A stretching device for imaging real-time molecular dynamics of live cells adhering to elastic membranes on inverted microscopes during the entire process of the stretch

Abstract

We present a device for stretching cells adhering to elastic membranes in equiaxial or uniaxial mode, meanwhile allowing real-time imaging of molecular dynamics of live cells at high resolution on an inverted microscope during the entire process of the stretch. We obtained high-resolution images of stress fibers at each stage of the stretch, and found that stress fibers were shortened after one stretching cycle. We, for the first time, captured real-time images of the process of stress fiber disassembly during stretching. Several adjacent stress fibers appeared to reassemble into a single one after stretching. All these indicated that mechanical stretching played important roles in the rearrangement of actin filaments. This device will be especially useful in studies of the molecular dynamics in the process of mechanotransduction. The device is fabricated on a glass slide through a simple procedure and is adaptable to most ordinary laboratories.

Mechanical Analysis of a Pneumatically Actuated Concentric Double-Shell Structure for Cell Stretching

“…Therefore, the proposed model can be used for estimating the loads needed to achieve a certain strain. Furthermore, Wang et al carried out the NIH/3T3 (National Institute of Health/3-day transfer, inoculum 3 × 10 5 cells) fibroblasts stimulation by applying high-level equiaxial strains (12%-27%) to the cells [8]. Moreover, they employed an inverted microscope (Leica DM16000, Leica Microsystems, Wetzlar, Germany) to view the cells, and performed the scanning to obtain clear cell images [8].…”

“…Furthermore, Wang et al carried out the NIH/3T3 (National Institute of Health/3-day transfer, inoculum 3 × 10 5 cells) fibroblasts stimulation by applying high-level equiaxial strains (12%-27%) to the cells [8]. Moreover, they employed an inverted microscope (Leica DM16000, Leica Microsystems, Wetzlar, Germany) to view the cells, and performed the scanning to obtain clear cell images [8]. In the device developed by Kreutzer et al [6], according to our prediction, to achieve such high-level strain, a loading up to 100 kPa is needed, which will result in an out-of-plane displacement as high as 895 μm.…”

“…The mechanical strain in this device was achieved by out-of-plane distension of the substrate membrane. Wang et al developed a cell-stretching device, which employed vacuum pressure to achieve either equiaxial or uniaxial strain field in the substrate membrane [8]. In a recent study, the authors of the present paper developed a pneumatically actuated device with concentric double-shell structure for cell stretching and demonstrated the applicability of the developed stretching device for differentiation of human pluripotent stem cells into cardiomyocytes in long-term cell experiments [6].…”

“…In the Matlab script of the computational model, Young's modulus E was determined as a range of [0. 8,4] MPa, and the model was revolved by COMSOL pre-processor. Moreover, to calculate the in-plane strain, we needed to convert the displacements given in the Cartesian coordinates (e.g., in Figure 3d,e) into the cylindrical coordinates as presented in Equation (17): 2 2 ε 100%…”

“…Recent studies of cardiac mechanobiology have shown that to a certain extent mechanical stimulation enhances the growth of stem-cell-derived cardiomyocytes in vitro [1][2][3][4][5][6][7][8]. Thus, different types of devices have emerged for the purpose of mechanical stimulation for stem cells, such as devices that use out-of-plane circular substrate distension, in-plane substrate stretching, or fluid shear stress (FSS) [7,9,10].…”