Abstract
Aging or injury in Drosophila intestine promotes intestinal stem cell (ISC) proliferation and enteroblast (EB) differentiation. However, the manner the local physiology couples with dynamic EB differentiation assessed by traditional lineage tracing method is still vague. Therefore, we developed a 3D-printed platform “FlyVAB” for intravital imaging strategy that enables the visualization of the Drosophila posterior midgut at a single cell level across the ventral abdomen cuticle. Using ISCs in young and healthy midgut and enteroendocrine cells in age-associated hyperplastic midgut as reference coordinates, we traced ISC-EB-enterocyte lineages with Notch signaling reporter for multiple days. Our results reveal a “differentiation-poised” EB status correlated with slow ISC divisions and a “differentiation-activated” EB status correlated with ISC hyperplasia and rapid EB to enterocyte differentiation. Our FlyVAB imaging strategy opens the door to long-time intravital imaging of intestinal epithelium.
Highlights
Aging or injury in Drosophila intestine promotes intestinal stem cell (ISC) proliferation and enteroblast (EB) differentiation
Our results suggest that the FlyVAB intravital imaging strategy did not affect flies’ digestion and had a small adverse physiologic effect on flies’ survival, indicating that the images of the flies acquired from the FlyVAB device represent the in vivo conditions
These results suggested that Notch activation is strongly dependent on the connection between ISC and EB, and Notch activation is reversible in EBs after their detachment from the ligand sending cells, the ISCs
Summary
Aging or injury in Drosophila intestine promotes intestinal stem cell (ISC) proliferation and enteroblast (EB) differentiation. We developed a 3D-printed platform “FlyVAB” for intravital imaging strategy that enables the visualization of the Drosophila posterior midgut at a single cell level across the ventral abdomen cuticle. Because lineage analysis is based on a single image taken at the end of a time window in days, the evaluation of the stem cell division rate, the order of progeny generation, and the dynamics of EB differentiation is difficult. The adult Drosophila is glued to cover glass, and this device enables the intravital tracing of stem cells lineage with intervals of 2–4 days Since this strategy uses glue as an intermedium and lateral compression is complicated to restore the gut position, “Bellymount” has its limits in tracking every cell division in ISC lineage over days or revealing the dynamics of EB differentiation. Our intravital imaging strategy reveals a differentiation-poised EB status and a differentiation-activated EB status that was closely correlated with the local intestinal physiology conditions
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