Mechanosensation and mechanotransduction is a cellular process that converts a mechanical stimulus to a biochemical response. Typically identified with hair cells in the inner ear and the endothelial lining of the vasculature, mechanosensation has been shown to occur in nearly every cell within an organism. In addition to the role of the actin cystoskeleton and integrin receptors, the mechanism identifies a particular organelle, the primary cilium, as a seat of mechanosensation and mechanotransduction. Ciliopathies are clinical disease states involving the primary cilium, and number over 120. The Ciliopathies form a class of genetic disease whose etiology lies not with dysfunction in single genes and their products, but with dysfunction in an integrated aspect of cellular physiology. One ciliopathy, Autosomal Dominant Polycystic Kidney Disease (ADPKD), is driven by gene products polycystin 1 and polycystin 2 which are localized to the primary cilium. A large body of evidence demonstrates that a physiologically relevant function of the primary cilium in renal epithelia is to respond to the presence (or absence) of fluid flow by initiating multiple signaling cascades, one of which involves nuclear translocation of the cytoplasmic tail of polycystin 2. The current picture of ADPKD progression thus connects disease to both gene product and fluid flow via the primary cilium. Our long term goal is to study the role of the primary cilium in transducing mechanical stresses into a biological response. We use polarized, terminally differentiated monolayers of cultured mouse renal epithelial cells as our disease model. As our first aim, we will measure the bending modulus of the primary cilium using optical trapping to apply a known acute stimulus to the cilium while imaging the resultant deformation. Simultaneous trapping and Calcium imaging will be performed to ensure physiological relevance. As our second aim, we will examine the hypothesis that chronic steady and unsteady flows provoke different epithelial cell responses. We will correlate ADPKD biomarkers (e.g. polycystin2, STAT6) with precisely specified flow properties. Well-characterized physiological readouts (e.g. ENaC activity, cilium length) will also be used to correlate flow with cell phenotype. Detailed properties of the flow will be chosen in a rational manner based on the Peclet and Strouhal numbers. Together, these approaches will clarify and quantify how the primary cilium senses the state of fluid flow to act as a complex signaling center, which will have broad impact to multiple fields of biomedical research.
Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a progressive disease, typically appearing in the 5th decade of life and is one of the most common monogenetic inherited human diseases, affecting approximately 600,000 people in the United States. Considerable research using mouse models for the progression of ADPKD indicates that pathology correlates with defective primary cilia. It is also thought that a physiological role of the primary cilium is to sense fluid flow. The experiments proposed here will examine a hypothesis that connects these two lines of study- that environmental factors targeting the primary cilium, specifically fluid flow, is an important modifier of ADPKD progression.
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