(Left) Rendered engineering image of the bioreactor used in Professor Ruberti’s mechanobiology experiments. (Right) Sequence of light microscope images of the behavior of primary human corneal fibroblasts cultured in the device. A) Initial seeding; B) confluent single layer of cells forms; C) second layer of cells grows over the top of the first layer; D) cell sheet sliding occurs where the second layer of fibroblasts slides over the first layer.
The Fox Center for Vision Restoration organizes an exciting lecture series focusing on ocular regeneration and new therapies.
Distinguished national and international speakers present their innovative and multidisciplinary approaches to finding cures for vision impairment. The objective of this lecture series is to accelerate research through knowledge sharing, partnership building and out of the box thinking.
This lecture series should be of interest to: clinicians with an interest in ophthalmology; scientists and engineers interested in tissue engineering, cellular therapies and assistive technologies; students, postdoctoral fellows, residents and research staff.
Live Imaging of the Mechanobiology of Matrix Production by Corneal Fibroblasts
Jeff Ruberti, PhD
Director of the Extracellular Matrix Engineering Research Laboratory
Dr. Ruberti received his BSE in Biomedical Engineering from Tulane University in 1986 and his Ph.D. from Tulane in 1998. In the intervening years (‘86-‘91) he worked at Sikorsky aircraft (helicopter electrical systems engineering), at the Southwest Foundation for Biomedical Research (nenonatal pulmonary function research) and at Hamilton Standard (space station life support systems engineering).
His PhD thesis employed mathematical and experimental methods to extract the membrane transport parameters from the corneal endothelium. His post-doctoral research was performed at MIT in the Fluid Mechanics Laboratory and continued at Northwestern University in the Department of Biomedical Engineering where he used Quick Freeze Deep Etch imaging to capture the age-related deposition of lipids between the retina and its blood supply (possibly leading to Age-Related Macular Degeneration).
He returned to industry in 2001 as an associate consultant at Cambridge Polymer Group where he began work on controlling the organization of collagen and controlling the structure of physically associated polymeric gels. He has been at Northeastern University in the Mechanical and Industrial Engineering Department since the Fall of 2004 where he is a tenured Associate Professor, Chair of the Bioengineering PhD program and director of the Extracellular Matrix Engineering Research Laboratory.
Dr. Ruberti holds seven issued patents (with 4 patents pending) and has been published on numerous topics including membrane transport, experimental blood flow modeling, nanobubble evolution, implant wear patterns, corneal biomechanics, corneal tissue engineering and collagen mechanochemistry. His research has been funded by the NIH (NIAMS, NEI and NIBIB), NSF and the US DOD.
It was recognized over one hundred years ago by scientists such as Julius Wolff and Wilhelm Roux that mechanical force plays an important role in the shaping of biological structure. In his remarkable tome published in 1917, Darcy Thompson wrote about biological systems in the following way “Cell and tissue, shell and bone, leaf and flower, are so many portions of matter, and it is in obedience to the laws of physics that their particles have been moved, moulded and conformed….” .
In spite of an early and auspicious start, while biologists explored the role of genes and molecular signaling pathways as determinants of biological form, function and disease, the field of mechanobiology lay fallow for the better part of a century. Recently however, there has been a resurgence in the investigation of the influence of mechanics on the behavior of living systems with the number of publications increasing exponentially since the beginning of this decade. Concurrent with this increase in scientific enquiry, a series of new tools has been developed (some in our lab) which permits the precise mechanical probing of tissue, cells, molecular aggregates, single molecules and systems of interacting molecules. What we are beginning to learn is that there are numerous exquisitely force-sensitive mechanomic systems of molecules (e.g. vinculin and talin association in focal adhesion formation) which are strongly implicated in the development of biological form and function.
While the ocular globe does not carry a significant load under normal adult conditions, we do know that mechanical force is critical to the proper development and growth of the entire ocular tunic, including the cornea. In a remarkable experiment, Neath et al (1991) found that intentional reduction of the intraocular pressure in embryonic chick eyes dramatically reduces the growth of the cornea and the entire ocular globe.
In this presentation, I will describe our efforts to examine the role of externally applied force on the behavior of a primary human corneal fibroblast (PHCF) culture system which is known to produce stroma-like extracellular matrix. To perform the experimental series, we first developed a mechanobioreactor with permits direct observation of cell culture dynamics for extended periods (two weeks +). Using the device, we are able to follow the behavior of the PHCF culture in realtime, from initial seeding to matrix production, while applying a fixed uniaxial tensile load (6% strain) to the substrate on which the cells were grown.
The results of this investigation suggest that applied tension influences the direction of PHCF alignment over long length scales (45 degrees to the applied strain) relative to unloaded controls. We also found that the synthesized matrix (fibronectin and collagen) follows the cell orientation and direction. Further, complex population dynamics were observed which included differing phases of cell velocity and migration direction over the culture period.
We conclude from the work, that mechanical strain is a potentially important “signal” which has the ability to control the behavior of human primary cells both before and during the production of matrix. The mechanobioreactor used in this investigation is an important tool which should enable scientists to both probe and evaluate the effects of mechanics on populations of interacting cells. It is becoming increasingly clear that ignoring mechanics in biological systems may invite misinterpretation of observed effects.
Location and Address
Eye and Ear Boardroom
Eye and Ear Institute