壓電刺激對人類間質幹細胞增生、分化與聚合的影響

The effect of piezoelectric stimulation on human mesenchymal stem cells: proliferation, differentiation and aggregation

黃文顥(Wen-Hao Hwang)

指導教授 ： 王兆麟

國立臺灣大學/工學院/醫學工程學研究所/碩士(2020年)

https://doi.org/10.6342/NTU202001926

摘要

不管是經過氣管手術或者關節受損，軟骨組織修復的速度都非常緩慢。在再生醫學的領域當中，有許多研究都嘗試提高幹細胞的增生速度與軟骨分化速度，希望藉此解決軟骨修復難題，本研究亦為此目標對人類的骨髓間質幹細胞進行研究。
實驗使用自製超音波驅動的壓電刺激系統，探討壓電刺激對幹細胞增生數量、軟骨分化程度以及細胞聚合形態的影響。使用西方墨點法與免疫螢光染色對刺激後的細胞內部phospho-ERK與SOX9蛋白質含量進行分析，並以細胞極性與細胞核β-catenin濃度探討壓電刺激與典型和非典型Wnt訊息傳遞路徑之間的關係。
本研究的結果顯示壓電刺激造成幹細胞內部phospho-ERK提升並有效地加快細胞增生。在LIC與LIC-Pro兩種刺激裝置不同的超音波能量層級之下 (約0.6 mW/cm^2與18 mW/cm^2) 皆促進細胞聚合與軟骨分化，且無需使用軟骨分化培養液。在壓電刺激後活化了細胞中典型Wnt訊息傳遞路徑與非典型Wnt細胞平面極性路徑，使細胞產生極性並依照電場較高的區域聚合，隨著軟骨標記蛋白SOX9的提升，聚合的細胞進行軟骨分化。
骨骼中存在許多膠原蛋白使其具有壓電的特性，然而其與體內軟骨形成的關聯性尚不明確。以本研究中的結果推測，壓電環境很有可能在人體內形成軟骨的過程中起作用。在體內產生壓電效應後，幹細胞向著產生電場的區域遷移與聚合，接著在軟骨分化因子的作用下進行分化。
本研究已經對人類骨髓間質幹細胞進行增生、分化與聚合的分段實驗，但並不確定從細胞低密度時持續培養至軟骨分化，壓電刺激是否能夠整合並貫穿在所有過程。各階段細胞內部其他蛋白質變化以及實際在臨床上的應用為未來主要研究方向。

關鍵字

壓電刺激 ； 微能量超音波 ； 人類骨髓間質幹細胞 ； 增生 ； 軟骨分化 ； 聚合

並列摘要

Cartilage repair is a growing market due to aging population. Among various approaches, mesenchymal stem cells have been considered as a promising solution for cartilage tissue engineering and regeneration as chondrocytes extracted from aging patients cannot proliferate easily. The objective of the study is to investigate the effect of piezoelectric stimulation (PE) on bone marrow-derived human mesenchymal stem cells (hMSCs) with respect to proliferation and differentiation.
The results revealed that PE promoted the proliferation of hMSCs possibly through elevation of phospho-ERK and its translocation into nucleus. Furthermore, PE can induce chondrogenesis of hMSCs even without the use of differentiation medium. In particular, PE caused clustering of hMSCs in a pattern highly correlated with the charge distribution on the quartz plate. By analyzing the orientation of primary cilia with respect to cells, we found both ultrasound and piezoelectric can induce cell polarity but required differentiation media to exhibit a visible representation.
Although piezoelectricity is abundantly available in bones, the mechanism between piezoelectricity and cartilage formation is unclear. The results in the study suggest that piezoelectricity might play a role in hMSC proliferation and chondrogenesis. Furthermore, our results indicate that piezoelectric stimulation might offer a cheaper and faster way to induce chondrogenesis from hMSCs.

並列關鍵字

piezoelectric stimulation ； very-low-intensity ultrasound ； bone marrow-derived human mesenchymal stem cells ； proliferation ； chondrogenesis ； aggregation

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