国际眼科杂志
國際眼科雜誌
국제안과잡지
INTERNATIONAL JOURNAL OF OPHTHALMOLOGY
2014年
8期
1386-1390
,共5页
余曼%林伟%陈波%吴峥峥
餘曼%林偉%陳波%吳崢崢
여만%림위%진파%오쟁쟁
二十碳五烯酸%花生四烯酸%ELOVL4基因%Stargardt病%超长链多不和脂肪酸
二十碳五烯痠%花生四烯痠%ELOVL4基因%Stargardt病%超長鏈多不和脂肪痠
이십탄오희산%화생사희산%ELOVL4기인%Stargardt병%초장련다불화지방산
eicosapentaenoic acid%arachidonic acid%ELOVL4 gene%Stargardt’s disease%very long chain polyunsaturated fatty acid
目的：ELOVL4是常染色体显性Stargardt氏黄斑变性的致病基因,ELOVL4蛋白酶参与n3和n6超长链多不饱和脂肪酸( very long chain polyunsaturated fatty acid,VLC-PUFA)的合成,比较两者的生物合成效率,对治疗Stargardt氏黄斑变性有指导意义。<br> 方法：构建携带ELOVL4基因和绿色荧光蛋白的重组腺病毒,转入培养的PC12细胞,将细胞分成三组：PC12、PC12+Ad-GFP 和 PC12+Ad-ELOVL4,前两组为对照组,通过qRT-PCR定量分析 ELOVL4基因的表达量, Western Blot检测 ELOVL4蛋白的表达；等浓度(1：1)加入 EPA ( n3 PUFA)和AA(n6 PUFA),孵育48h之后进行脂肪酸提取,通过气相色谱-质谱法(gas chromatography-mass spectrometry, GC-MS)分析超长链脂肪酸的成分。<br> 结果：GC-MS检测到分别用EPA和AA处理后的PC12+Ad-ELOVL4的细胞中有n3 VLC-PUFA的表达,34：5 n3和36：5n3是20：5n3/EPA的主要产物,分别为0.71%和1.6%；34：4 n6和36：4 n6是20：4 n6/AA的主要产物,分别为0.46%和0.61%；EPA所产生的n3 VLC-PUFAs总和是AA所产生的n6 VLC-PUFAs总和的2倍。<br> 结论：在ELOVL4蛋白作用下,EPA合成VLC-PUFAs的效率高于AA,饮食中给予适当比例的n3/n6 PUFAs,可能是治疗STGD3疾病的方式之一。
目的：ELOVL4是常染色體顯性Stargardt氏黃斑變性的緻病基因,ELOVL4蛋白酶參與n3和n6超長鏈多不飽和脂肪痠( very long chain polyunsaturated fatty acid,VLC-PUFA)的閤成,比較兩者的生物閤成效率,對治療Stargardt氏黃斑變性有指導意義。<br> 方法：構建攜帶ELOVL4基因和綠色熒光蛋白的重組腺病毒,轉入培養的PC12細胞,將細胞分成三組：PC12、PC12+Ad-GFP 和 PC12+Ad-ELOVL4,前兩組為對照組,通過qRT-PCR定量分析 ELOVL4基因的錶達量, Western Blot檢測 ELOVL4蛋白的錶達；等濃度(1：1)加入 EPA ( n3 PUFA)和AA(n6 PUFA),孵育48h之後進行脂肪痠提取,通過氣相色譜-質譜法(gas chromatography-mass spectrometry, GC-MS)分析超長鏈脂肪痠的成分。<br> 結果：GC-MS檢測到分彆用EPA和AA處理後的PC12+Ad-ELOVL4的細胞中有n3 VLC-PUFA的錶達,34：5 n3和36：5n3是20：5n3/EPA的主要產物,分彆為0.71%和1.6%；34：4 n6和36：4 n6是20：4 n6/AA的主要產物,分彆為0.46%和0.61%；EPA所產生的n3 VLC-PUFAs總和是AA所產生的n6 VLC-PUFAs總和的2倍。<br> 結論：在ELOVL4蛋白作用下,EPA閤成VLC-PUFAs的效率高于AA,飲食中給予適噹比例的n3/n6 PUFAs,可能是治療STGD3疾病的方式之一。
목적：ELOVL4시상염색체현성Stargardt씨황반변성적치병기인,ELOVL4단백매삼여n3화n6초장련다불포화지방산( very long chain polyunsaturated fatty acid,VLC-PUFA)적합성,비교량자적생물합성효솔,대치료Stargardt씨황반변성유지도의의。<br> 방법：구건휴대ELOVL4기인화록색형광단백적중조선병독,전입배양적PC12세포,장세포분성삼조：PC12、PC12+Ad-GFP 화 PC12+Ad-ELOVL4,전량조위대조조,통과qRT-PCR정량분석 ELOVL4기인적표체량, Western Blot검측 ELOVL4단백적표체；등농도(1：1)가입 EPA ( n3 PUFA)화AA(n6 PUFA),부육48h지후진행지방산제취,통과기상색보-질보법(gas chromatography-mass spectrometry, GC-MS)분석초장련지방산적성분。<br> 결과：GC-MS검측도분별용EPA화AA처리후적PC12+Ad-ELOVL4적세포중유n3 VLC-PUFA적표체,34：5 n3화36：5n3시20：5n3/EPA적주요산물,분별위0.71%화1.6%；34：4 n6화36：4 n6시20：4 n6/AA적주요산물,분별위0.46%화0.61%；EPA소산생적n3 VLC-PUFAs총화시AA소산생적n6 VLC-PUFAs총화적2배。<br> 결론：재ELOVL4단백작용하,EPA합성VLC-PUFAs적효솔고우AA,음식중급여괄당비례적n3/n6 PUFAs,가능시치료STGD3질병적방식지일。
AIM:To compare the synthesis efficiency of n3 and n6 very long chain polyunsaturated fatty acid ( VLC-PUFA ) by overexpressing ELOVL4 protein, providing guidance for treating Stargardt-like macular dystrophy (STGD3). <br> METHODS:To establish recombinant adenovirus with the ELOVL4 protein and green fluorescent protein, transferred into cultured PC12 cells. The cells were divided into 3 groups: PC12, PC12 + Ad- GFP and PC12 + Ad-ELOVL4, former two groups serve as controls. ELOVL4 gene expression was quantified by qRT-PCRs. ELOVL4 protein was analyzed by Western - Blot ( WB ) . The transduced cells were treated with both EPA and AA (1:1). After 48h of incubation, cells were collected, total lipids extracted and fatty acid methyl esters prepared and analyzed by gas chromatography-mass spectrometry ( GC-MS) . <br> RESULTS:When supplemented together, 20:5n3 (EPA) and 20:4n6 ( AA) were efficiently taken up at almost the same amounts in the PC12 cells regardless of ELOVL4 expression. The ELOVL4-expressing cells elongated both EPA and AA to a series of n3 and n6 VLC-PUFAs. From 20:5n3/EPA, 34:5n3 and 36:5n3 account for 0. 71% and 1.6%, respectively. From 20:4n6/DHA, 34:4n6 and 36:4n6 were only 0. 46% and 0. 61%, respectively. The total relative mol% of n3 VLC-PUFAs synthesized from EPA was almost two times that of n6 VLC-PUFAs synthesized from AA. <br> CONCLUSION: ELOVL4 protein preferentially elongates n3 PUFA to VLC - PUFAs over n6 PUFA. Dietary supplementation of appropriate n3/n6 PUFAs may provide STGD3 patients with some therapeutic benefits.
