OVEREXPRESSION OF HUMAN LECITHIN CHOLESTERYL ACYLTRANSFERASE IN TRANSGENIC MICE

为了评估LCAT在HDL代谢中的作用，使用6.2 kb的 利用人LCAT基因的片段（hLCAT）来开发三种人LCAT基因。 在血浆水平过表达hLCAT的单独转基因小鼠系 分别是对照组（1 μ g/ml）的10、14和100倍。LCAT活动 45只杂合子和纯合子小鼠，范围为582 ± 92， 3695q340 nmol/ml/h（NL=31q4 nmol/ml/h）。北方印迹杂交 分析表明hLCAT在小鼠肝脏中的组织特异性表达。 与24个年龄和性别匹配的兄弟姐妹相比，转基因小鼠具有更高的 血浆TC（NL的133-237%）、CE（NL的141-267%）和HDL-C（NL的123-209%） NL），但甘油三酯、PL、含B 脂蛋白，apoA-I和apoA-II。hLCAT TG血浆的FPLC分析 显示了富含CE和PL的较大尺寸的HDL颗粒。 年龄/性别匹配的转基因动物（M=7，F=8）和对照动物（M=14，F=13） 高胆固醇-脂肪（HF）饮食21天，以研究 LCAT在调节饮食反应中的潜在作用。饮食前脂质 对照组TC= 97 ± 11，HDL= 72 q10， 转基因组TC=121 ± 16，HDL-C=84 ± 17。饮食后 对照组的血脂值为TC= 290 ± 55，HDL= 85 ± 15， 转基因组TC= 313 ± 83，HDL= 115 ± 27。因此，在本发明中， 在HF饮食中，转基因小鼠具有显著较高的（小于0.05） 与对照组相比，HDL-C以及TC/HDL比值降低，无差异 Tg、PL、CE、LCAT质量和活性。转基因小鼠的FPLC分析 血浆HDL-C、CE和PL显著升高， IDL/LDL-C、CE和PL降低。 进行小鼠125 I-apoA-I和131 I-apoA-II动力学研究， 研究LCAT过表达导致的潜在机制 HDL水平升高。apoA-I和apoA-II的催化剂延迟， 与对照组（FCR=2.6）相比，转基因组（N=5）（FCR=1.9和3.4d-1 和4.2d-1）小鼠。因此，转基因小鼠的高脂蛋白血症 hLCAT的过表达是HDL的延迟催化的结果。

In order to evaluate the role of LCAT in HDL metabolism, a 6.2 kb fragment of the human LCAT (hLCAT) gene was utilized to develop three separate transgenic mouse lines overexpressing hLCAT at plasma levels 10,14 and 100 fold higher than control (1 ug/ml) mice. LCAT activity in 45 heterozygous and homozygous mice ranged from 582plus/minus 92 to 3695q340 nmol/ml/h (NL=31q4 nmol/ml/h). Northern blot hybridization analysis demonstrated tissue specific expression of hLCAT in mouse liver. Compared to 24 age and sex-matched siblings, transgenic mice had elevated plasma TC (133-237% of NL), CE (141-267% of NL) and HDL-C (123-209% of NL) but similar plasma levels of triglycerides, PL, B-containing lipoproteins, apoA-I and apoA-II. FPLC analysis of hLCAT TG plasma revealed larger sized HDL particles enriched in CE and PL. Age/sex matched transgenic (M=7, F=8) and control (M=14, F=13) animals were then placed on a high chol-fat (HF) diet for 21 d to investigate the potential role of LCAT in modulating dietary responses. Pre-diet lipid values (mg/dl) in controls were TC=97plus/minus11, HDL=72q10 and in transgenics were TC=121 plus/minus16, HDL-C=84 plus/minus17. Post-diet lipid values in controls were TC=290plus/minus55, HDL= 85plus/minus15, and in transgenics were TC=313plus/minus83, HDL=115plus/minus27. Thus, on the HF diet transgenic mice had significantly higher (pless than0.05) HDL-C as well as reduced TC/HDL ratios than controls, without differences in Tg, PL, CE, LCAT mass and activity. FPLC analysis of transgenic mouse plasma revealed significant increases in HDL-C, CE and PL with reciprocal decreases in IDL/LDL-C, CE and PL. Mouse 125I-apoA-I and 131I-apoA-II kinetic studies were performed to investigate the underlying mechanisms by which LCAT overexpression leads to increased HDL levels. ApoA-I and apoA-II catabolism was delayed in transgenic (N=5) (FCR=1.9 and 3.4d-1) when compared to control (FCR=2.6 and 4.2d-1) mice. Thus, hyperalphalipoproteinemia in transgenic mice overexpressing hLCAT results from delayed catabolism of HDL.