Generation and Characterization of a Novel Mouse Line, Keratocan-rtTA (KeraRT), for Corneal Stroma and Tendon Research.
Generation and Characterization of a Novel Mouse Line, Keratocan-rtTA (KeraRT), for Corneal Stroma and Tendon Research.
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DOI：
10.1167/iovs.17-22661
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发表时间：
2017-09-01
影响因子：
4.4
通讯作者：
Liu CY
中科院分区：
文献类型：
--
作者：
Zhang Y;Kao WW;Hayashi Y;Zhang L;Call M;Dong F;Yuan Y;Zhang J;Wang YC;Yuka O;Shiraishi A;Liu CY
我们创建了一种新的诱导型小鼠系Keratocan-rtTA（KeraRT），其允许在发育期间和成人中对角膜角膜细胞和腱细胞进行特异性遗传修饰。构建基因靶向载体（Kera-IRES 2-rtTA 3），并通过基因靶向技术将其插入小鼠Kera等位基因的终止密码子之后。将得到的KeraRT小鼠与tet-O-Hist 1H 2B-EGFP（TH 2B-EGFP）杂交，获得KeraRT/TH 2B-EGFP复合转基因小鼠，其中表达Kera的细胞通过多西环素（Dox）诱导用绿色荧光蛋白（GFP）标记。在KeraRT/TH 2B-EGFP中检测GFP和内源性Kera的表达模式。此外，将KeraRT与tet-O-TGF-α繁殖以产生双转基因小鼠KeraRT/tet-O-TGF-α，以在Dox诱导后在角膜基质细胞中过表达TGF-α。当KeraRT/TH 2B-EGFP小鼠喂食Dox食物时，在角膜基质、四肢和尾部检测到强GFP标记的细胞。在任何单个转基因KeraRT或TH 2B-EGFP小鼠中没有GFP。组织学分析显示，角膜中的GFP仅限于KeraRT/TH 2B-EGFP的基质角膜细胞，这与Kera表达一致。GFP的诱导在24小时内发生，并在Dox诱导后7天达到平台期。在KeraRT/TH 2B-EGFP诱导后3个月可检测到GFP。TGF-α在角膜基质细胞中的异位表达导致角膜上皮和基质的增生。新的Dox诱导型KeraRT驱动小鼠系是一种有用的遗传工具，用于基因操作和阐明胚胎发育、稳态和发病过程中角膜基质和四肢和尾部肌腱中的基因功能。
We created a novel inducible mouse line Keratocan-rtTA (KeraRT) that allows specific genetic modification in corneal keratocytes and tenocytes during development and in adults. A gene-targeting vector (Kera- IRES2-rtTA3) was constructed and inserted right after the termination codon of the mouse Kera allele via gene targeting techniques. The resulting KeraRT mouse was crossed to tet-O-Hist1H2B-EGFP (TH2B-EGFP) to obtain KeraRT/TH2B-EGFP compound transgenic mice, in which cells expressing Kera are labeled with green fluorescence protein (GFP) by doxycycline (Dox) induction. The expression patterns of GFP and endogenous Kera were examined in KeraRT/TH2B-EGFP. Moreover, KeraRT was bred with tet-O-TGF-α to generate a double transgenic mouse, KeraRT/tet-O-TGF-α, to overexpress TGF-α in corneal keratocytes upon Dox induction. Strong GFP-labeled cells were detected in corneal stroma, limbs, and tail when KeraRT/TH2B-EGFP mice were fed Dox chow. There was no GFP in any single transgenic KeraRT or TH2B-EGFP mouse. Histological analysis showed that GFP in the cornea was limited to stromal keratocytes of KeraRT/TH2B-EGFP, which is consistent with Kera expression. Induction of GFP occurred in 24 hours and reached a plateau by 7 days after Dox induction. GFP could be detected 3-months after induction of KeraRT/TH2B-EGFP. Ectopic expression of TGF-α in corneal keratocytes caused hyperplasia in the corneal epithelium and stroma. The novel Dox inducible KeraRT driver mouse line is a useful genetic tool for gene manipulation and elucidating gene functions in corneal stroma and tendons of limbs and tail during embryonic development, homeostasis and pathogenesis.
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影响因子：
6
作者：
Zheng, B;Zhang, ZP;Denton, CP
通讯作者：
Denton, CP
影响因子：
--
作者：
Cho, Andrew;Haruyama, Naoto;Kulkarni, Ashok B
通讯作者：
Kulkarni, Ashok B
影响因子：
7
作者：
Rees, S. G.;Waggett, A. D.;Hughes, C. E.
通讯作者：
Hughes, C. E.
影响因子：
4
作者：
Zhang, Yujin;Kao, Winston W. -Y.;Liu, Chia-Yang
通讯作者：
Liu, Chia-Yang
DOI：
10.1111/j.1749-6632.1999.tb08574.x
发表时间：
1999-01-01
期刊：
CHARCOT-MARIE-TOOTH DISORDERS
影响因子：
--
作者：
Feltri, ML;D'Antonio, M;Wrabetz, L
通讯作者：
Wrabetz, L