Imaging and quantification of subbasal nerve plexus in healthy volunteers and diabetic patients with or without retinopathy

Abstract

Background: The alterations of subbasal nerve plexus (SBP) innervation and corneal sensation were estimated non-invasively and compared with the values in healthy volunteers. Additionally, this study addressed the relation of SBP changes to the retinal status, glycemic control and diabetes duration.

Methodology/principal findings: Eighteen eyes of diabetic patients with peripheral diabetic neuropathy aged 68.8±8.8 years and twenty eyes of healthy volunteers aged 66.3±13.3 yrs. were investigated with in vivo confocal laser-scanning microscopy (CLSM). An adapted algorithm for image analysis was used to quantify the morphological and topological properties of SBP. These properties were correlated to incidence of diabetic retinopathy (DR) and corneal sensation (Cochet-Bonnet esthesiometer). The developed algorithm allows a fully automated analysis of pre-segmented SBP structures. Altogether, 10 parameters were analysed, and all of them revealed significant differences between diabetic patients and healthy volunteers. The nerve fibre density, total fibre length and nerve branches were found to be significantly lower in patients with diabetes than those of control subjects (nerve fibre density 0.006±0.002 vs. 0.020±0.007 mm/mm(2); total fibre length 6223±2419 vs. 19961±6553 µm; nerve branches 25.3±28.6 vs. 141.9±85.7 in healthy volunteers). Also the corneal sensation was significantly lower in diabetic group when compared to controls (43±11 vs. 59±18 mm). There was found no difference in SBP morphology or corneal sensation in the subgroups with (DR) or without (NDR) diabetic retinopathy.

Conclusions/significance: SBP parameters were significantly reduced in diabetic patients, compared to control group. Interestingly, the SBP impairment could be shown even in the diabetic patients without DR. Although automatic adapted image analysis simplifies the evaluation of in vivo CLSM data, image acquisition and quantitative analysis should be optimised for the everyday clinical practice.

Figure 1. Seven standard-field fundus photography with corresponding CLSM images.

Regular SBP pattern and normal fundus appearance in control (A). Patches of retinal whitening and rarefication of nerve fibers in SBP in a patient with DR (B). Relatively impaired appearance of SBP in a patient without DR (C).

Figure 2. Confocal microscopy and quantification of micromorphological parameters.

Initial image of SBP in the central cornea obtained with in vivo CLSM in a healthy volunteer (A) with corneal sensation 60 mm and diabetic patient (D) with corneal sensation 40 mm and NDS = 8 (image size: 400 x 400 µm). B and E represent the results of segmentation from the corresponding SBP images in control and diabetic subjects, respectively. C and F show graphs displaying the geometry of SBP in a final surface reconstruction. Total fibre length of 4706 and 545.4 µm, nerve fibre density 0.034 mm/mm² and 0.004 mm/mm², and single nerve fibre count 68 and 3 were measured in control subject and diabetic patient, respectively.