全基因组扫描2型糖尿病KK/Ta小鼠白蛋白尿易感基因定位

Genome-wide Linkage Analysis of a Quantitative Trait Locus for the Development of Albuminuria in Diabetic KK/Ta Mice

目的利用全基因组扫描方法进行2型糖尿病KK/Ta小鼠白蛋白尿易感基因定位。方法以近交纯品系雌性BALB/c小鼠与雄性KK/Ta小鼠交配建立第一代杂交小鼠(F1),以雄性(KK/Ta×BALB/c)F1小鼠与雌性KK/Ta小鼠回交。提取208只雄性回交小鼠基因组DNA。根据扩增的101个微卫星DNA分离片段大小决定回交小鼠基因型,同时测定体质量、血糖、尿白蛋白及尿肌酐等表现型。采用基因定位专用软件(MAPMAKER/QTL,Map manager)进行数量性状位点(QTL)分析。结果 KK/Ta小鼠的体质量、空腹血糖水平、经腹腔葡萄糖耐量试验空腹与注射葡萄糖后2 h血糖之和显著高于BALB/c和F1小鼠(P<0.01)。20,28周龄的KK/Ta小鼠平均尿白蛋白水平显著高于BALB/c和F1小鼠(P<0.01)。20周龄与28周龄的白蛋白尿水平与第2条染色体83.0 cM D2Mit311微卫星DNA标记附近区域显著连锁,最大对数优势积分值(LOD)为3.5,我们将这一易感基因座位命名为尿白蛋白1(UA-1)。KK/KK组回交小鼠20周龄和28周龄的尿白蛋白水平显著高于KK/BALB组回交小鼠。结论与2型糖尿病KK/Ta小鼠尿白蛋白水平紧密连锁的易感基因座位位于第2条染色体83.0 cM处D2Mit311微卫星DNA标记附近区域(UA-1)。UA-1区域附近的生长激素释放激素基因GHrH、生长抑素受体4基因Smstr4、血栓调节蛋白(TM)基因Thbd等为糖尿病肾病的可能易感基因。

Objective To identify the Quantitative Trait Locus（QTL） for the development of albuminuria in diabetic KK/Ta mice.Methods Genome-wide linkage analysis was conducted.BALB/c female mice were mated with KK/Ta males to produce the F1 hybrid mice.KK/Ta ×（BALB/c×KK/Ta） F1 backcross mice were obtained by crossing female KK/Ta mice with male（BALB/c×KK/Ta） F1 mice.Genome DNA was extracted from 208 male KK/Ta×（BALB/c×KK/Ta） F1 backcross mice.Genome screening of KK/Ta-derived loci contributing to the development of albuminuria entailed genotyping of 208 male KK/Ta×（BALB/c×KK/Ta） F1 backcrossed mice with 101 microsatellite markers polymorphic between the two strains.Phenotypic values including body weight,blood glucose level and Urinary albumin：creatinine ratio etc.were monitored.Linkage analysis was performed using the Map Manager QT package program.Results The body weight,fasting blood glucose and the sum of blood glucose levels in the intraperitoneal glucose tolerance test（IPGTT） of KK/Ta mice were significantly higher than those of BALB/c and F1 mice（P 〈 0.01）.Levels of the mean urinary albumin/creatinine ratio in KK/Ta mice were significantly higher than those in F1 hybrid and BALB/c mice at 20 and 28 weeks of age（P 〈 0.01）.A genome-wide analysis of susceptibility loci for albuminuria with microsatellite-based chromosomal maps showed a contributing KK/Ta locus,provisionally designated UA-1（urinary albumin 1）,with a significant linkage with the interval on chromosome 2 at 83.0 cM close to the microsatellite marker D2Mit311 with a maximum log odds score（LOD） of 3.5.The progeny homozygous for KK/Ta allele for UA-1-linked D2Mit311 showed significantly higher urinary albumin levels than the other heterozygous for KK/Ta and BALB/c allele.Conclusion The susceptibility locus（UA-1） contributing to the development of albuminuria in diabetic KK/Ta mice locates on chromosome 2 at 83.0 cM close to the microsatellite marker D2Mit311.There are several potentially important candidate genes that may be relevant to diabetic nephropathy in the vicinity of UA-1,including growth hormone releasing hormone（GHrH）,somatostatin receptor 4（Smstr4） and thrombomodulin（Thbd） gene et al.