ABSTRACT
Background Researches determined that ELOVL4 gene is a disease-causing gene of Stargard tmacular dystrophy and is a elongation enzyme of very long chain fatty acids.Stargardt type Ⅲ(STGD3) may be associated with the metabolism of extensive enzyme of very long chain fatty acids.To explore the effect of ELOVL4 in STGD3 and its relationship with the metabolism of extensive enzyme of very long chain fatty acids is of important clinical significance.Objective This study was to determine the role of ELOVL4 gene for the pathogenesis of STGD3.Methods Recombinant adenovirus type 5 carrying mouse ELOVL4 gene and green fluorescent protein (GFP) was transfected into pheochromocytoma cells (PC12 cells),and then the cells were divided into PC12 group,PC12+Ad5-GFP group and PC12+AdS-ELOVL4 group.Ad-GFP or Ad-ELOVL4 was added into the culture medium for 24 hours with the virus plasmid 1 × 104-2× 104 pfu/ml.Expression of ELOVL4 mRNA in the PC12 was quantified by quantitative real time PCR(qRT-PCR) and was described as relative value to the expression of RPL19.ELOVL4 protein was assayed by Western blot.The transfected cells were treated with arachidonic acid (AA,20:4n6),eicosapentaenoic acid (EPA,20:5n3) and docosahexaenoic acid (DHA,22:6n3) individually for 48 hours.The cells were collected,and total lipids were extracted,and fatty acid methyl esters were prepared and analyzed by gas chromatography-mass spectrometry (GC-MS).Results The relative expression levels of ELOVL4 mRNA in PC12 cells in the PC12+Ad5-ELOVL4 group,PC12+Ad5-GFP group and PC12 group were 0.833± O.138,0.027t±0.002 and 0.024 ±0.002,with a significant difference among the 3 groups (F =102.700,P =0.000),and relative expression levels of PC12+Ad-ELOVL4 were 30-40 folds more than those in the PC12 group and the PC12+Ad-GFP group.Western blot assay showed a stronger response band for ELOVL4 protein in the PC12+Ad-ELOVL4 group.GC-MS found that abundant polyunsaturated fatty acids (C28-C38) were synthesized by PC12 cells in the PC12+Ad-ELOVL4 group,with the more levels in C34 and C36.Conclusions ELOVL4 can promote the synthesis of C28-C38 polyunsatured fatty acid in PC12 cells,which offers a novel clue for the treatment of STGD3.
ABSTRACT
Objective To establish an experimental retinal artery occlusion (RAO) model by means of photochemistry therapy with 532 nm frequency-doubled laser, and to evaluate the feasibility of this method. Methods Ten eyes from ten healthy adult New Zealand rabbits were intravenously injected 5% rose bengal solution, then their monocular retinal artery was irradiated by 532 nm frequency-doubled laser. Both eyes of these rabbits were examined by direct ophthalmoscope and fundus photographic camera on the 1st hour, 24th hour and 3rd day. Meanwhile, the experimental eyes were examined by fundus fluorescein angiography (FFA). These rabbits were killed on the 3rd day, experimental eyes were enucleated, the retina was observed under light microscope. Results There were 9 eyes approved having RAO by direct ophthalmoscope and FFA. Thrombi in retinal arterials could be found by light microscope in all these 9 experimental eyes, and the walls of blood vessel were intact without any obvious necrosis or attenuation. Conclusions RAO animal model can be induced by photochemistry therapy with 532 nm frequency-doubled laser. There are thrombi in the retinal artery without trauma of the artery wall. Photochemistry therapy is an efficient, secure and reproducible method to induce animal model of RAO.