| ObjectiveCongenital heart disease ( CHD) is a kind of congenital malformation that does severe harm to inborn' health and the morbidity is from 4%c to 50%c. About 80% percent of congenital heart diseases only manifest malformations in cardiovascular system without any abnormality of other systems, which is called simple congenital heart disease. It is considered that congenital heart disease results from the abnormal development of the embryonic cardiovascular system and the genetic factors play an important role in the pathogenesis of CHD. The heritabil-ity of CHD is from 55% to 65% , but it is not clear about the hereditary' patterns and penetrance, and little advances have been made in the researches on the susceptibility genes of CHD. Therefore, it is of great significance that the molecular genetic mechanisms of CHD should be discussed, which can provide the theorical bases for the genetic interfere of CHD, enrich the theory and practical application of human genomics and offer an useful method from which researches on other congenital malformations can draw lessons.Cardiac development is a complicated progress that involves not only spatial and temporal expression of many genes but also the migration, differentiation, proliferation of cells and accurate interaction between cells. With the studies on the cardiac Development of Drosophila, zebrailsh, chick embryo and mouse, some cardiac development - related genes have been identified by the means of fluorescence in situ hybridization, immumohistochemistry or gene knockout, such as Nkx2 -5, Pitx2, erbB2, MEF2C, dHAND, Endothelin, Irx4, Ufd1, NF - ATc, Sox - 6 and so on. The highly conservative characteristics of thesegenes provide an useful clue for us to look for cardiac development - related genes.At present, researches on human CHD mainly focus on the environmental factors at early stage of pregnancy and many syndromes with cardiac malformations. Candidate regions for cardiac malformations such as 22qll, 10pl3, 4pl6 and CHD related genes such as TBX5, Jagged - 1, TFAP2B, are identified. However, only Nkx2 - 5/Csx is confirmed with regard to the pathogenesis of simple CHD. Therefore, little has been known about the pathogenesis of human simple CHD and further researches are urgent.Previous researches in our group showed that D12S1056 locus in the region of 12ql3 was remarkably associated with simple CHD ( P < 0. 01) , suggesting there might be simple CHD susceptibility genes around D12S1056. In order to confirm it, ten microsatellite DNA markers were chosen around D12S1056, that is, D12S1056, D12S1293, D12S83, D12S1655, D12S1662, D12S334, D12S137, D12S102, D12S1702 and D12S1691. Genotyping and transmission disequilibrium test were done in 186 members from 62 core pedigrees of simple CHD. For the first time, the simple CHD susceptibility gene was narrowed to 3.4cM in 12ql3. At the same time, Gli gene in the candidate region was firstly confirmed associated with simple CHD using candidate cloning stragety.MethodsSamples186 members from 62 core pedigrees include 37 patients with ventricular septal defect, 8 patients with atrial septal defect, 10 patients with Patent Ductus Arterisus, and 7 patients with Fallot s Tetralogy. All patients had typical manifestation and were confirmed by cardiac ultrasonics and surgical operation. Vein-ous blood of 186 members from 62 core pedigrees were kept at -20C for use after anti - coagulation.Choose of microsatllite DNA markersAccording to the number of alleles, heterozygosity and polymorphic information content, 10 microsatellite DNA markers in 12q13 were chosen for TDT.Among these microsatellite DNA markers, D12S1056 and D12S1293 were from the Cooperative Human Linkage Center (CHLC) , while D12S83, D12S1655, D12S1662, D12S334, D12S137, D12S102, D12S1702 and D12S1691 were from Genome DataBase (GDB).GenotypingDNA was extracted from veinous blood routinely and all microsatellite DNA markers were amplified by fluorencent - labeling polymerase chain reaction. PCR products were pu...
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