Web Page Jennifer Barnett Moss 65 mg/dL
Day 0
303 mg/dL
Day 3 125 mg/dL
Day 7
72 mg/dL
Day 14
Regenerating red fluorescent beta cells regrow along green fluorescent vasculature after conditional ablation of
insulin-nitroreductase zebrafish (mg/dL = blood glucose). The panel depicts adult zebrafish pancreas regeneration at
four timepoints using mutiphoton confocal imaging. We use regenerating adult zebrafish to investigate beta cell re-growth
and function after damage to the pancreas. Beta cell damage in humans occurs by several mechanisms and we have
evaluated the effects of genetic or chemical ablation as well as pancreatectomy using zebrafish as models for human
diabetes. We have developed a conditional genetic line where the zebrafish insulin promoter drives bacterial nitroreductase
expression only in beta cells. When the prodrug metronidazole is added to the water, nitroreductase is activated and
zebrafish beta cells stop expressing insulin and mCherry. Blood glucose levels increase 5-7 fold after three days but return to
normal after 2 weeks. We are using this unique animal model to harvest information about gene expression, signaling
mechanisms and metabolic readouts during beta cell regeneration. Small molecules added to the water are being studied for
their ability to potentiate beta cell regeneration in vivo. Information about efficacy, whole body dose and toxicity of small
molecules guides studies in rodent and human islets.
Living zebrafish larvae can be used as a basis for chemical screens. A new
transparent zebrafish line (casper) allows us to image the pancreas in a living animal.
High content imaging can enable the screening of small molecule compounds in larval
zebrafish or in mammalian islets expressing adenoviral constructs. A SPIM lightsheet
microscope to be located in the Light Microscopy Core Facility at Duke has the capacity of
collecting rapid readouts from living, fluorescence-tagged islet cultures or living zebrafish
larvae. The green fluorescent (GFP) vasculature and red fluorescent (mCherry) beta cells
can be followed over time as depicted in this image of a living zebrafish pancreas.
Primary cultures of mammalian islets provide translational platforms for the factors
we discover in zebrafish. Small molecules generating positive signals in zebrafish screens
are being tested in islet cultures for proliferation. The islet shown has a nuclear stain (blue)
and beta cells are labeled with adenovirus-expressed GFP in this multiphoton confocal
image. Although mammalian islets lose blood vessels in culture, we have been able to
detect potentiated beta cell proliferation in the presence of small molecules that affect the
mTOR and PTEN signaling pathways.
Engineered vasculature and islet “hydrogels”. In a collaboration with Jennifer
West’s lab, beta cells (red) derived from freshly isolated, dispersed rat islets are cocultured with primary human endothelial cells and pericytes in the presence of a PEG
hydrogel that was prepared by Duke Biomedical Engineers. The confocal section
depicts immunohistochemical fluorescence of (red) insulin+ rat beta cells and (green)
CD31+ human endothelial cells growing within the 3D hydrogel. Survival as well as
insulin secretion was observed after 7 days. We are developing the use of
vascularized islet hydrogels as optimized platforms for small molecule screens.