組織專一性破壞自分泌運動因子(Autotaxin)導致斑馬魚神經管發育異常

自分泌運動因子(Autotaxin, Atx)是一種存在於大部分的體液當中之分泌性溶血磷脂酶，在胚胎發育的過程中扮演著重要的角色。已知在老鼠胚胎中剔除Atx導致血管與神經系統發育不全，並於胚胎發育期9.5天時死亡。然而，Atx調控神經系統發育的作用機制則尚未明瞭，主要由於哺乳動物胚胎發育不易觀察，因此我嘗試利用體外發育且通體透明之斑馬魚建立Atx缺失模式以利機轉探討。首先我觀察到atx表現於出生後11小時至36小時的斑馬魚胚之神經系統，利用嗎

關鍵字

斑馬魚；自分泌運動因子；神經管發育；神經中間線形成

並列摘要

Autotaxin (Atx), a secreted form of lysophospholipase D, is present in most body fluids. Atx is vital for different developmental processes. Loss of Atx causes vascular and neural defects and embryonic lethality at E9.5 in mice. However, how Atx affects neural development in vivo remains largely unclear mainly due to the difficulty of studying early development in mammals. To address this point, I used translucent zebrafish embryos as a model. I observed that atx expressed in neural tissues from 11 to 36 hours post fertilization (hpf), corresponding to the period of early neural development. Knockdown of atx with morpholino caused defects in formation of midbrain-hindbrain boundary (MHB) and brain ventricles in 24-hpf embryos. I further examined midbrains of atx morphants by cryosection at various developmental stages and observed that neural tube and midline formation were severely affected. It suggests that the Atx loss of function cause defects in neural cell migration toward the midline during neurulation. Knockdown of atx by MO is known to inhibit the formation of Küpffer vesicle, left-right asymmetry and subsequent cardiac looping in zebrafish. These defects may secondarily interfere with neurulation. To avoid this complication, I generated a neural-specific CRISPR construct targeting atx driven by a gfap promoter and successfully disrupted atx in neural tissue. I found brain and neural tube defects similar to atx morphants in the 48 hpf F0 embryos-injected with the atx-targeting plasmid. The formation of neural tube midline was severely delayed or malformed. Taken together, I unequivocally demonstrate the specific role of Atx in the neural tube formation and brain development in zebrafish. Further investigation using this model could help to elucidate the regulation of neural tube formation by Atx.

並列關鍵字

Autotaxin ； midline formation ； neural tube formation ； neurulation ； zebrafish

參考文獻

Ablain, J., E. M. Durand, S. Yang, Y. Zhou and L. I. Zon (2015). "A CRISPR/Cas9 vector system for tissue-specific gene disruption in zebrafish." Dev Cell 32(6): 756-764.

Araya, C., L. C. Ward, G. C. Girdler and M. Miranda (2016). "Coordinating cell and tissue behavior during zebrafish neural tube morphogenesis." Dev Dyn 245(3): 197-208.

Bachner, D., M. Ahrens, N. Betat, D. Schroder and G. Gross (1999). "Developmental expression analysis of murine autotaxin (ATX)." Mech Dev 84(1-2): 121-125.

Barbayianni, E., E. Kaffe, V. Aidinis and G. Kokotos (2015). "Autotaxin, a secreted lysophospholipase D, as a promising therapeutic target in chronic inflammation and cancer." Prog Lipid Res 58: 76-96.

Bernardos, R. L. and P. A. Raymond (2006). "GFAP transgenic zebrafish." Gene Expr Patterns 6(8): 1007-1013.

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組織專一性破壞自分泌運動因子(Autotaxin)導致斑馬魚神經管發育異常

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