## 4 Discussion
Although the observer was blind to the LASIK group classification when subjectively assessing nerve tortuosity, the obtained scores were not significantly different among groups (Table 2). In contrast, the automated tortuosity index showed that the LASIK pain group suffered significantly higher tortuosity than the non-symptomatic LASIK group (Table 2). However, the difference between both symptomatic LASIK groups was not significant, although the score of the LASIK pain group was higher. [PERSON] et al. [54] have reported that autoimmune DED patients suffering from corneal pain show increased nerve tortuosity in comparison with healthy controls. These authors used a subjective grading system, which might be the reason why they did not find increased tortuosity in Meibomiam gland dysfunction patients suffering from corneal pain, although their tortuosity values were also increased. In the present study, we observed a moderate (\(r=0.49\)) correlation between the subjective and automated tortuosity grading methods, but the automated tool is expected to provide more reliable data. In addition, it was also observed that the values of the automated tortuosity index showed a positive relationship with OSDI and NRS pain scores (Figure 4). These findings show that subjective ocular surface perception is obviously related to the nervous system, specifically the ophthalmic division of the trigeminal nerve. In fact, it has been reported that, after corneal refractive surgery, the regeneration of corneal nerves correlates strongly with the recovery of corneal sensation [55]. On the contrary, in the present study, CFS and TBUT did not show any relationship with the automated tortuosity index. This finding supports previous reports highlighting that, in LASIK patients suffering from chronic corneal pain, conventional DED tests do not explain the persistent ocular surface symptoms [56; 57]. The limitations of the current study are related to the nature of the sample used to evaluate the automated method. First, our recruited volunteers were post-LASIK patients with DED symptoms. These selection criteria contributed to reducing the heterogeneity of the image features in the sample. However, it would be interesting to evaluate the performance of this new automatic tool in other patients suffering from different anomalies. Future studies should assess if the proposed automated method can perform similarly using corneal sub-basal plexus images with additional information such as inflammatory cells or gliomas. However, our tool could be optimized to detect these features on IVCM images to increase the accuracy of the outcomes in other anomalies. Second, the sample size was relatively small, especially in the post-LASIK DED group. This shortfall would have an impact on the statistical power of the hypothesis test to identify as significant the differences between groups. Despite these limitations, this proof-of-concept study designed to describe an automatic method has provided data showing its adequateness for the assessment of corneal nerve tortuosity. Future studies should corroborate this study's findings using larger sample populations, including patients with diverse ocular anomalies.