Abstract
The specific functional properties of a tissue are distributed amongst its component cell types. The various cells act coherently, as an ensemble, in order to execute a properly orchestrated physiologic response. Thus, modern approaches to dissect physiologic mechanism would benefit from an ability to identify specific cell types in live tissues and image them in real time. Current techniques require the use of fluorescent genetic reporters that are not only cumbersome, but which only allow the simultaneous study of 2 or 3 cell types. We report a non-invasive imaging modality that capitalizes on the endogenous autofluorescence signatures of the metabolic cofactors NAD(P)H and FAD. By marrying morphological characteristics with autofluorescence signatures, all seven of the airway epithelial cell types can be distinguished simultaneously in real time. Furthermore, we find that this methodology for direct cell type specific identification avoid potential pitfalls with the use of ostensibly cell type-specific markers that can be altered by clinically relevant physiologic stimuli. Finally, we utilize this methodology to interrogate real-time physiology using a clinically relevant model of cholinergic stimulation and identify dynamic secretory cell associated antigen passages (SAPs) that are highly reminiscent of previously reported goblet cell associated antigen passages (GAPs) in the intestine.
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Competing Interest Statement
The authors have declared no competing interest.
Footnotes
Competing interests: No competing interests declared
Data Availability: All data is included in the manuscript and generated custom scripts are included as Appendix. N A
Ethics: Human Subjects: No Animal Subjects: Yes Ethics Statement: Mice were maintained in accordance with the Association for Assessment and Accreditation of Laboratory Animal Care-accredited animal facility at the Massachusetts General Hospital (MGH). All procedures were performed with Institutional Animal Care and Use Committee (IACUC)-approved protocols (#2009N000119). MGH is accredited by AAALAC International, has an assurance with the Office of Laboratory Animal Welfare (OLAW) and is registered with the United States Department of Agriculture (USDA). Euthanasia was performed via house line CO2-mediated asphyxiation and confirmatory cervical dislocation consistent with the recommendations of the American Veterinary Medical Association.
