Intravital imaging of CTLs killing islet cells in diabetic mice
Ken Coppieters, … , Natalie Amirian, Matthias von Herrath
Ken Coppieters, … , Natalie Amirian, Matthias von Herrath
Published December 1, 2011
Citation Information: J Clin Invest. 2012;122(1):119-131. https://doi.org/10.1172/JCI59285.
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Intravital imaging of CTLs killing islet cells in diabetic mice

Abstract

Type 1 diabetes (T1D) is caused by autoimmune destruction of the insulin-producing β cells in the pancreatic islets, which are essentially mini-organs embedded in exocrine tissue. CTLs are considered to have a predominant role in the autoimmune destruction underlying T1D. Visualization of CTL-mediated killing of β cells would provide new insight into the pathogenesis of T1D, but has been technically challenging to achieve. Here, we report our use of intravital 2-photon imaging in mice to visualize the dynamic behavior of a virally expanded, diabetogenic CTL population in the pancreas at cellular resolution. Following vascular arrest and extravasation, CTLs adopted a random motility pattern throughout the compact exocrine tissue and displayed unimpeded yet nonlinear migration between anatomically nearby islets. Upon antigen encounter within islets, a confined motility pattern was acquired that allowed the CTLs to scan the target cell surface. A minority of infiltrating CTLs subsequently arrested at the β cell junction, while duration of stable CTL–target cell contact was on the order of hours. Slow-rate killing occurred in the sustained local presence of substantial numbers of effector cells. Collectively, these data portray the kinetics of CTL homing to and between antigenic target sites as a stochastic process at the sub-organ level and argue against a dominant influence of chemotactic gradients.

Authors

Ken Coppieters, Natalie Amirian, Matthias von Herrath

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Figure 3

Profound leakiness of postcapillary islet venules is unrelated to endothelial apoptosis.

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Profound leakiness of postcapillary islet venules is unrelated to endoth...
(A and B) Isosurfacing rendition of vascular staining from region depicted in Figure 2, H and I, before and 10 minutes after injection of the dye, respectively. w/h = 1.52/d = 5/z = 26. (C and D) Naive control animal imaged before and 14 minutes after injection with dextran–Texas Red (dextran-TxRed). w/h = 1.52/d = 5/z = 33. (E) Quantitative analysis of vascular leakiness as observed in A and B compared with baseline in C and D. Leakiness was defined as increase in isosurfaced dextran–Texas Red–derived fluorescent signal and was normalized to T = 0 immediately after injection. Representative of observations from 3 individual prediabetic animals and 3 controls. (F and G) TUNEL staining was performed to reveal ongoing apoptosis in the transfer model during the late prediabetic phase. TUNEL+ nuclei are exclusively observed within the β cell mass and inflammatory infiltrate. Red arrows indicate vascular structures and the lack of ongoing apoptosis within these regions. Images are representative of at least 3 consecutive sections from 4 individual prediabetic animals. Scale bars: 100 μm.