Transgenic mouse as a model of lipoprotein glomerulopathy (LPG), and analysis of LPG

脂蛋白肾小球病（LPG）是1988年新发现的一种肾脏疾病。其特征是肾小球毛细血管明显扩张的脂蛋白沉积异常，血浆载脂蛋白（apo） e浓度升高。最近，我们在三个日本LPG亲属中发现了apoE变体apoE2 Sendai （Arg 145Pro）。此外，我们还发现了三种不同的apoE变体，apo E2 Kyoto （Arg25Cys）、apo E1 （del 141-143）和apo E1 (del 156-173)，它们与LPG密切相关。建立载脂蛋白e1 （del 141-143）转基因小鼠作为液化石油气动物模型。采用定点诱变方法将ApoE1 （del 141 ~ 143）突变引入正常的apoE基因及其启动子区（11.5kb）。纯化了HindIII酶切的人apoE变异基因片段，并将其微注射到C57BL/6小鼠受精卵原核中。注射的卵子通过手术转移到麻醉的代孕雌性的输卵管中。然而，转基因小鼠由于感染了致病性病毒而被杀死。第二次显微注射完成后，产下了7只幼崽。从它们的尾巴上分离出基因组DNA。用引物对人apoE基因基因组尾部DNA进行聚合酶链反应分析。从一只雌性小鼠身上检测到人apoE基因。人apoE基因在Fl小鼠中的检出率很低，目前在2只雄性小鼠中证实了人apoE基因。建立apoE1 （del 141-143）小鼠纯合子后，计划分析其肾小球病变和血清脂蛋白。

Lipoprotein glomerulopathy (LPG) is a newly recognized renal disease in 1988. It is characterized by abnormal lipoprotein deposition in the marked dilated glomerular capillaries, and elevated plasma concentration of apolipoprotein (apo) E.Recently, we identified an apoE variant, apoE2 Sendai (Arg 145Pro) in three Japanese kindred with LPG.Furthermore, we discovered three different novel apoE variants, apo E2 Kyoto (Arg25Cys) apo E1 (del 141-143) and apo E1 (del 156-173), which are closely related to LPG.Establishment of an apo E 1 (del 141-143) transgenic mouse was tried as an animal model of LPG.ApoE1 (del 141-143) variant was introduced into normal apoE gene including promoter region (11.5kb) using site directed mutagenesis. Fragment of the variant human apoE gene digested with HindIII was purified and microinjected into pronuclei of fertilized eggs of C57BL/6 mice. Injected eggs were surgically transferred to oviducts of anesthetized surrogate females. However, the transgenic mice were killed because they were infected with pathogenic virus. Second microinjection was done, and seven pups were born. Genomic DNA was isolated from their tails. Polymerase chain reaction analysis was performed on genomic tail DNA using primer set of human apoE gene. Human apoE gene was detected from a female mouse. Human apoE gene detection rate of Fl mice was very low, and human apoE gene was confirmed currently in two male mice. After homozygotes of apoE1 (del 141-143) mice are established, analysis of their glomerulus lesions and serum lipoproteins are planned.