ABSTRACT
Objective To investigate the effect of alteration of CRELD1 gene expression on the related genes in the endocardial cushion development.Methods Over-expression and silence of CRELD1 gene were realized by the construction of lentiviral vector.Afterwards,the lentiviral vectors were used to infect the human fetal lung fibroblasts (HFL)-I.All of the cells were divided into the following 5 groups:the blank control group,the negative control group of interference,the interference group,the negative control group of over-expression,and the over-expression group.Western blot and real-time fluorescent quantitative polymerase chain reaction were applied to examine the mRNA and protein expression of CRELD1,Sox9,Aggrecan,Scleraxis and Tenascin-C.Results The DNA sequences of 2 recombinant plasmids pLV3-shRNA-CRELD1 and pLV4-CRELD1 matched very well with those which were designed according to the DNA sequence analysis.HFL-I was successfully infected with lentiviral vectors and displayed fluorescent green light under inverted fluorescence microscope.The results of real-time PCR detection and Western blot test were consistent:expressions of Sox9 and Aggrecan in the interference group were significantly higher than those in the negative control group of interference,while the expressions of the 2 genes in the over-expression group were significantly lower than those in the negative control group of over-expression.Expressions of Scleraxis increased in both the interference group and the over-expression group when compared with the negative control groups respectively.Compared to the corresponding negative control groups,Tenascin-C expression decreased markedly in the interference group,whereas it increased significantly in over-expression group.Conclusions CRELD1 gene has negative effect on the expression of the related genes Sox9 and Aggrecan in the endocardial cushion development,whereas it has positive effect on the Tenascin-C expression.It serves as a theoretical framework to illustrate the effect of CRELD1 gene on the atrioventricular septal defect.
ABSTRACT
Objetivo: Comparar o crescimento volumétrico total do coraçäo (C), do miocárdio ventricular (MV) e dos coxins endocárdicos (CE) entre si e com o crescimento do embriäo. Coleçäo de embriöes humanos seccionados em série e corados (27 embriöes) no período pós-somítico. Os volumes de C, MV e CE foram determinados morfometricamente e correlacionados estatisticamente ao comprimento vertex-coccyx (V-C) dos embriöes pela equaçäo alométrica Y = a x**b. O crescimento cardíaco e de seus componentes MV e CE apresentou correlaçöes estatisticamente significativas (p<0,01), mas alometricamente negativas, em relaçäo ao crescimento de V-C. Isto indica que o coraçäo apresenta, mais intensamente modificaçöes na forma que no tamanho. Entretanto, em relaçäo ao coraçäo, MV é a estrutura que apresenta maior aumento relativo no segundo mês gestacional. Comparativamente, CE decresce nesse período e termina com menos de 3% do volume final do coraçäo. Os CE näo têm a importância anteriormente atribuída no desenvolvimento das valvas e dos septos cardíacos. Pelo contrário, alteraçöes no crescimento do MV devem interferir profundamente no desenvolvimento cardíaco
Purpose: To compare the volumetric growth of the whole heart (WH), ventricular myocardium (VM), and endocardial cushions (EC) among each other and in relation to the growth of the embryo. Patients and Methods: Collection of human embryos serially sectioned and stained (27 embryos) in post-somitic period. The volumes of WH, VM and EC were morphometrically determined and statistically correlated to the embryos crown-rump length (C-R) by using the allometric equation Y = a xb. Results: The cardiac growth, and the growth of the VM and EC, presented significant (p < 0.01) but allometically negative correlations in relation to C-R length. It indicates that cardiac changes in shape are more pronounced than in size. However, relative to the heart itself VM is the component that presents greater volumetric increase during the second month of gestation. EC decreases and end the embryonic period proper with less of 3% of the cardiac volume. Conclusion: Our quantitative results agree with more recent morphological studies which consider EC with small significance in valves and septa development, probably functioning more as a plastic component than in genesis of cardiac structures. On the contrary some lack in growth of the VM should disturb deeply cardiac development.