Researchers clear the way for well-rounded view of cellular defects

Cellular dynamics research studies living cells and their life, death, division, and multiplication. Over the past several years, Nain has taken many journeys down the microscopic roads where cells live. His past work has analyzed how cells move and even included projects with colleagues to measure cell forces and nucleus shapes and to electrify cells and observe how they heal.

A cell divided is how we stand

His latest collaboration investigates how cells divide, particularly in the fibrous environment of living tissue. Cells are typically studied in a flat environment, and the difference between flat and fibrous landscapes opens new windows into the behavior of cells and the diseases that impact them. The findings were published in the Proceedings of the National Academy of Sciences on Feb. 27. The work received funding from the National Science Foundation and support from both the Virginia Tech Institute for Critical Technology and Applied Science and the Virginia Tech Macromolecules Innovation Institute.

Cell division, called mitosis, is essential for developmental, repair, and disease biology. A cell, at its most fundamental level, duplicates its chromosomes, which are then separated and distributed equally between two daughter cells, each with its own complete set of genetic information. As new cells perform the same function over and over, they form organs, heal wounds, and replace dead cells, sustaining the cycle of healthy tissues and organs.

But cell division doesn't always happen this smoothly. Sometimes, cells divide unevenly, or chromosomes can become unevenly split. When those misfires occur, the resulting cell will continue to duplicate copies of its faulty self, creating genetic defects that could cause widespread problems in a living body. These abnormalities account for many prenatal birth defects and can contribute to the origins of cancer.

Better understanding cellular mitosis increases our chances of diagnosing, treating, and preventing those mitotic defects. Nain's discovery puts valuable information in the hands of researchers by painting a complete picture of what's going on at the cellular level within the body's fibrous environment.

Movement, multiplication, and division

At the microscopic level, cells move by way of an extracellular matrix (ECM), a three-dimensional lattice of organic material that provides the framework for cells to form organs by underlaying a strong foundation that holds them together.

Nain'sfoundational research focuses on re-creating and studying that lattice, and his team's past studies on cellular motion have shown how cells travel along it. For a single fiber, a cell pulls itself along at each end, walking the fiber like a tightrope. Two fibers running parallel allow the cell to double those connections.

A dividing cell also makes use of the fibers around it. For a single fiber, each end of the cell adheres and pulls to create the division. If a cell is in an environment with multiple fibers, it will likely attach to those as well. The ECM may cross above and below the cell, providing a three-dimensional web onto which cells connect.

The number of fibers available for cells to attach to affects the timing of cell division and the types of defects a cell may produce. Cells take longer to divide on single fibers, and mitotic errors change with more attachments, creating a complex picture of the myriad ways in which a cell might fail.

This discovery affects future research because the complex view of cell division errors has not been previously investigated in fibrous environments.

A new dimension for research

"Cellular biology has predominantly been studied on a Petri dish, which is a flat, two-dimensional surface," said Nain. "Flat 2D is limited in physiological output because there are very few places in the body where the environment can be considered two-dimensional."

The team found that observing cells in the 3D environment of an ECM yielded new results beyond the capability of 2D Petri dishes. In this work, the team asked a central question: how does the shape of a cell affect its dividing behavior?

Cell shape depends on how a cell adheres to underlying substrates. For example, on a flat, two-dimensional Petri dish, a cell resembles a pancake. In a fibrous environment such as an ECM, shapes range from elongated aerofoils to kites, depending on the number of fibers and their architecture. While a cell might adhere above and below the fiber plane on suspended fibers, a flat surface causes the cell to flatten out and form connections outward. That flattening causes the cell to behave differently when it balls up and undergoes division.

Schematic of a rounded cell body attached to a single fiber and held by actin retraction fiber cables (red) connecting adhesion clusters (green) with the cell cortex (blue). Image courtesy of Amrinder Nain.

As a rounded cell body divides, it's held in place by organic cables that attach the cell body, or cortex, to the fibers. On single fibers, near-perfect spherical cell bodies are held in place by two sets of cables, giving maximum freedom for the rounded cell body to move in 3D. As the number of fibers in the lattice increases, so does the number of places to which a cell can adhere. This results in multiple cable complexes that limit 3D movement of the rounded cell body.

This simple mechanical effect highlights the significant difference between the Petri dish and the ECM. On a Petri dish, monopolar spindle defects, which represent incomplete spindle pole (or centrosome) separation, do not often occur. However, when a cell is in a single-fiber environment with two cable attachment sites, monopolar spindle defects increase.

These results turn cell study quite literally on its head: in the environment of a Petri dish, some defects that occur during cellular mitosis cannot happen in the same way as they do in a living body.

"While bipolar division, the most common and error-free division mode, dominates division outcomes in fibrous environments, our work shows a switch in monopolar and multipolar defects by changing the number of fibers cells attach to," said Nain. "It offers a glimpse into how cell division might occur in actual living tissues."

Nain hopes that the fresh perspective provided by this foundational experimental-computational work will yield insights on how to treat disease and genetic disorders.

"With fiber networks, we provide more detail on a comprehensive in vivo picture, filling in some missing information and using our multi-disciplinary approach, we would like to ask some precise questions in mitotic biology as we move forward," he said.

The multi-disciplinary team assembled for this project comprises leading experts, including cell division biologist Jennifer DeLuca, a professor at Colorado State University; theoretician and biophysicist Nir Gov, a professor at the Weizmann Institute of Science, Israel; and theoretician and computational expert Raja Paul, a professor at the Indian Association for the Cultivation of Science (IACS), India. The first author for the publication was Aniket Jana, now a postdoctoral fellow at the University of Maryland, College Park. Other student members involved in this study include Hoanan Zhang and Atharva Agashe from the Virginia Tech Department of Mechanical Engineering, Ji Wang from the Virginia Tech Department of Biomedical Engineering and Mechanics, and Apurba Sarkar from IACS, India.