Knouse Lab

The damage or death of terminally differentiated cells in organs that lack active stem cell populations underlies the morbidity of numerous diseases including diabetes, heart attack, stroke, and neurodegeneration as well as many aspects of aging. In these contexts, the differentiated cells permanently exited the cell cycle in the process of differentiation and therefore any remaining functional cells cannot replenish the lost cells. Undoubtedly, a means of enabling terminally differentiated cells to renew and proliferate could alleviate myriad diseases and counteract aging across organ systems.

Although proliferation and differentiation might seem mutually exclusive, the liver—and the hepatocytes responsible for the majority of its mass and function—stand out as notable and informative exceptions. In the uninjured liver, essentially all functional hepatocytes are quiescent and may remain in this non-proliferative state for months to years. However, if liver mass or function is ever compromised, these cells will immediately re-enter the cell cycle and proliferate to regenerate the organ. Although cellular quiescence and liver regeneration are long-studied phenomena, much of our understanding of quiescence is based on artificial cell culture systems and we still do not understand why the liver, but not other organs, has regenerative capacity. Using the mouse liver as a physiologic and tractable system, we are developing and employing tools to probe the molecular regulation of these processes in their native context from the level of single cells to whole organism. Our investigations will first and foremost fill critical gaps in our understanding of the quiescent state and liver regeneration and ultimately uncover new avenues for enabling regeneration across organ systems.

(*) How does an organism sense and respond to organ injury?
Organ damage disrupts both local and systemic homeostasis and the body responds with some combination of inflammation, fibrosis, or repair. In the case of the liver, it is unknown how the organism senses liver injury and initiates regeneration. We are employing organism-wide approaches to identify the tissues and cell types that respond to liver insufficiency, the mechanisms by which insufficiency is sensed, and the physiologic consequences of these responses. By understanding how an organism responds to injury with regeneration, we bring ourselves closer to the ability to modulate injury responses in presently non-regenerative tissues such that injury is met with regeneration rather than fibrosis.

(*) How do quiescent cells retain proliferative capacity?
Quiescent cells, unlike terminally differentiated cells, retain the ability to re-enter the cell cycle after prolonged dormancy.  However, both quiescent and terminally differentiated cells have similar expression of canonical cell cycle genes at baseline. Thus, the functional distinction between quiescence and terminal differentiation lacks a molecular explanation. We are employing a variety of tools to better understand the molecular requirements for the quiescent state and proliferative capacity. This knowledge will lay the foundation not only for attempts to confer proliferative capacity to terminally differentiated cells but also efforts to target quiescent cancer cells that evade traditional chemotherapies.

(*) What genes regulate regenerative capacity?
The ability to understand and manipulate mammalian physiology and disease has long been limited by a lack of tools to perform unbiased, comprehensive genetic dissection directly in a living organism. The lab recently developed genome-wide CRISPR screening in the mouse liver. We are now leveraging this tool to understand the genetic regulation of the liver’s remarkable regenerative capacity. Beyond the liver, we are developing new approaches for transgene delivery to enable genome-wide screening in other organs. With these novel tools, we hope to identify and overcome barriers to regeneration in other tissues.