[Status of iron metabolism and erythropoietic proliferation in children with various genotypes of thalassemia].

OBJECTIVE: To study the status of iron metabolism and erythropoietic proliferation in children with various genotypes of thalassemia. METHODS: Serum concentrations of ferritin (SF), transferrin receptor (sTfR) and erythropoietin (EPO) were measured in 158 children with thalassemia. The differences in the concentrations of the three indices among children with different genotypes of thalassemia were compared. The correlations of the hemoglobin level with sereum SF, sTfR and EPO levels were assessed. RESULTS: Among the 158 children with thalassemia, 52(32.9%) were diagnosed with alpha-thalassemia minor, 27(17.1%) with HbH disease, 59(37.4%) with beta-thalassemia minor, 13(8.2%) with beta-thalassemia major, and 7(4.4%) with combining alpha beta thalassemia. The SF levels in children with HbH disease or beta-thalassemia major were significantly higher than those in the other thalassemia groups (P<0.01). The sTfR levels in children with beta-thalassemia major were the highest when compared with those in the other thalassemia groups (P<0.05). The EPO levels in children with beta-thalassemia major were also the highest when compared with those in the other thalassemia groups (P<0.01). There was a negative correlation between hemoglobin and EPO levels in children with HbH disease (r=-0.656, P<0.01) and beta-thalassemia major (r=-0.641; P<0.05). CONCLUSIONS: The status of iron metabolism and erythropoietic proliferation is different in children with different genotypes of thalassemia. A combined measurement of SF, sTfR and EPO may reflect the status of erythropoietic proliferation.

[Application of PCR-DGGE technique in G-6-PD deficiency].

OBJECTIVE: To detect gene mutations of children with glucose-6-phosphorate dehydrogenase (G-6-PD) deficiency and of carriers of G-6-PD deficiency gene with the technique of polymerase chain reaction and denatured gradient gel electrophoresis (PCR-DGGE), and to explore the value of the technique in the diagnosis of G-6-PD deficiency and G-6-PD deficiency gene carrying. METHODS: cDNAs were harvested by reverse transcription method after RNAs had been extracted from peripheral blood of 43 children with G-6-PD deficiency and of their family members (36 lineages). Electrophoresis behaviors of the fragment from exons 11-12 of G-6-PD cDNA were detected with the technique of PCR-DGGE. Gene sequencing was then performed for the abnormal electrophoresis bands. RESULTS: Abnormal electrophoresis bands were found in the 1304-1520 fragment of G-6-PD cDNA in 33 out of 36 family lineages. The G-6-PD/6-PGD ratio was below 1.00 in 9 mothers of patients. Three of them had the G-6-PD/6-PGD ratio lower than 0.50. The PCR-DGGE bands were the same in the 3 mothers. Gene sequencing showed double heterozygote in the 3 mothers, but the maternal carriers of G-6-PD deficiency gene who had normal G-6-PD/6-PGD ratio showed mono-heterozygote in gene sequencing. Three mutational sites were found in the 1304-1520 fragment, i.e., C1311TG1376T and G1388A. The electrophoresis behaviors were different among the 3 gene mutational sites. CONCLUSIONS: PCR-DGGE is a sensitive and reliable technique in the screening of gene mutations. It is useful in the diagnosis of G-6-PD deficiency, especially in the diagnosis of female G-6-PD deficiency gene carrying.

[Expression of G-6-PD mRNA in children with G-6-PD deficiency].

OBJECTIVE: To detect the level of glucose-6-phosphate dehydrogenase (G-6-PD) mRNA expression in children with G-6-PD deficiency and their lineal family members in order to explore possible mechanisms of the disease at the transcriptional level. METHODS: RNA was extracted from peripheral blood of 41 children with G-6-PD deficiency and of their lineal family members (29 father lineages and 40 mother lineages). cDNA was then harvested using the reverse transcription method. G-6-PD mRNA expression was detected by quantitative real-time PCR (QRT-PCR). The detection results of G-6-PD mRNA expression in three groups (Patient, Father lineage and Mother lineage) were compared using Statistical Software SPSS 10.0 system. RESULTS: The mean G-6-PD mRNA value in the Patient, Father lineage and Mother lineage groups were 0.57 +/- 0.19, 0.74 +/- 0.21 and 0.67 +/- 0.21 respectively. The G-6-PD mRNA value in the Patient group was significantly lower than both Father lineage (t = -3.18, P < 0.01) and Mother lineage groups (t = -2.54, P < 0.05). CONCLUSIONS: The expression of G-6-PD mRNA decreased in children with G-6-PD deficiency, suggesting that the pathogenesis of this disorder relates to the varying levels of gene transcription.