Label free autofluorescence imaging permits comprehensive and simultaneous assignment of cell type identity and reveals the existence of airway secretory cell associated antigen passages (SAPs)

Abstract

AbstractThe 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.eLife’s Review ProcesseLife works to improve the process of peer review so that it more effectively conveys the assessment of expert reviewers to authors, readers and other interested parties. In the future we envision a system in which research is first published as a preprint and the outputs of peer review are the primary way research is assessed, rather than journal title.Our editorial process produces two outputs: i) an assessment by peers designed to be posted alongside a preprint for the benefit of the readers;i) detailed feedback on the manuscript for the authors, including requests for revisions and suggestions for improvement.Therefore we want to change how we construct and write peer reviews to make themuseful to both authors and readers in a way that better reflects the work you put into reading and thinking about a paper.eLife reviews now have three parts:Anevaluation summary(in two or three sentences) that captures the major conclusions of the review in a concise manner, accessible to a wide audience.Apublic reviewthat details the strengths and weaknesses of the manuscript before you, and discusses whether the authors’ claims and conclusions are justified by their data.A set of privaterecommendations for the authorsthat outline how you think the science and its presentation could be strengthened.All three sections will be used as the basis for an eLife publishing decision, which will, as always, be made after a consultation among the reviewers and editor. Each of thepublic reviewswill be published (anonymously) alongside the preprint, together with a response from the authors if they choose. In the case of papers we reject after review, the authors can choose to delay posting until their paper has been published elsewhere.If this is your first time going through this new process, we ask that you take some time to read ourReviewer Guide, which discusses how we see each section will be used, what it should contain, and what we hope it accomplishes. And we remind you that, with the shift of reviews from private correspondence to public discourse, it is more important than ever that reviews are written in aclear and constructive mannerappropriate for a public audience and mindful of the impact language choices might have on the authors.