Mead acid supplementation does not rescue rats from cataract and retinal degeneration induced by N-methyl-N-nitrosourea

Abstract

Fatty acids and their derivatives play a role in the response to ocular disease. Our current study investigated the effects of dietary mead acid (MA, 5,8,11-eicosatrienoic acid) supplementation on N-methyl-N-nitrosourea (MNU)-induced cataract and retinal degeneration in Sprague-Dawley rats. Experiment 1 was designed to inhibit cataract formation, with the dams fed a 2.4% MA or basal (<0.01% MA) diet during lactational periods. On postnatal day 7, male pups received a single intraperitoneal (ip) injection of 50 mg/kg MNU or vehicle. Lens opacity and morphology were examined 7 and 14 days after the MNU injection. Experiment 2 was designed to inhibit retinal degeneration and was performed with female postweaning rats. In this experiment, dams were fed the 2.4% MA or basal diet during the lactational periods. Thereafter, the female pups were continuously fed the same diets during their postweaning periods. On postnatal day 21 (at weaning), pups received a single ip injection of 50 mg/kg MNU. Retinal morphology was examined 7 days after the MNU injection. In experiment 3, six-week-old female rats were fed the 2.4% MA or basal diet starting at one week before the MNU injection and were then continuously fed the same diets until sacrifice. Rats at 7 weeks of age were given a single ip injection of 40 mg/kg MNU, and the retina was then examined morphologically one week after the MNU injection. In experiment 1, mature cataract was found in all of the MNU-treated groups, with or without MA supplementation. In experiments 2 and 3, atrophy of both the peripheral and central outer retina occurred in all rats exposed to MNU, with or without MA supplementation, respectively. The severities of the cataracts and retinal atrophy in the rats were similar regardless of MA supplementation. Dietary mead acid, which is used as a substitute in essential fatty acid deficiency in the body, does not modify MNU-induced cataract and retinal degeneration in rat models.

Keywords: N-methyl-N-nitrosourea; arachidonic acid; cataract; mead acid; rats; retinal degeneration.

Fig. 1.

Schema of the experimental protocol. Experiment 1 was designed to inhibit cataract formation in the MNU-induced cataract model using male neonatal rats. Experiments 2 and 3 were designed to confirm the effects on the MNU-induced retinal degeneration models using female postweaning and adult rats^, .

Fig. 2.

Lens opacity and mature cataract in experiment 1. (a) A severe degree of opacity in the whole lens is seen two weeks after injection of N-methyl-N-nitrosourea (MNU) in a rat fed a basal diet. (b) Similar lens opacity is seen in an MNU-treated rat fed a 2.4% mead acid (MA) diet. (c) Histopathologically, swelling, vacuolation and liquefied fibers occur in the whole lens at two weeks after injection of MNU in a rat fed a basal diet. These changes corresponded to the diagnosis of mature cataract. (d) Mature cataract is similarly seen in the whole lens of a MNU-treated rat fed an MA diet. Hematoxylin and eosin (HE) staining, ×30.

Fig. 3.

Effect of the mead acid (MA) diet on 50 mg/kg N-methyl-N-nitrosourea (MNU)-induced retinal damage in the postweaning rats in experiment 2. (a) Histology of the central retina in rats with or without MNU that were fed 2.4% MA and basal diets. At 7 days after a single ip injection of MNU, the outer nuclear layer and photoreceptor layer disappeared in the rats fed the MA and basal diets. GCL, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL, outer plexiform layer; ONL, outer nuclear layer; PRL, photoreceptor cell layer; and PEL, pigment epithelial cell layer. HE staining, ×200. (b) Photoreceptor cell ratio in the central retina 7 days after a single ip injection of MNU in rats fed 2.4% MA and basal diets. Rats treated with 50 mg/kg MNU and fed MA the diet do not have any statistically significant changes in their photoreceptor ratios at the central retina compared with the MNU-treated basal diet group. The index is calculated as [(outer retinal thickness / total retinal thickness) ×100]. (c) Retinal damage ratio in MNU-treated rats fed 2.4% MA and basal diets. Rats fed the MA diet do not show any statistically significant changes in the retinal damage ratio compared with the MNU-treated basal diet group. The index was evaluated as [(length of retina composed of less than four photoreceptor cells / whole retinal length) × 100]. The mean ± SE of the five rats in each treatment group is shown.

Fig. 4.

Effect of the mead acid (MA) diet on 40 mg/kg N-methyl-N-nitrosourea (MNU)-induced retinal damage in adult rats in experiment 3. (a) Histology of the central retina in rats with or without MNU who were fed the 2.4% MA and basal diets. At 7 days after a single ip injection of MNU, the outer nuclear layer and photoreceptor layer disappeared in the rats fed the MA and basal diets. GCL, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL, outer plexiform layer; ONL, outer nuclear layer; PRL, photoreceptor cell layer; and PEL, pigment epithelial cell layer. HE staining, ×200. (b) Photoreceptor cell ratio in the central retina 7 days after a single ip injection of MNU in rats fed the 2.4% MA and basal diets. MNU-treated rats fed the MA diet have a statistically significant decrease in their photoreceptor ratio at the central retina, which is similar to that seen in the MNU-treated basal diet group. The index is calculated as [(outer retinal thickness / total retinal thickness) ×100]. (c) Retinal damage ratio in MNU-treated rats fed the 2.4% MA and basal diets. Rats fed the MA diet do not show any statistically significant changes in the retinal damage ratio, which is similar to that seen in the MNU-treated basal diet group. The index is evaluated as [(length of retina composed of less than four photoreceptor cells / whole retinal length) ×100]. The mean ± SE of the six or seven rats in each treatment group is shown.

Fig. 5.

Fatty acid composition of the serum, lens and retina of the vehicle-treated rats. In the serum of the rats fed the 2.4% mead acid (MA) diet, the MA composition of the total fatty acids was higher than that seen in the serum of the rats fed the basal diet. In the lens and retina of the rats fed the 2.4% (MA) diet, although the MA composition was higher, the differences were not remarkable when compared with the serum values. The mean ± SE of four rats in the basal group and three rats in the MA diet group are shown.