Acoustic Tweezing Cytometry Induces Rapid Initiation of Human Embryonic Stem Cell Differentiation

Tuğba Topal,Zhenzhen Fan,Ninad Kanetkar,Jianping Fu,Xufeng Xue,Paul H Krebsbach,Cheri X Deng,Xiaowei Hong,Joe T Nguyen

doi:10.1038/s41598-018-30939-z

Abstract

Mechanical forces play critical roles in influencing human embryonic stem cell (hESC) fate. However, it remains largely uncharacterized how local mechanical forces influence hESC behavior in vitro. Here, we used an ultrasound (US) technique, acoustic tweezing cytometry (ATC), to apply targeted cyclic subcellular forces to hESCs via integrin-bound microbubbles (MBs). We found that ATC-mediated cyclic forces applied for 30 min to hESCs near the edge of a colony induced immediate global responses throughout the colony, suggesting the importance of cell-cell connection in the mechanoresponsiveness of hESCs to ATC-applied forces. ATC application generated increased contractile force, enhanced calcium activity, as well as decreased expression of pluripotency transcription factors Oct4 and Nanog, leading to rapid initiation of hESC differentiation and characteristic epithelial-mesenchymal transition (EMT) events that depend on focal adhesion kinase (FAK) activation and cytoskeleton (CSK) tension. These results reveal a unique, rapid mechanoresponsiveness and community behavior of hESCs to integrin-targeted cyclic forces.

Highlights

Periodontics, University of California, Los Angeles School of Dentistry, Los Angeles, CA, 90095, USA
Since focal adhesion kinase (FAK) is an important mechanotransductive component downstream of force-activated integrin[24], we examined whether acoustic tweezing cytometry (ATC) application influenced changes in FAK activity in human embryonic stem cell (hESC)
No change was observed in T and Snai[1] gene expression (Fig. S11C). These results demonstrated that 30 min application of ATC-mediated cyclic forces/strains to integrin induced rapid initiation of differentiation and epithelial-mesenchymal transition (EMT) in hESC colonies, in stark contrast with the multiple-day process of differentiation and EMT in hESCs in the presence of morphogenic factors or regulated by matrix rigidity[27,28,29]

Summary

Introduction

Periodontics, University of California, Los Angeles School of Dentistry, Los Angeles, CA, 90095, USA. For single hESCs, cellular contractility increased immediately after ATC application in the cells with attached MBs (Fig. S2A,B), suggesting robust cellular responses to integrin-targeted external forces/strains.

Results

Conclusion