血管内皮细胞条件性敲除 Cx43 杂合子小鼠模型建立及其血管功能变化的研究
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1. 陆军军医大学大坪医院 野战外科研究部,战伤休克与输血研究室,创伤、烧伤与复合伤国家重点实验室,重庆 400042;2. 陆军军医大学大坪医院 野战外科研究部, 武器杀伤生物效应评估研究室,创伤、烧伤与复合伤国家重点实验室,重庆 400042

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Vascular function in a heterozygous mouse model with conditional knockout of connexin 43 in vascular endothelial cells
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1.State Key Laboratory of Trauma,Burns and Combined Injury ,Department of Shock and 'Transfusion,ResearchInstitute of Surgery ,Daping Hospital,Army Medical University,Chongqing 400042,China.2. State Key Laboratoryof Trauma,Burms and Combined Injury,Department of Weapon Bioeffect Assessment,Research Institute of Surgery, Daping Hospital ,Army Medical University ,Chongqing 400042

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    摘要:

    目的 建立血管内皮细胞连接蛋白 43(connexin 43, Cx43)条件性敲除小鼠模型,并对其血管舒张/收缩功能进行检测。 方法 将 8 只 Cx43flox / flox小鼠与 C57BL/ 6 品系野生型(wild type, WT)小鼠交配,子代小鼠继续与 C57BL/ 6 小鼠回交 9 代进行基因背景转换;再将 Cx43flox / +小鼠与血管内皮细胞特异性表达 Tie2-Cre 重组酶小鼠(以下简称 Cre 小鼠)交配,获得 Tie2-Cre / Cx43flox / +小鼠。 利用鼠尾 PCR 试剂盒进行基因型鉴定,Western Blot 法和免疫荧光法检测小鼠肠系膜上动脉(superior mesenteric artery, SMA)中 Cx43 表达。利用离体张力测定技术检测SMA 血管舒张/ 收缩功能,包括去甲肾上腺素诱导的收缩反应性和乙酰胆碱诱导的舒张反应性。 结果 PCR 电泳和血管蛋白表达检测结果证实 Tie2-Cre / Cx43flox / +小鼠成功构建,肠系膜上动脉中 Cx43 蛋白表达与野生型小鼠相比明显降低(P< 0. 01)。 与 WT 小鼠相比,部分敲除血管内皮细胞 Cx43 后使乙酰胆碱诱导的内皮依赖的血管舒张反应性显著降低(P< 0. 01)。 结论 采用 Cx43flox / flox小鼠与血管内皮细胞特异性表达 Tie2-Cre 重组酶小鼠成功构建血管内皮细胞条件性敲除 Cx43 杂合子小鼠,该小鼠的肠系膜上动脉中 Cx43 表达降低,且舒张反应性明显降低,预期可为研究 Cx43 及其相关结构在血管功能调节中的作用提供动物模型。

    Abstract:

    Objective To establish an endothelial conditional connexin 43 (Cx43)-knockout mouse model and use it to examine vascular relaxation / constriction. Methods Eight Cx43flox / flox mice were crossed with C57BL/ 6 wild type (WT) mice and the offspring were backcrossed with C57BL/ 6 mice. This backcross process was repeated nine times, after which the Cx43flox / + mice were crossed with vascular endothelial-cell-specific Tie2-Cre recombinase mice to obtain Tie2-Cre /Cx43flox / + mice. The genotype was confirmed by mouse tail direct polymerase chain reaction (PCR). Expression of Cx43 in the superior mesenteric artery ( SMA) was measured by Western Blot and immunofluorescence. SMA rings were used to measure vascular relaxation and the contractile response to acetylcholine (ACh) and norepinephrine. Results Tie2-Cre /Cx43flox / +offspring were obtained and identified by PCR, Western Blot and immunofluorescence. Cx43 expression and AChinduced endothelium-dependent relaxation reactivity of the SMA were significantly decreased in Tie2-Cre / Cx43flox / + mice compared with WT mice ( P < 0. 01). Conclusions A mouse model with endothelium-specific Cx43-knockout was successfully established by intercrossing Tie2-Cre mice with Cx43flox / flox mice. Cx43 expression in vascular tissues and the relaxation reactivity of SMA were decreased by conditional Cx43 knockout. These mice may thus provide an animal model for studying the link between Cx43 and vascular function.

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彭小勇,周远群,张紫森,邓蒙生,雷艳,李涛,刘良明,王建民,康建毅,杨光明.血管内皮细胞条件性敲除 Cx43 杂合子小鼠模型建立及其血管功能变化的研究[J].中国实验动物学报,2022,30(7):880~886.

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  • 收稿日期:2022-05-24
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  • 在线发布日期: 2023-03-10
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