Abstract Details

Title Sweat Gland Nerve Fiber Density: Development of a Novel, Unbiased 3D Reconstruction Methodology

Topic Neuromuscular and Clinical Neurophysiology (EMG)

Presentation(s) P3 - Poster Session 3 (5:30 PM-6:30 PM)

Poster/Presentation
Number 7-056

Objective

To develop a reliable and reproducible technique to quantify SGNFD using 3D reconstruction of nerve fibers.

Background Dermal structures are densely innervated by autonomic nerve fibers. Over the past decade skin biopsies have sparked great interest in the evaluation of autonomic disorders. Different groups have reported on sweat gland nerve fiber density (SGNFD) as a means to quantitate sudomotor innervation. However, methodologies vary significantly from imaging to analyzing techniques and although conventional stereology is commonly used, no standard technique has been established. In an attempt to bridge this gap, we sought to develop a reliable and reproducible technique to quantify SGNFD.

Design/Methods Four-millimeter skin punch biopsies were collected from healthy individuals. Biopsies were stained with PGP 9.5 and Collagen IV antibodies to visualize nerve fibers and sweat gland (SG) tubules, respectively. SGs were imaged using confocal and wide-field microscopes. Volumes were estimated using contouring functions and images were optimized using deconvolution algorithms. Nerve fibers were reconstructed and nerve fiber length (NFL) was quantified using automated 3D software. SGNFD was obtained by dividing NFL by volume. SGNFD was also assessed using stereology for comparison.

Results A total of 92 SGs from 10 subjects were analyzed by 2 independent observers using both microscopy techniques. Using confocal microscopy, the software reliably traced nerve fibers surrounding sweat glands requiring minimal operator adjustment. The rendering of nerve fibers was inferior when using wide-field microscopy, often raising questions about true nerve signal versus artifact, requiring more operator input. This is reflected in the intraclass correlation coefficient (ICC), which was ICC=0.98 using confocal images versus ICC=0.82 using wide-field images. Correlation between 3D-reconstructon and stereology was poor (ICC=0.38).

Conclusions

The newly developed technique of SGNFD quantitation using deconvolution, 3D reconstruction and confocal microscopy reliably traces SG nerve fibers, shows outstanding reproducibility, is almost completely unbiased, and arguably superior to conventional stereology methods.

Authors/Disclosures
Karla Minota, MD PRESENTER	No disclosure on file
Ann M. Schmeichel	Ann M. Schmeichel has nothing to disclose.
	No disclosure on file
Jayawant N. Mandrekar, PhD	Dr. Mandrekar has nothing to disclose.
Phillip A. Low, MD, FAAN (Mayo Clinic)	Dr. Low has nothing to disclose.
Wolfgang Singer, MD, FAAN (Mayo Clinic)	Dr. Singer has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Biohaven. The institution of Dr. Singer has received personal compensation in the range of $10,000-$49,999 for serving as a Consultant for Lundbeck. Dr. Singer has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Ionis. Dr. Singer has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Yoda. Dr. Singer has received personal compensation in the range of $5,000-$9,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Theravance. Dr. Singer has received personal compensation in the range of $5,000-$9,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Ferrer. The institution of Dr. Singer has received research support from NIH. The institution of Dr. Singer has received research support from FDA. The institution of Dr. Singer has received research support from Michael J. Fox Foundation. Dr. Singer has received intellectual property interests from a discovery or technology relating to health care.

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