探索機械拉伸對上皮細胞分裂的調控潛力：以 MDCK 細胞為模型

Abstract

在2022年，中央研究院細胞生物研究所陳振輝教授實驗室於斑馬魚幼魚的表皮細胞中發現了一種特殊的細胞分裂現象，稱為無合成分裂 (asynthetic fission)。這一現象不同於傳統的有絲分裂，因為細胞在分裂過程中不經過 DNA 合成 (S期)，而是直接進入分裂階段。研究人員提出，這種現象可能與斑馬魚幼魚在其生長階段中，皮膚細胞所承受的巨大張力有關，這樣的張力可能促使細胞發生非典型的分裂過程。本研究旨在探討是否可以在哺乳類表皮細胞中觀察到類似的無合成分裂現象。我們選擇了表現 H2B-mCherry的MDCK (Madin-Darby Canine Kidney)細胞株並利用自製的拉伸器(Fig. 1a)來模擬張力。該拉伸器通過夾緊PDMS薄膜兩端並控制插銷寬度來施加拉伸，拉伸程度大約為30-40%。我們通過共軛焦顯微鏡，使用10倍物鏡，每10分鐘拍攝一次1024x1024像素的影像，每組實驗時間為兩小時。雖然在實驗中觀察到了多次分裂事件，但目前尚未能夠確定 MDCK 細胞發生無合成分裂。依賴拉伸配合EdU staining來推測細胞所處的分裂週期階段也尚未找到MDCK無合成分裂的證據，且在拉伸條件的環境設置下如何清晰地觀察細胞分裂也是一大挑戰。未來可以探索更多細胞週期相關的 MDCK cell line 進行更細緻的實驗。此外，拉伸儀器的精度和薄膜的拉伸速度也將成為未來優化實驗設計的重點，以便更精確地模擬細胞在自然條件下的反應。

In 2022, a research team led by Professor Chen Chen-Hui at the Institute of Cellular and Organismic Biology, Academia Sinica, reported a novel mode of cell division termed “asynthetic fission” in the epidermal cells of zebrafish larvae. Unlike conventional mitosis, this type of division bypasses the DNA synthesis (S) phase and proceeds directly to the mitotic stage. The researchers proposed that this process may be driven by the substantial mechanical tension experienced by epidermal cells during the larval growth phase, which may induce such non-typical divisions. The aim of our study is to investigate whether a similar asynthetic fission phenomenon can be observed in mammalian epithelial cells. We utilized a H2B-mCherry-expressing MDCK (Madin-Darby Canine Kidney) cell line and applied mechanical tension using a custom-built stretching device (Fig. 1a). This device operates by clamping a PDMS membrane at both ends and varying the plug distance to induce stretching, achieving an estimated strain of approximately 30–40%. Imaging was performed using a confocal microscope with a 10× objective lens, acquiring 1024×1024 pixels images every 10 minutes over a two-hour observation period. Although multiple cell division events were observed under these conditions, no definitive evidence of asynthetic fission has been identified in MDCK cells. Attempts to infer the cell cycle phase based on EdU staining following mechanical stretching remain inconclusive. Moreover, clearly visualizing cell division events under stretching conditions continues to pose significant technical challenges. Future studies may benefit from the use of MDCK cell lines with fluorescently labeled cell cycle markers to enable more precise analysis. Additionally, improving the mechanical resolution of the stretching device and optimizing the membrane extension rate will be essential for better mimicking physiological mechanical stimuli and enhancing experimental reproducibility.