A Dual-Labeled Annexin A5 is not Suited for SPECT Imaging of Brain Cell Death in Experimental Murine Stroke

Abstract

Cell death is one of the pathophysiological hallmarks after stroke. Markers to image cell death pathways in vivo are highly desirable. We previously showed that fluorescently labeled Annexin A5 (AnxA5), which binds specifically to phosphatidylserine (PS) on dead/ dying cells, can be used in experimental stroke for monitoring cell death with optical imaging. Here we investigated whether dual-labeled AnxA5 (technetium and fluorescence label) can be used for single-photon emission computed tomography (SPECT) of cell death in the same model. C57Bl6/N mice were subjected to 60-minute middle cerebral artery occlusion (MCAO) and underwent SPECT imaging at 24, 48, and 72 hours afterwards. They were injected intravenously with either PS-binding AnxA5 or the nonfunctional AnxA5 (negative control), labeled with 99mTc and Alexa Fluor 568, respectively. After SPECT imaging, brain sections were cut for autoradiography and fluorescence microscopy. Ethanol-induced cell death in the femur muscle was used as positive control. We detected dual-labeled AnxA5 in the model of ethanol-induced cell death in the femur muscle, but not after MCAO at any time point, either with SPECT or with ex vivo autoradiography or fluorescence microscopy. Dual-labeled AnxA5 appears to be unsuited for visualizing death of brain cells in this MCAO model.

Phenoxide-Bridged Zinc(II)-Bis(dipicolylamine) Probes for Molecular Imaging of Cell Death

“…Hematoxylin/eosin staining of kidney histology slices from probe-treated rats did not show any abnormalities (Figure S11), suggesting that the kidney retention was not due to probe-induced cell death. A similar kidney retention effect has been observed with Annexin V, 44,45 where there is histological evidence that Annexin V localizes to the distal tubular cells in healthy kidneys 46,47 due to relatively high levels of PS in the cortical regions of the kidney. 48 The favorable biodistribution profile of fluorescent probe 1 prompted us to conduct a comparative study of two radiotracer probes, 9 and 10 (Scheme 4).…”

Single photon emission computed tomography imaging of cerebral blood flow, blood–brain barrier disruption, and apoptosis time course after focal cerebral ischemia in rats

“…26,27 In vivo experiments showed that 99m Tc-Anx-V128 may be superior than 99m Tc-Annexin V-HYNIC which has a high uptake in kidneys and the liver, that might be limiting to an acceptable target to background ratio, 28 and to HIS-cys-Anx-AF568 which was too hydrophilic to reach the cerebral compartment. 29…”

Annexin Animal Models—From Fundamental Principles to Translational Research

“…As mentioned above, 99m TC radiolabeled AnxA5 and AnxA5-conjugated gold nanoparticles were valuable diagnostic imaging tools in mouse models for atherosclerosis [ 253 , 254 , 255 ] ( Table 5 d). AnxA5 dual-labeled with 99m TC and Alexa Fluor 568 allowed detection of ethanol-induced cell death in the femur muscle but failed to visualize cell death in a murine model for stroke [ 272 ]. Imaging of apoptosis using 111 In- and DOTA-peptide -labeled AnxA5 monitored efficacy of suicide gene therapy after treatment with ganciclovir in NG4TL4 sarcoma expressing herpes simplex virus thymidine kinase [ 273 ].…”