Résumé
Objective To explore the pathogenesis of Russell-Silver syndrome (RSS). Methods Two milliliter peripheral blood samples were collected from 6 male patients aged 6 to 8 years with suspected RSS phenotype, the parents of 2 patients and 5 healthy boys. Mononuclear cells were isolated and genomic DNA was extracted. The methylation level of the H19 imprinting control region(ICR)1 on chromosome 11p15.5 was detected by pyrosequencing.The methylation status and the copy number variation in the corresponding region of one RSS patient with positive results by pyrosequencing were analysed by methylation-specific multiplex-ligation-dependent probe amplification assay (MS-MLPA). Results Pyrosequencing analysis revealed that the methylation rates on the 6 CpG targeting sites in H19 differentially methylated region(DMR)in the 6 RSS patients were about 11%~29%, which were significantly lower than those in their parents and normal controls (44%~59%). The MS-MLPA results of one patient with positive pyrosequencing showed that the methylation rates of 4 sites in H19-DMR were about 10%,which was obviously lower than the normal level.The methylation rates of the 4 sites in KCNQ1OT1 gene were about 50%, which was in the normal range. The copy number variations from all samples detected were in the normal range. Conclusion There is methylation aberration of H19-DMR in ICR1 in children with RSS.
Résumé
Objective To evaluate the application of high-performance liquid chromatography (HPLC) in diagnosis and screening of thalassemia. Methods Automated HPLC was used to measure HbF and HbA2 in 100 genetically diagnosed thalas-semic patients and 35 normal children. The results were compared with those from traditional tests including alkali denaturation test and cellulose acetate electrophoresis. The diagnose accordance rates, sensitivity and specificity were compared. Results Seventy-fourβthalassemia, 64 were heterozygous with single mutations and 10 were compound heterozygous with double muta-tions. Twenty-sixαthalassemia, 25 were compound mutations and one was heterozygous with single mutation. The HbF percent-age from HPLC was higher than that from alkali denaturation tests in either thalassemia or normal children (P<0.01). HbF level from HPLC inα-thalassemia was signiifcantly different from that in the normal children (P=0.011). The percentage of HbA2 from HPLC was higher than that from cellulose acetate electrophoresis (P=0.010). HbA2 in the single heterozygousβ-thalassemia were twice higher than that in the double heterozygous mutatedβ-thalassemia (P<0.01). The combination of HbF-HbA2 (≥4.0%) from HPLC with MCV (<80 lf) and MCH (<27 pg) had high accordance rates (99.3%), sensitivity (99.0%) and speciifcity (100.0%) in diagnosis of thalassemia. Conclusions When the results of HPLC are combined with MCV and MCH, it can be applied to the diagnosis of thalassemia with high speciifcity, high sensitivity and has high diagnostic accordance rate with genetic results. HPLC can be an ideal approach to screenβthalassemia.