High-resolution 3D volumetry versus conventional measuring techniques for the assessment of experimental lymphedema in the mouse hindlimb

Abstract

Secondary lymphedema is a common complication of cancer treatment characterized by chronic limb swelling with interstitial inflammation. The rodent hindlimb is a widely used model for the evaluation of novel lymphedema treatments. However, the assessment of limb volume in small animals is challenging. Recently, high-resolution three-dimensional (3D) imaging modalities have been introduced for rodent limb volumetry. In the present study we evaluated the validity of microcomputed tomography (μCT), magnetic resonance imaging (MRI) and ultrasound in comparison to conventional measuring techniques. For this purpose, acute lymphedema was induced in the mouse hindlimb by a modified popliteal lymphadenectomy. The 4-week course of this type of lymphedema was first assessed in 6 animals. In additional 12 animals, limb volumes were analyzed by μCT, 9.4 T MRI and 30 MHz ultrasound as well as by planimetry, circumferential length and paw thickness measurements. Interobserver correlation was high for all modalities, in particular for μCT analysis (r = 0.975, p < 0.001). Importantly, caliper-measured paw thickness correlated well with μCT (r = 0.861), MRI (r = 0.821) and ultrasound (r = 0.800). Because the assessment of paw thickness represents a time- and cost-effective approach, it may be ideally suited for the quantification of rodent hindlimb lymphedema.

Quantification of tissue volume in the hindlimb of mice using microcomputed tomography images and analysing software

“…We have also included how good positioning affects interrater agreement compared to poor positioning, and we examined the intrarater agreement. Frueh et al had shown that the interrater agreement was high (r = 0.975, p < 0.001) for µCT analysis 17 . Their BA LoA where a lot wider than ours when examining the interrater agreement.…”

“…The population of interest consisted of C57BL/6 mice from two prior experiments, and the rater population of interest was comprised of medical staff with a BSc degree. Interrater agreement for µCTanalysis has previously been shown to be high (r = 0.975, p < 0.001) 17 . While the study by Frueh et al focused on the comparison of µCT with several different measuring techniques, reporting additionally on interrater agreement, our study targeted intra-and interrater agreement in µCT measuring, and the Guidelines for Reporting Reliability and Agreement Studies (GRRAS) 22 were applied.…”

“…prone to bias, especially when measuring small animals such as rodents. Paw thickness has been used as a surrogate parameter for volume of the hindlimb 13 , and has been suggested to be the most suitable method to assess the course of lymphedema in mice as it is an inexpensive, fast and reproducible method 17 . Both paw thickness and circumferential measurement are inexpensive and easy to use, but it should be noted that these methods only assess the change in volume and do not allow the researcher to directly quantify the volume of the hindlimb.…”

“…In recent studies, three-dimensional (3D) image analysing software has been used to quantify tissue volume in the hindlimb of rodents using both µCT- [17][18][19] and magnetic resonance imaging (MRI) 17,20 . It has been shown that both µCT and MRI analysis provide reliable measurements of hindlimb volume when compared to conventional measuring techniques, such as caliper-measured paw thickness 17 . The limitations of using µCT and MRI scans to measure volume are that they are expensive and relatively time-consuming when compared to the alternative techniques.…”

Magnetic Resonance Lymphography at 9.4 T Using a Gadolinium-Based Nanoparticle in Rats

“…Caliper-measured paw thickness represents a valid surrogate parameter for rodent hindlimb volumes. 25 Repetitive measurements in isoflurane anesthesia were performed over a period of 14 weeks.…”