Purpose: To investigate the thicknesses and areas of Henle fiber layer (HFL), outer nuclear layer, and outer plexiform layer in the eyes of patients with diabetes with no diabetic retinopathy, in eyes with nonproliferative diabetic retinopathy without diabetic macular edema, and in healthy eyes using a modified directional optical coherence tomography strategy.Methods: In this prospective study, the no diabetic retinopathy group included 79 participants, the nonproliferative diabetic retinopathy group comprised 68 participants, and the control group had 58 participants. Thicknesses and areas of Henle fiber layer, outer nuclear layer, and outer plexiform layer were measured on a horizontal single optical coherence tomography scan centered on the fovea using directional optical coherence tomography.Results: The foveal, parafoveal, and total HFL were significantly thinner in the nonproliferative diabetic retinopathy group than in the no diabetic retinopathy group and the control group (all P , 0.05). The no diabetic retinopathy group had significantly thinner foveal HFL thickness and area compared with the control group (all P , 0.05). The nonproliferative diabetic retinopathy group had significantly thicker outer nuclear layer thickness and area in all regions than the other groups (all P , 0.05). The outer plexiform layer measurements did not differ between the groups (all P . 0.05).Conclusion: Directional optical coherence tomography provides isolated thickness and area measurement of HFL. In patients with diabetes, the HFL is thinner, and HFL thinning begins before the presence of diabetic retinopathy.