Label-free imaging immune cells and collagen in atherosclerosis with two-photon and second harmonic generation microscopy

Atherosclerosis has been recognized as a chronic inflammation disease,in which many types of cells participate in this process,including lymphocytes,macrophages,dendritic cells(DCs),mast cells,vascular smooth muscle cells(SMCs).Developments in imaging technology provide the capability to observe cellular and tissue components and their interactions.The knowledge of the functions of immune cells and their interactions with other cell and tissue components will facilitate our discovery of biomarkers in atherosclerosis and prediction of the risk factor of rupture-prone plaques.Nonlinear optical microscopy based on two-photon excited autofluorescence and second harmonic generation(SHG)were developed to image mast cells,SMCs and collagen in plaque ex vivo using endogenous optical signals.Mast cells were imaged with two-photon tryptophan autofluorescence,SMCs were imaged with two-photon NADH auto fluorescence,and collagen were imaged with SHG.This development paves the way for further study of mast cell degranulation,and the effects of mast cell derived mediators such as induced synthesis and activation of matrix metalloproteinases(MMPs)which participate in the degradation of collagen.