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Nils Wendel, Weiwei Wang, Alistair Barber; Corneal sensory nerve fiber swelling in streptozotocin-diabetic rats. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3242.
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© ARVO (1962-2015); The Authors (2016-present)
Patients with diabetes often develop corneal nerve fiber atrophy, associated with comorbidities such as reduced corneal wound-healing, but other changes that precede nerve fiber atrophy are not well established. The aim of this study was to characterize early changes in the corneal sensory nerve fibers of experimentally diabetic rats.
Male Long-Evans rats (175-200g) were made diabetic by streptozotocin (STZ, Sigma) injection (i.v., 100 mg/kg, citrate buffer, pH 4.5; n=12) and housed in parallel with vehicle-injected age-matched controls (CNT, n=7). All experiments were in compliance with the ARVO statement for the use of animals in ophthalmic research and procedures were reviewed by the institutional animal care and use committee. Rats were sacrificed after 7 (STZ n=4), 9 (CNT n=7, STZ n=4) and 10 weeks (STZ n=4) and corneas were dissected for whole-mount imaging. Immunofluorescent labelling was by primary antibody to β3-tubulin (Abcam) and secondary antibody-tagged Alexa 488 (Jackson ImmunoResearch). 2X2 montage Z-stacks of the anterior nerve fibers in the central whorl of each flat-mounted cornea were acquired by confocal microscopy (Leica SP8, 20x objective). 3D object generation, surface area and volume analysis was performed with Imaris (Bitplane) image analysis software. Statistical analysis was by t-test and linear regression.
STZ-diabetic rats had significantly greater total nerve fiber volume (N-V) (p<0.005), and normalized surface area:volume (N-SA:V) (p<0.0005) compared to CNT. No significant difference was observed between CNT and STZ groups in terms of normalized total nerve fiber surface area (p=0.13). Analysis of weight and blood glucose in the subset of animals sacrificed at 9 weeks showed that weight at sacrifice correlated significantly with both N-V (r2=0.50, p=0.015) and N-SA:V (r2 = 0.38, p=0.044). Blood glucose also correlated significantly with N-V (r2=0.51, p=0.014) and N-SA:V (r2=0.47, p=0.02).
The results suggest that corneal sensory nerve fibers swell after short durations of diabetes, evidenced by their increased volume and surface area:volume ratios. Axonal swelling is a well-established feature of early nerve injury in other neurodegenerative diseases, and may precede subsequent atrophy. Correlation with body weight and blood glucose suggest that the amount of swelling is closely related to the severity of diabetes.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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