RESUMO
BACKGROUND: Phenylketonuria is caused by gene mutation of phenylalanine hydroxylasel (PAH), which is mainly induced by permutation, short segments and insertion of base.OBJECTIVE: To evaluate the gene mutation of phenylalanine hydroxylasel in phenylketonuria in Hui nationality.DESIGN: Open study.SETTING: Urumqi General Hospital of Lanzhou Military Area Command of Chinese PLA; Capital Pediatrics Institute.PARTICIPANTS: A boy of Hui nationality in China and aged 3.1 years was selected in this study. The boy had intellect hysteresis in his one year and received medical treatment in his three years, while he was diagnosed as cerebral paralysis. After repeatedly inefficient treatment, he was hospitalized in our hospital on December 13, 2004. Iron sesquichloride in urine was strongly positive and concentration of serum phenylalanine was 1 680 μmol/L; therefore, he was diagnosed as the typical phenylketonuria.METHODS: 5 mL venous blood was selected from the boy and his parents, respectively, and anticoagulated with EDTA-Na2. DNA in gene group was extracted by using typical phenol/chloroform method. In addition, polymerase chain reaction (PCR) primer sequence of extron 7, 6, 11, 3, 12 and 5 of PAH gene was designed based on references. And then, PCR products were detected with 2% agarose gel electrophoresis. 5 μL PCR products were mixed with the same volume of degenerated buffer solution, degenerated at 97 ℃ for 5 minutes, put in iced bath and performed with 80 g/Lnon-degenerated polyacrylamide gel electrophoresis. After that, the products were dealt with sliver staining routinely, and single strand DNA banding patterns were analyzed and recorded. ABI377 automatic sequenator (PE Company) was used to detect PCR sequence and purify PCR product in Shanghai Boya Biotechnology Company.MAIN OUTCOME MEASURES: Iron sesquichloride in urine, concentration of serum phenylalanine and mutant gene types of phenylalanine hydroxylase.RESULTS: Extron 7, 6, 11, 3, 12 and 5 of PAH gene were analyzed in the boy and his parents. The results demonstrated that SSCP electrophoresis in extron 6 was different from that in the normal control group. Site of electrophoresis strip of his father was coincident with that of his mother, but different from that of the boy. Sequencing results indicated that point mutation (cytosine replaced by thymine), which was a R176X mutant heterozygote, occurred at the 526th site of cDNA of phenylalanine hydroxylase gene in his parents; however, two chromosomes of the boy had mutation at the same site, which was R176X mutant homozygote.CONCLUSION: Mutation of R176X homozygote of phenylketonurea is firstly reported in Hui nationality in China.
RESUMO
Objective To study the expression of a virulent novel gene F10 associated with hydatidiform mole in the hepatoma carcinoma cells(HCC),and to explore the effects of F10 on the occurrence and development of primary hepatocellular carcinoma(PHC).Methods The full-length cDNA sequence of F10 was obtained from the samples of early pregnant chorionic villi and hydatidiform mole by employing suppression subtractive hybridization and cDNA microarray techniques.The hepatocarcinoma tissues and their surroundings were examined by in situ hybridization with digoxigenin-labeled of F10 gene.The real-time-fluorescence quantitative polymerase chain reaction(FQ-PCR)was used to quantitatively detect the expression level of F10 gene in primary hepatocellular carcinoma.F10 gene RNA copies in the samples were examined in the hepatocarcinoma tissues samples and their surroundings with PHC by PE ABI 7000 Sequence Detector.Results F10 mRNA was detected in hepatocarcinoma cells with positive signals found in kytoplasm and cell membrane,but not detected in the tissues surrounded the hepatocarcinoma.In 6 of 8 PHC samples,the expression Level of F10 gene was higher in tumor tissues than that in the tissues surrounded the tumor tissue.Among them 2 cases showed more than 4 times over-expression of F10 gene in tumor cells than in the surrounded tissues.Conclusion The virulent new F10 gene associated with hydatidiform mole was mainly expressed in hepatocellular carcinoma cells.The results of present study suggested that the higher expression of F10 gene not only showed a close relation to trophoblastic tumor,but also indicated that the F10 gene may play an important role in the occurrence and development of primary hepatocellular carcinoma.
