Studying the Effects of Confinement on Cell Division

On Microfluidic Future I like reviewing advancements in therapeutic or diagnostic devices because I’m really drawn to those areas of research. Every once in a while, however, I take interest in research for the sake for knowledge, like the Root Chip. I recently came across an article from Dino Di Carlo of UCLA that describes a microfluidic device used to study cancer cells. The article, “Increased Asymmetric and Multi-Daughter Cell Division in Mechanically Confined Microenvironments” appeared in PLoS ONE, which is an open access journal (very cool!).

Specifically, Di Carlo’s device is used to study the effects of the mechanical environment on cancer cells during division. It’s commonly known that the course of the cell cycle is affected by soluble factors, but the cell’s mechanical interaction with the environment also affects its morphology, differentiation and cell cycle. Changes in confinement and substrate elasticity were tested using the HeLa cervical cancer cell line in this study. The authors looked for several deviations from standard cell division including delayed mitosis, multi-daughter mitosis events, unevenly sized daughter cells and induction of cell death.

Device Design

A. The device features posts to confine cells with continuous perfusion. B. The device is deformable, bringing the two layers into contact. C. The device has variable elasticity and confinement height.

A. The device features posts to confine cells with continuous perfusion. B. The device is deformable, bringing the two layers into contact. C. The device has variable elasticity and confinement height.

Di Carlo’s device has a bottom layer and an elevated PDMS layer supported by posts with varying height for control over cell confinement. In a relaxed state, there is a 15 µm clearance between the posts and bottom layer. When pressure is applied to the device, the two layers meet which confines the cells between posts and reduces the clearance to 3 µm or 7 µm. Additionally, the top layer has an elasticity of 130 KPa or 1 MPa. The device is designed to allow media to flow throughout all the confining chambers, eliminating the possibility of cell death due to a toxic environment.

Results

Induced multi-daughter division resulted in 3, 4 and even 5 daughters

Induced multi-daughter division resulted in 3, 4 and even 5 daughters

In an unconfined environment, a HeLa cell would normally ball up into a sphere during mitosis, which would take no longer than 140 minutes. But with increased confinement and stiffness, the authors witnessed multi-daughter mitosis (one cell dividing into three or four daughter cells), unbalanced daughter sizes, prolonged mitosis and cell death. Resulting control cells from division would often be spheres with a diameter of 20 µm, while the confined cells would be highly asymmetric with diameters 40-80 µm. Increases in stiffness and confinement generally increased the odds of abnormal cell division, with some clear observed patterns. Under low compression of 7µm and stiffness of 130 KPa, 90% of multi-daughter divisions resulted in three cells. When a stiffness of 1 MPa was applied to the same low compression, 85% of multi-daughter divisions resulted in four cells. The authors believe that the cells aren’t able to effectively deform the stiffer substrate and are limited in how spherical they can be before mitosis. The confined shape may also affect chromosomes lining up at the metaphase plane(s), resulting in a bias toward multi-daughter divisions. The multi-daughter cell divisions can produce viable cells, which subsequently can undergo their own multi-daughter division, and also generate daughters that re-fuse after division.

The authors also hypothesized that when the cells are forced to divide in a discoid shape, signaling and regulation may be affected. Diffusion or active transport of signals would take much longer to traverse the large cross-section of the cell, and the force of cytoskeletal elements might be diminished across the same large distance.

Discussion

This work has produced some findings that may not be totally surprising, but are definitely peculiar. A follow-up to the findings generated here would surely add to the increasing knowledge base of cancer cell behavior. In its current form, this information wouldn’t lead to any new treatments, although under high confinement 70% of cell cycles resulted in cell death, which holds potential in therapeutic applications. Studying diseases help us learn more about healthy cells because we can see what goes wrong when specific elements fail, but I’m also interested in seeing how healthy cells react under the same mechanical conditions. The microfluidic device itself also has potential beyond the study of cellular life cycles: One area in particular includes investigating the effects of mechanical strain in osteocyte and chondrocyte differentiation.

Reference:

ResearchBlogging.org

Henry Tat Kwong Tse, Westbrook McConnell Weaver, & Dino Di Carlo (2012). Increased Asymmetric and Multi-Daughter Cell Division in Mechanically Confined Microenvironments PLoS ONE, 7 (6)