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Imaging and Analysis of Neurofilament Transport in Excised Mouse Tibial Nerve.

ABSTRACT: Neurofilament protein polymers move along axons in the slow component of axonal transport at average speeds of ~0.35-3.5 mm/day. Until recently the study of this movement in situ was only possible using radioisotopic pulse-labeling, which permits analysis of axonal transport in whole nerves with a temporal resolution of days and a spatial resolution of millimeters. To study neurofilament transport in situ with higher temporal and spatial resolution, we developed a hThy1-paGFP-NFM transgenic mouse that expresses neurofilament protein M tagged with photoactivatable GFP in neurons. Here we describe fluorescence photoactivation pulse-escape and pulse-spread methods to analyze neurofilament transport in single myelinated axons of tibial nerves from these mice ex vivo. Isolated nerve segments are maintained on the microscope stage by perfusion with oxygenated saline and imaged by spinning disk confocal fluorescence microscopy. Violet light is used to activate the fluorescence in a short axonal window. The fluorescence in the activated and flanking regions is analyzed over time, permitting the study of neurofilament transport with temporal and spatial resolution on the order of minutes and microns, respectively. Mathematical modeling can be used to extract kinetic parameters of neurofilament transport including the velocity, directional bias and pausing behavior from the resulting data. The pulse-escape and pulse-spread methods can also be adapted to visualize neurofilament transport in other nerves. With the development of additional transgenic mice, these methods could also be used to image and analyze the axonal transport of other cytoskeletal and cytosolic proteins in axons.

SUBMITTER: Boyer NP

PROVIDER: S-EPMC8006945 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundDiabetic peripheral neuropathy (DPN) is the most common diabetes-associated complication and imposes a significant burden to healthcare systems. Thus, early diagnosis of DPN is extremely critical for management and outcome of diabetic patients. Supersonic Shear Wave Imaging (SSI) enables the noninvasive measurement of nerve stiffness. However, previous studies on SSI in the diagnosis of DPN were limited in sample sizes and reported various results. In this meta-analysis, we aimed to obtain comprehensive evidence on the value of tibial nerve stiffness measurement by SSI in the diagnosis of DPN.MethodsA comprehensive literature search in English and Chinese electronic database was conducted for studies (published until January 25, 2022) that investigated the diagnostic performance of tibial nerve stiffness measurement by SSI for detecting DPN. Summary receiver operating characteristics (SROC) modelling was constructed to conduct the meta-analysis of diagnostic accuracy of SSI for detecting DPN.ResultsFinally, a total of 12 eligible studies with 1325 subjects were included for evaluation, and a meta-analysis was conducted to evaluate the diagnostic performance of tibial nerve stiffness measurement by SSI for detecting DPN. For tibial nerve stiffness measurement by SSI, the summary sensitivity and specificity for the diagnosis of DPN were 80% (95% confidence interval [CI]: 73%-86%) and 86% (95% CI: 82%-89%), respectively. The summary area under the ROC curve (AUROC) value of the SROC was 0.90 (95% CI: 0.87-0.92), for diagnosing DPN. A subgroup analysis of 11 SSI studies from China revealed similar diagnostic performance, with a summary sensitivity of 79% (95% CI: 72%-85%), specificity of 86% (95% CI: 82%-89%) and summary AUROC value of the SROC of 0.90 (95% CI: 0.87-0.92) for diagnosing DPN.ConclusionsOur meta-analysis suggests that a tibial nerve stiffness measurement by SSI shows good performance in diagnosing DPN and has considerable potential as a noninvasive tool for detecting DPN.

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Imaging and Analysis of Neurofilament Transport in Excised Mouse Tibial Nerve.

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