Person:

Keliher, Edmund J.

The ability to isolate pure pancreatic ß-cells would greatly aid multiple areas of diabetes research. We developed a fluorescent exendin-4-like neopeptide conjugate for the rapid purification and isolation of functional mouse pancreatic β-cells. By targeting the glucagon-like peptide-1 receptor with the fluorescent conjugate, β-cells could be quickly isolated by flow cytometry and were >99% insulin positive. These studies were confirmed by immunostaining, microscopy and gene expression profiling on isolated cells. Gene expression profiling studies of cytofluorometrically sorted β-cells from 4 and 12 week old NOD mice provided new insights into the genetic programs at play of different stages of type-1 diabetes development. The described isolation method should have broad applicability to the β-cell field.

Acute bacterial endocarditis is a rapid, difficult to manage and frequently lethal disease. Even potent antibiotics often cannot efficiently kill Staphylococcus aureus (S. aureus) that colonizes the heart’s valves. S. aureus relies on virulence factors to evade therapeutics and the host’s immune response. For instance, S. aureus usurps the host’s clotting system by activating circulating prothrombin with staphylocoagulase and von Willebrand factor-binding protein. An insoluble fibrin barrier then forms around the bacterial colony, shielding the pathogen from immune cell clearance. Targeting virulence factors may provide new avenues to better diagnose and treat endocarditis. To tap into this unused therapeutic opportunity, we co-developed therapeutics and multimodal molecular imaging to probe the host-pathogen interface. We introduced and validated a family of small-molecule optical and PET reporters targeting active thrombin in the fibrin-rich environment of bacterial colonies. The imaging agents, based on the clinical thrombin inhibitor dabigatran, bound to heart valve vegetations in mice. Aortic or tricuspid S. aureus endocarditis in piglets was successfully imaged with clinical PET/MRI. We employed optical imaging to monitor therapy with antibodies neutralizing staphylocoagulase and von Willebrand factor binding protein in mice with S. aureus endocarditis. This treatment deactivated bacterial defenses against innate immune cells, decreased in vivo imaging signal and improved survival. Our data map a route towards adjuvant immunotherapy for endocarditis and provide efficient tools to monitor this drug class for infectious diseases.