第一型膠原蛋白在斑馬魚表皮發育中扮演之角色

膠原蛋白，為細胞外間質中最主要的成分。不僅參與結構組織的形成，也調控細胞極化、固著、黏附、遷移和分化這類型的生理過程。魚類的第一型膠原蛋白已被證實如同哺乳類動物一般，為皮膚中最主要的膠原蛋白。過去在研究表皮發育機制中，除了指出真皮中纖維母細胞合成膠原蛋白外，是否還有哪一類型表皮細胞參與合成，目前鮮少資料提及也尚未有證據證實。到目前為止，關於第一型膠原蛋白在魚類皮膚中的基因表現和功能尚未完全清晰。本實驗目的為闡明第一型膠原蛋白在斑馬魚胚胎表皮上所扮演的角色及功能。斑馬魚的第一型膠原蛋白有三種isoforms，分別為alpha 1a 、alpha 1b與alpha 2。我們以斑馬魚作為實驗材料，利用RT-PCR的技術檢測發現，col1a1a mRNA大量表現在皮膚中並且在受精後2小時的階段就有表現產生，另外兩種isoforms則是受精後8小時才有基因表現的訊號產生。定位雜交(in situ hybridization)技術顯示三種isoforms在胚胎表皮上有著類似的分布型態。利用反義核酸(morpholino oligos)抑制三種第一型膠原蛋白isoforms短肽的合成後，發現表皮細胞中的表皮幹細胞(ESCs)、離子細胞、角質細胞和黏液細胞數量都有顯著性下降。另外也觀測到col1a1a mRNA與表皮幹細胞的免疫染色共定位於一起，第一型膠原蛋白的抗體免疫染色也顯示第一型膠原蛋白表現在黏液細胞中。在此研究中，我們證明了第一型膠原蛋白於早期的胚胎發育時期表現，並且通過表皮幹細胞來生成。而根據第一型膠原蛋白表現在黏液細胞中的結果，我們推測或許第一型膠原蛋白也參與了皮膚上的防禦機制。綜合以上所述，這篇研究對於第一型膠原蛋白在皮膚的發育中所扮演的角色提供了不同的理解和新穎的知識。

關鍵字

斑馬魚；第一型膠原蛋白；表皮幹細胞；黏液細胞；表皮發育

並列摘要

Extracellular matrix majorly composed of collagen. Collagens not only function as structural component but also participate in many physiological processes such as cell polarity, anchorage, adhesion, migration, and differentiation. Similar to mammals, teleosts collagen type 1 has been identified as major collagen in skin. Previous studies linked collagen type 1 in epithelial development but none can provide the specific mechanism behind such as the type of epidermal cell synthesizing collagen type 1 besides fibroblast is yet to be unidentified. So far, the knowledge of collagen type 1 isoforms gene expression and function in fish skin is not completely clear. The purpose of the present study is to examine the role of collagen type 1 in zebrafish embryos epidermal development. There are 3 isoforms of collagen type 1 in genome database, the collagen type 1 alpha 1a (col1a1a), alpha 1b (col1a1b) and alpha 2 (col1a2), all were detected by RT-PCR and cloned. Tissue scan indicated that col1a1a mRNA expressed abundantly in the skin and the expression begins to be endogenously transcribed from 2 h post fertilization (hpf) followed by other isoforms from 8 hpf. Expression of collagen type 1 isoform mRNAs showed similar spatial patterns via in situ hybridization. The knockdown of col1a1a, col1a1b, and col1a2 results showed that all of the ESCs, ionocytes, keratinocytes, and mucous cells number were decreased. mRNA in situ hybridization of col1a1a co-localized with ESC and immunostaining showed collagen type 1 is present only in agr2/mucous cells. The presence of collagen in ESCs and keratinocytes was not established in this study due to the limitation of commercial antibodies used. Nevertheless, we demonstrated that collagen type 1 is present very early during the embryogenesis and is being synthesized by the ESCs, clarifying the timing and location of expression. The presence of collagen in mucous cells may indicate possible role in this defense mechanism of the skin. Hence, this work provides new knowledge and additional understanding of the collagen type 1 role in skin development.

並列關鍵字

zebrafish ； collagen type 1 ； epidermal stem cell (ESC) ； mucous cells ； epidermal development

參考文獻

Whitfield, T, T., White, S. J. (2011). Functional and developmental expression of a zebrafish Kir1. 1 (ROMK) potassium channel homologue Kcnj1. The Journal of Physiology, 589(6), 1489-1503.

Alberts, B., Johnson, A., Lewis, J., Raff, M., Roberts, K., Walter, P, (2008). Molecular

Zebrafish ΔNp63 Is a Direct Target of Bmp Signaling and Encodes a Transcriptional Repressor Blocking Neural Specification in the Ventral Ectoderm. Developmental cell, 2(5), 617-627.

Ehlers-Danlos syndromes: revised nosology, Villefranche, 1997.American journal of medical genetics, 77(1), 31-37.

Bottcher-Haberzeth, S., Biedermann, T., Reichmann, E. (2010). Tissue engineering of

國際替代計量

第一型膠原蛋白在斑馬魚表皮發育中扮演之角色

全文下載

主題瀏覽