Imaging Meets Cytometry: Analyzing Heterogeneous Functional Microscopic Data from Living Cell Populations

Abstract

Biological tissue consists of populations of cells exhibiting different responses to pharmacological stimuli. To probe the heterogeneity of cell function, we propose a multiplexed approach based on real‐time imaging of the secondary messenger levels within each cell of the tissue, followed by extraction of the changes of single‐cell fluorescence over time. By utilizing a piecewise baseline correction, we were able to quantify the effects of multiple pharmacological stimuli added and removed sequentially to pancreatic islets of Langerhans, thereby performing a deep functional profiling for each cell within the islet. Cluster analysis based on the functional profile demonstrated dose‐dependent changes in statistical inter‐relationships between islet cell populations. We therefore believe that the functional cytometric approach can be used for routine quantitative profiling of the tissue for drug screening or pathological testing.

Taurine rescues pancreatic β‐cell stress by stimulating α‐cell transdifferentiation

“…All counts were determined in a blinded manner with 25–150 islets analyzed per treatment group, as indicated in the figure legends. Image processing for live‐cell recordings (ROI mapping, intensity‐vs.‐time computation) was performed in Fiji, with subsequent time series data analysis done in IgorPro (Wavemetreics), as detailed in 44 . Statistical analysis was performed using GraphPad PRISM 5.0 (GraphPad, U.S.A.) and/or R4.1.0 45 .…”

“…Image processing for live-cell recordings (ROI mapping, intensity-vs.-time computation) was performed in Fiji, with subsequent time series data analysis done in IgorPro (Wavemetreics), as detailed in. 44 Statistical analysis was performed using GraphPad PRISM 5.0 (GraphPad, U.S.A.) and/or R4.1.0. 45 For cluster analysis (Figure 7F), we used the mean values for 25 phenotypic characteristics presented in this study (Figures 1-4), inclusive of global animal physiological indexes as well as proliferative, apoptotic and differentiation parameters from current study and our earlier works.…”

Classical and non-classical islet peptides in the control of β-cell function

“…GLP-1 secretion has a recognised glucose-dependent insulinotropic action [70] and, although the lifetime of the active form of GLP-1 (7-36) is limited by the activity of the intra-islet dipeptidyl peptidase-IV, the inactive form of GLP-1 (9-36), has been claimed to elevate cytosolic cAMP, at high concentrations [213]. This suggests a potential role for both major forms of locally produced islet GLP-1.…”