Vascular endothelium-specific overexpression of human catalase in cloned pigs.
Vascular endothelium-specific overexpression of human catalase in cloned pigs.
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DOI：
10.1007/s11248-010-9473-7
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发表时间：
2011-10
影响因子：
3
通讯作者：
Laughlin MH
中科院分区：
文献类型：
--
作者：
Whyte JJ;Samuel M;Mahan E;Padilla J;Simmons GH;Arce-Esquivel AA;Bender SB;Whitworth KM;Hao YH;Murphy CN;Walters EM;Prather RS;Laughlin MH
本研究的目的是开发以内皮特异性方式过表达人过氧化氢酶（hCat）的转基因尤卡坦小型猪。过氧化氢酶代谢过氧化氢（H2 O2），这是一种重要的血管张力调节剂，有助于动脉粥样硬化和先兆子痫等疾病。因此，研究减少内皮源性H2 O2的大型动物模型将产生关于健康和疾病中血管调节的有价值的转化数据。将用人过氧化氢酶（Tie 2-hCat）和eGFP表达构建体稳定共转染的尤卡坦小型猪胎儿成纤维细胞分离成单细胞群。通过PCR确定成纤维细胞的单个集落中Tie 2-hCat转基因的存在。将转基因成纤维细胞用于通过电融合将核转移到去核卵母细胞中。每头代孕母猪至少移植140个克隆胚胎（n = 4）。所有四个代理人保持怀孕和仔猪通过剖腹产分娩。来自四窝中的三窝的九只雄性仔猪携带Tie 2-hCat转基因。通过RT-PCR和Western blot分别验证转基因仔猪分离的脐静脉内皮细胞中人过氧化氢酶mRNA和总体升高的过氧化氢酶蛋白的表达，并通过免疫组织化学证实内皮定位。与野生型内皮细胞相比，转基因内皮细胞中过氧化氢酶的酶活性增加是基于培养物中H2 O2水平显著降低而推断的。猪和人类心血管解剖学和生理学的相似性将使该猪模型成为关于内皮源性H2 O2在血管舒张中的假定作用以及血管健康和疾病的潜在机制的有价值的信息来源。
The objective of this study was to develop transgenic Yucatan minipigs that overexpress human catalase (hCat) in an endothelial-specific manner. Catalase metabolizes hydrogen peroxide (H2O2), an important regulator of vascular tone that contributes to diseases such as atherosclerosis and preeclampsia. A large animal model to study reduced endothelium-derived H2O2 would therefore generate valuable translational data on vascular regulation in health and disease. Yucatan minipig fetal fibroblasts stably co-transfected with human catalase (Tie2-hCat) and eGFP expression constructs were isolated into single-cell populations. The presence of the Tie2-hCat transgene in individual colonies of fibroblasts was determined by PCR. Transgenic fibroblasts were used for nuclear transfer into enucleated oocytes by electrofusion. A minimum of 140 cloned embryos were transferred per surrogate sow (n = 4). All four surrogates maintained pregnancies and piglets were delivered by cesarean section. Nine male piglets from three of the four litters carried the Tie2-hCat transgene. Expression of human catalase mRNA and overall elevated catalase protein in isolated umbilical endothelial cells from transgenic piglets were verified by RT–PCR and western blot, respectively, and endothelial localization was confirmed by immunohistochemistry. Increased enzymatic activity of catalase in transgenic versus wild-type endothelial cells was inferred based on significantly reduced levels of H2O2 in culture. The similarities in swine and human cardiovascular anatomy and physiology will make this pig model a valuable source of information on the putative role of endothelium-derived H2O2 in vasodilation and in the mechanisms underlying vascular health and disease.
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影响因子：
3.1
作者：
Kumar S;Sun X;Wiseman DA;Tian J;Umapathy NS;Verin AD;Black SM
通讯作者：
Black SM
影响因子：
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作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
Wang, Wen-Hui
影响因子：
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作者：
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通讯作者：
King, W. Allan
影响因子：
14.8
作者：
Baudin, Bruno;Bruneel, Arnaud;Vaubourdolle, Michel
通讯作者：
Vaubourdolle, Michel