Background: Diabetic peripheral neuropathy (DPN) is a progressive complication of diabetes that initially affects the small nerve fibres in the peripheral nervous system. Early detection of DPN is essential to slow or reverse the progression. Structural changes to the corneal nerves have been observed using in vivo corneal confocal microscopy and serve as a biomarker for small nerve fibre damage in DPN. Also, nerve excitability is a non-invasive technique that allows the assessment of ion-channel function in the nodal and internodal regions of the axon. Previous studies have shown that type 1 diabetic patients with and without DPN have impaired ion-channel activity in the axonal membrane. The relationship between corneal nerve morphology and axonal function in type 1 patients with or without DPN is yet to be established.
 
Aims: To investigate the relationship between corneal nerve structure and axonal ion-channel function in patients with and without diabetic peripheral neuropathy.

Methods: Thirty-six participants (23M:13F) with an average type I diabetes duration of 16.1 ± 10.2 years and a mean age of 35.94 ± 15.20 years were enrolled. Average HbA1c and BMI was 64.4 ± 16.1 mmol/mol and 24.6 ± 3.4 kg/m2. All participants underwent confocal microscopy to assess corneal nerve fibre length (CNFL), fibre density (CNFD), branch density (CNBD), fibre tortuosity (CNFT), fibre total branch density (CTBD), fibre area (CNFA), fibre width (CNFW) and inferior whorl length (IWL). The presence and severity of DPN was assessed by the total neuropathy score (TNS) which involved comprehensive neurological assessments including nerve conduction studies, and assessment of symptoms and signs of DPN. Based on the TNS, participants were stratified into those with neuropathy (TNS > 1; n = 10) and those without neuropathy (TNS < 1; n = 26). Motor excitability studies were conducted on the median nerve to assess axonal ion-channel function in nodal and internodal regions of the axonal membrane. 
Results: Overall, IWL (r=-0.42; p<0.05) decreased with increasing TNS. The latency and peak response was positively correlated with neuropathy severity (r=0.64, p<0.01; r=-0.4, p<0.05). CNBD, CNFL, CTBD and CNFA showed strong positive correlations with excitability measures that reflect nodal sodium ion-channel activity (rheobase, r=0.46, p<0.01; r= 0.36, p<0.05; r=0.38, p<0.05; r=0.38, p<0.05, respectively) as did CNBD, CTBD and CNFA (stimulus for 50% peak, r=0.5, p<0.01; r=0.4, p<0.05; r=0.37, p<0.05, respectively). CNFW showed a negatively correlated with potassium ion-channel activity in the internodal region (hyperpolarising threshold electrotonus (90-100ms), r=-0.36, p<0.05). Subjects with DPN had a significant reduction in CNFD and peak response compared to subjects without DPN (24.83 ± 6.73 vs. 19.74 ± 4.06 fibres/mm2, p=0.03; 11.8 ± 2.57 vs. 8.2 ± 2.46, p<0.01). In those with DPN, CNFL was positively correlated with internodal potassium ion-channel activity (depolarising threshold electrotonus (90-100ms), r=0.65, p<0.05; depolarising electrotonus (40-60ms), r=0.64, p<0.05). A negative correlation was observed between IWL and measures of internodal potassium ion-channel function (superexcitability, r=-0.91, p<0.01). In those without neuropathy, CNFD, CNBD, CNFL and CTBD positively correlated with sodium ion-channel function (rheobase, r=0.41, p<0.01; r=0.60, p<0.001; r=0.51, p<0.001; r=0.6, p<0.01, respectively). IWL was negatively associated with other measures of sodium ion-channel activity (refractoriness, r=-0.62, p<0.01).

Discussion: Structural changes of corneal nerve fibres correlated with electrophysiological measures of nerve function in patients with type 1 diabetes. These findings suggest that corneal nerve structural abnormalities may provide an indication of different aspects of axonal membrane function prior to the onset and during the progression of DPN. These correlations also indicate that there are diffuse changes throughout the nervous system in patients with type 1 diabetes with and without DPN. The changes in corneal nerve fibre morphology are more pronounced in patients with severe DPN. In conclusion, abnormal structural changes of corneal nerve fibres are associated with neuropathy severity and altered axonal function in patients with type 1 diabetes.

The relationship between structural and electrophysiological measures of peripheral nerves in human type 1 diabetes