Gastrointestinal physiology and pharmacology Cystic fibrosis, epithelial transport and stem cell biology. Dr. Clarke’s laboratory investigates abnormalities of acid-base transporters in cystic fibrosis (CF) and other genetic diseases as they relate to intestinal stem cell biology (ISCs) and differentiation/function of secretory cells. Studies of mice with gene-targeted deletion of CFTR (the cystic fibrosis gene) and other acid-base transporters such Na+/H+ and Cl–/HCO3– exchangers are performed using mice or regenerating murine or human intestinal organoids grown in 3D gel culture. As an anion channel, CFTR is responsible for epithelial fluid secretion which provides the proper environment for nutrient absorption and maintains normal viscoelastic properties of mucus. Functional activity of acid-base or nutrient transporters is measured in real time using fluorescence confocal or conventional microscopy and electrophysiological methods. Gene or protein expression is measured using quantitative real-time PCR, immunoblots, immunofluorescence and laser capture microdissection. Currently, three major projects in the laboratory are funded by NIDDK, the Cystic Fibrosis Foundation (CFF) or CFF Therapeutics, Inc. (CFFT). NIDDK-funded projects investigate the role of CFTR, a cAMP-regulated Cl– and HCO3– ion channel, in down-regulating cell cycle dynamics and Wnt/b-catenin signaling in intestinal stem cells (ISCs). Loss of this regulation in CF results in hyperproliferation of ISCs which forms the platform for a six-fold increase in the incidence of gastrointestinal cancer in young CF patients. A second project investigates the acid-base transporters expressed in ISCs that determine intracellular pH (pHi). Manipulation of pHi is used to control ISC proliferation to offset “bystander” damage to ISCs resulting from therapeutic doses of chemotherapeutic reagents and radiation during cancer treatment. Two CFF-funded projects investigate goblet cell (mucus secreting) dysfunction in the CF intestine, i.e., mucoviscidosis. The goals of the first project are to determine the role of CFTR in goblet cell dysfunction and evaluate the impact of abnormal goblet cell antigen presentation to dendritic cells on CF immune status. The goal of the second project is produce a CF mouse that expresses a pH-sensitive fluorescent mucin to evaluate the abnormalities of mucus pH during processing and release. To facilitate the translational potential of the above CFF projects, a CFFT-funded project has the goal to develop a human CFTR “rescue” mouse model in which murine CFTR is replaced by the human ortholog of the gene. This humanized CFTR mouse will enable pharmacological testing and gene-editing strategies for correcting defective function of CFTR in CF patients or to combat diarrheal diseases in humans and animals that are caused by enterotoxigenic bacteria such a Vibrio cholerae.