Identification of four novel mutations in the phenylalanine hydroxylase gene in patients with PKU / 中华检验医学杂志

Objective To investigate the distributions of PAH gene mutation and provide guidance for gene diagnosis and prenatal diagnosis of patients with PKU in Xinjiang of China.Methods A total of 15 patients (aged from 2 to 10 years, all with blood Phe concentration over 700 μmol/L) who visited Urumqi general hospital of Lanzhou Command were clinically diagnosed as PKU and were included in this study. PCR followed by DNA sequencing was performed to analyze the promoters, all the 13 exons and their flanking introns of PAH gene in these 15 PKU patients.Results PAH gene of 15 PKU patients was amplified by PCR, and PCR products were subjected to DNA sequencing directly.Four PAH gene mutation types, including 5′- Flanking-626G > A, 5′-Flanking-480DelACT, S196fsX4 and IVS8+1G>C, were identified in each of four PKU patients.Consequently reverse DNA sequencing showed G>A at -626 site, ACT deletion at -480 position in the promoter of PAH gene, an insertion at 584 site in the coding region and G>C at the border between exon 8 and intron 8 of PAH gene, respectively. After inquirying from PAH website and international PAH database (www.pahdb.mcgill.ca), these four PAH gene mutation types were verified as novel PAH gene mutations. Additionally, four patients carrying either of these four PAH gene mutation aged 3-5 years old were characterized by typical clinical phenotypes including blood Phe levels between 1 572-1 782 μmol/L, mental retardation, yellow hair and mousy odor of hair, skin and urine. Conclusions 5′-Flanking-626G>A, 5′-Flanking-480DelACT, S196fsX4 and IVS8+1G > C are identified as four novel PAH gene mutations to cause PKU directly probably either by disrupting the normal 3-D structure and affecting enzymatic activity of PAH or depressing the transcription and translation of PAH gene.Together, our identification of four novel PAH gene mutations will provide important clues for future gene diagnosis and prenatal diagnosis of PKU.

Clinical value of multiple tumor marker protein chip detective system in diagnosis and detection of post-operative recurrence of breast cancer / 国际肿瘤学杂志

Objective To evaluate the clinical value of multiple tumor marker protein chip in diagno-sis and detection of postoperative recurrence of breast cancer.Methods The serum levels of 12 tumor makers (CA199,NSE,CEA, CA2A2,Ferritin,β-HCG,AFP,f-PSA,PSA,CA125,CA153 and HGH)were measured in 70 preoperative breast cancer patients, 32 recurrence patients,52 non-recurrence patients and 76 normal con-trois.Results ①The breast cancer group had significantly higher positive rate than that of the controls (P<0.05).The positive rates and serum levels of CA199,CEA,CA242,Ferritin,CAI25 and CA153 in breast cancer patients had those of control significant differences compared with groups (P<0.05).②The recurrence group had significantly higher positive rate than that of non-recurrence group (P<0.05).The positive rates and se-rum levels of CA199, CEA, Ferritin, CA125 and CA153 in the recurrence patients had significant differences compared with those of non-recurrence patients(P<0.05).③The positive rate of recurrence group had signif-icant difference compared with that of breast cancer group(P<0.05).Moreover,The positive rate and serum level of Ferritin in the recurrence patients had significant difference compared with that of breast cancer pa-tients.Conclusion The multiple tumor marker protein chip detective system has valid value of clinical appli-cation in the diagnosis and detection of postoperative recurrence of breast cancer.The combination detection of CA199, CEA, Ferritin ,CA125 and CA153 may be the economical and effective in the diagnosis and detection of postoperative recurrence of breast cancer.

Identification of mutations in phenylalanine hydroxylase gene of Xinjiang minority nationality phenyiketonuria patients / 中华检验医学杂志

Objective To study the mutation characteristics in phenylalanine hydroxylase gene of Xinjiang minority nationality phenylketonuria (PKU) patients and provide a scientific basis for PKU prevention and cure strategy.Methods Mutations in phenylalanine hydroxylase gene were detected by Dolymerase chain reaction-single strand comformation polymorphism (PCR/SSCP) and gene sequencing in 12 minoritv nationality patients.Results Thirteen different mutations,including 8 missense mutations,1 nonsense mutation and 3 splice mutations were found in 24 alleles.The moat common mutations were EX696A＞G and P281 L.which were respectively prevalent in Asia and Europe populations.The common mutations were R243Q,R111X,R176X and F161S.The mutation frequency of R243Q was the highest and R111X was the third highest in Northern China.R176X and F161S were two rare mutations world wide.Especially.F161S was a Chinese-specific mutation because it was for the second time that it was found in China.The mutations detected in this study were first reported in these 3 minority nationality populations,which showed a distinct ethical characteristic.Condusions There is not only a consanguineous relation but also a distinct difference in PAH gene distribution between Xinjiang minority nationality population and yellow race and Latin-American.The results suggest that Xinjiang could probably be a special PAH gene distribution region.

R176X mutant homozygote in phenylketonuria firstly detected in Hui nationality in China: One case report / 中国组织工程研究

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.

Mutations in exon 5 of the phenylalanine hydroxylase (PAH) gene in patients with phenylketonuria / 解放军医学杂志

Objective To determine the mutations in exon 5 of the phenylalanine hydroxylase(PAH)gene in phenylketonuria(PKU)patients from Xinjiang.Methods The mutations in exon 5 and flanking sequence of PAH gene were detected by SSCP analysis and DNA sequencing.Results Four different mutations,including missence mutation F161S,splice mutation IVS4-1G→A,missence mutation R158Q and nonsence mutation Y166X were identified in 74 chromosomes from 37 PKU patients,with relative frequencies of 4.1%,1.4%,1.4% and 1.4%,respectively.The frequency of mutant alleles in exon 5 is 8.1%.Considering the previous reports and the present study,R158Q was the most prevalent form in PKU patients from European and Latin American countries,IVS4-1G→A was a common mutation inoriental PKU populations.However,F161S and Y166X are two characteristic forms in Chinese.Conclusion Characteristics of PAH gene mutations and their distribution were showed in Chinese PKU population from Xinjiang,where is a hinterland located between China and Europe.The results give a clue that Xinjiang might be an ideal genetic resource repertoire for studying diversity of gene mutations,heterogeneity of PAH gene,human genesis and migration.

Studies on mutations of exon 11 and 12 in phenylalaninase gene of phenylketonuria patients in Xinjiang / 解放军医学杂志

Objective To investigate the point mutation features of exon 11 and 12 in phenylalanine hydroxylase(PAH)gene of the patients with phenylketonuria(PKU)in Xinjiang.Methods PCR/SSCP and gene sequencing were used in present study.Results Five mutations were identified from 74 chromosomes of 37 patients.Among them two mutations were detected from exon 11 including nonsense mutation Y356X and splice site mutation V399V,and three mutations were detected from exon 12 including R413P,R408W and A434D,all which were missense mutation.The frequency of the five mutations were 5.4%,5.4%,4.1%,1.4%,1.4% and 1.4%,respectively.The allelomorphic frequency of exon 11 and 12 were 10.8% and 6.8%,respectively.Among the five mutations,R413P is common in Japan,Y356X and V399V are centered in north China,and R408W is the most often mutation in Europe and America.Conclusion Xinjiang Uygur Autonomous Region is situated on northwest China,contiguous to other countries of central Asia,and is such an area differed from other areas of China,and distributed special PAH gene mutation in PKU patients.