels of Ca²⁺i continued to slow down, and the level of HSFs Ca²⁺i did not return to baseline after 50 s of rhEndostatin withdrawal.HSFs Ca²⁺i was not affected by rhEndostatin when the cells were incubated with D-Hanks buffer.
ConclusionsRhEndostatin can induce the HSFs apoptosis by interfering with intracellular Ca²⁺ homeostasis and promoting extracellular Ca²⁺ influx to cause intracellular calcium overload.

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重组人内抑素对兔耳增生性瘢痕成纤维细胞内钙离子浓度的影响

Effect of recombinant human endostatin on the intracellular Ca2+ concentration in hypertrophic scar fibroblasts in a rabbit ear model

Effect of recombinant human endostatin on the intracellular Ca²⁺ concentration in hypertrophic scar fibroblasts in a rabbit ear model