Intumescence injury of tomato leaves is a physiological disorder in which white surface spots rise on the abaxial side of the leaf and then turn brown; when it worsens, the leaf dies. Scanning electron microscope observations showed individual cells were enlarged and raised due to intumescence injury. Vein epidermal cells and stomatal guard cells were also enlarged. Hypertrophy was observed by light microscopy in epidermal cells, spongy tissue cells, and palisade tissue cells. In thin leaf sections prepared with Technovit 7100 resin, the resin peeled from the epidermal surface on the adaxial and abaxial sides of normal leaf areas. However, resin did not peel off from the enlarged epidermis in areas of intumescence. Transmission electron microscopy imaging of the cuticular layer in the epidermal cells affected by intumescence injury showed disappearance of the outermost wax layer, abnormalities in intermediate layer, and decrease in cuticular layer. Toluidine blue O staining was used to identify abnormalities in the cuticular layer and the area of intumescence, which was stained purple. The percentage of staining area relative to total area of the leaf was used to quantify the degree of intumescence. In experiments using incubators under low-UV conditions, intumescence injury was less likely to occur in young leaves or fully expanded leaves and more likely to occur in expanding leaves. In addition, the water content of the leaves in which intumescence injury occurred was lower than that in normal leaves. Intumescence injury occurred less frequently under conditions of 70% relative humidity than under conditions of 90% relative humidity. UV and relative humidity are known to affect the development of the cuticular layer, and intumescence injury was severe under conditions in which the cuticular layer had difficulty developing. Our findings showed that intumescence injury of leaves was accompanied by abnormal cuticular layer, and suggest that occurrence of intumescence injury may be reduced under conditions in which the cuticular layer is well developed, such as with UV irradiation and low relative humidity.