Longitudinal monitoring of pancreatic islet damage in streptozotocin-treated mice with optical coherence microscopy

Abstract

Pancreatic islets regulate glucose homeostasis in the body, and their dysfunction is closely related to diabetes. Islet transplantation into the anterior chamber of the eye (ACE) was recently developed for both in vivo islet study and diabetes treatment. Optical coherence microscopy (OCM) was previously used to monitor ACE transplanted islets in non-obese diabetic (NOD) mice for detecting autoimmune attack. In this study, OCM was applied to streptozotocin (STZ)-induced diabetic mouse models for the early detection of islet damage. A custom extended-focus OCM (xfOCM) was used to image islet grafts in the ACE longitudinally during STZ-induced beta cell destruction together with conventional bright-field (BF) imaging and invasive glucose level measurement. xfOCM detected local structural changes and vascular degradation during the islet damage which was confirmed by confocal imaging of extracted islet grafts. xfOCM detection of islet damage was more sensitive than BF imaging and glucose measurement. Longitudinal xfOCM images of islet grafts were quantitatively analyzed. All these results showed that xfOCM could be used as a non-invasive and sensitive monitoring method for the early detection of deficient islet grafts in the ACE with potential applications to human subjects.

Fig. 1.

A system configuration of custom xfOCM. Bessel-beam illumination and Gaussian-beam detection paths are depicted in gray and red colors, respectively. Optical delay part in the reference arm and a spectrometer is presented in black and red dashed line boxes, respectively. FC: fiber coupler, COL: collimator, AP: angled prism, SM: scanning mirror, BS: non-polarizing beam splitter, VPHG: volume phase holographic grating, L1-L5: achromatic lens, L6: 10x objective lens.

Fig. 2.

Representative bright-field (BF) and xfOCM images of an islet graft in the ACE before and after STZ-induced damage. (a1-a2) BF images of the islet before and 3 days after STZ injection, respectively. Magnified BF islet images were presented as insets. (b1-b2) xfOCM images of the same islet graft before and 3 days after STZ injection, respectively. Yellow dashed lines in the x-z cross-sectional images indicated the depth of x-y en-face images. (c) Blood glucose levels of mice (n = 4) at the two different time points. (d-e) Representative confocal fluorescence images of a control islet graft and the one taken from the mice euthanized on day 5 after STZ injection, respectively. Confocal image of the islet graft on day 5 after STZ injection was the same one observed in the longitudinal BF and xfOCM images. Insulin (red) and glucagon (green) were visualized by immunofluorescence labeling. Scale bars indicate 100 µm.

Fig. 3.

Early detection of islet damage caused by STZ using xfOCM. (a-b) and (c-d) BF and xfOCM images of two representative islet grafts in the ACE before and in 12 hours after STZ injection. (a1-a2) and (c1-c2) BF images of the two islet grafts at the two time points. (b1-b2) and (d1-d2) xfOCM images of the two islet grafts. (e) Glucose levels in experimental mice (n = 6) at the two time points. (f-g) Confocal fluorescence images of islets before and in 12 hours after STZ injection. The islet in (g) was the same one shown in (a2) and (b2), and it was marked with asterisks. The confocal images showed cells in the islet with insulin (red) and glucagon (green) labeling. Scale bars indicate 100 µm.

Fig. 4.

Quantitative analysis of longitudinal xfOCM images of islet grafts in the early period of islet damage induced by STZ. (a) Representative xfOCM images and intensity histograms of an islet graft before and in 12 hours after STZ injection. Cyan arrow heads in the xfOCM image in 12 hours after the injection marked local reduced scattering regions. (b-c) Median intensity and skewness values of the intensity histograms from 11 islet grafts at the two different time points. (d) Ratio of vascular volume in the islet grafts between the two time points. Statistical significances were presented by number of asterisks depending on the p-value: *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.