Animals ; Biopterin ; analogs & derivatives ; therapeutic use ; Genetic Therapy ; Humans ; Phenylalanine Ammonia-Lyase ; administration & dosage ; Phenylketonurias ; therapy

OBJECTIVETetrahydrobiopterin (BH(4)) responsive phenylalanine hydroxylase (PAH) deficiency is one of the forms of phenylketonuria (PKU). The aim of this study was to screen and diagnose BH(4) responsive PAH deficiency, to further understand its clinical characteristics, and to provide evidence for applying BH(4) drug therapy.

METHODSBH(4) 20 mg/kg loading test was performed in 73 patients with hyperphenylalaninemia (HPA) (47 males and 26 females), the mean age was 1.93 months. Combined phenylalanine (100 mg/kg) and BH(4) loading test was performed if patients had a basic blood phenylalanine concentration less than 600 micromol/L. The urine pterin profile analysis and the dihydropteridine reductase (DHPR) activity in dry blood filter spot were analyzed simultaneously. The patients with BH(4) responsive PAH deficiency were treated with BH(4) tablets (10 - 20 mg/kg x d) under normal diet for 6 to 7 days. Their blood phenylalanine concentration was checked.

RESULTS(1) The characteristic curve of phenylalanine level was observed in 73 patients after BH(4) loading test. Twenty-two patients were diagnosed as classic phenylketonuria (PKU), 39 were moderate PKU and 12 were BH(4) deficiency. (2) Twenty-two (56.4%) of 39 moderate PKU patients were found to be responsive to BH(4) and the blood phenylalanine was decreased by at least 30%. (3) Six patients with BH(4) responsive PAH deficiency were treated with BH(4) for 6 to 7 days, 4 patients had a normal phenylalanine concentration after 10 mg/kg BH(4) supplement, while other 2 patients needed a treatment of BH(4) at 20 mg/kg.

CONCLUSIONSome patients with moderate PKU caused by phenylalanine hydroxylase deficiency were responsive to BH(4). Their blood phenylalanine significantly decreased after oral BH(4) loading. The BH(4) loading test is an effective diagnostic method to detect BH(4) sensitivity in PKU patients. BH(4) responsive PAH deficiency patient could be treated with BH(4) to replace low-phenylalanine diet treatment totally or partially, which may provide an optional treatment for the disease and improve the quality of life of the patients.

Biopterin ; analogs & derivatives ; deficiency ; therapeutic use ; Child, Preschool ; Female ; Humans ; Infant ; Male ; Phenylalanine Hydroxylase ; deficiency ; Phenylketonurias ; diagnosis ; drug therapy

BACKGROUNDA novel therapeutic strategy for phenylketonuria (PKU) has been initiated in Japan. Hyperphenylalaninemia (HPA) results from a phenylalanine hydroxylase (PAH) enzyme deficiency or a deficiency of its cofactor, tetrahydrobiopterin (BH4). BH4 can normalize blood phenylalanine levels in BH4 deficiency, but typically not in PKU. However, since 1999 it has been reported that many HPA patients (serum phenylalanine <20 mg/dL) showed a gradual decrease of serum phenylalanine levels after 24 hours from BH4 loading. The BH4 responsiveness seems to be regulated in mild PKU by PAH mutations, and affected by the BH4 dose and administration period.

METHODS AND RESULTSIn 2002 we formulated a provisional diagnostic criteria for patients with BH4-responsive PAH deficiency, and newly diagnosed 19 patients in 100 HPA cases between 2002 and 2006. The incidence in the recent 5 years for BH4-responsive mild PKU among patients with PAH deficiency was 25 %.

CONCLUSIONA total of 31 patients was detected in the past 10 years, and the incidence detected using the provisional diagnostic criteria had increased to 25% among PAH deficient patients. BH4 treatment for BH4-responsive mild PKU is a new and effective pharmacotherapy, which replaces or liberalises the phenylalanine-restricted diets for a considerable number of mild PKU patients.

Biopterin ; analogs & derivatives ; therapeutic use ; Humans ; Infant, Newborn ; Japan ; Phenylketonurias ; diagnosis ; drug therapy ; Severity of Illness Index ; Time Factors

OBJECTIVETo investigate the development of differential diagnosis of tetrahydrobiopterin (BH4) deficiency among patients with hyperphenylalaninemia (HPA) in provinces or cities of China and to investigate the incidence of BH4 deficiency.

METHODSOf the thirteen hundreds and ninety-two patients with HPA received, the differential diagnosis for BH4 deficiency during 1993 - 2007 were enrolled in this study. Of which, 591 patients came from outpatient and 801 patients' samples from other provinces or cities were sent to author's laboratory to investigate the case number of differential diagnosis for BH4 deficiency in provinces or cities of China according to the data from both outpatient case histories and laboratory as to investigating the development of differential diagnosis in the whole country. To discuss the diagnostic criteria for BH4 deficiency was according to the results of urinary pterin analysis, determination of dihydropteridine reductase (DHPR) activity and the tetrahydrobiopterin loading test as well as to get the incidence of BH4 deficiency and find some provinces or cities with higher incidence of BH4 deficiency in China.

RESULTS(1) The number of HPA patients, who were performed by urinary pterin analysis and the determination of DHPR activity, were remarkably increased in last three years (2005 - 2007). The patient numbers of both urinary pterin analysis and DHPR activity determination were 217 and 198 respectively in 2005. And in 2007 they increased to 511 and 458, which was about 2.3 times than that in 2005. The patients came from 29 provinces or cities in 2007. (2) The urinary biopterin and biopterin percent were key marks for diagnosis of 6-pyruvoyl tetrahydropterin synthase (PTPS) deficiency. The less than 5% [(1.41 +/- 1.10)%] biopterin percent and very low biopterin level [(0.14 +/- 0.17) mmol/mol Cr] were found in 96.83% (61/63) patients with PTPS deficiency in this study. The blood phenylalanine level was remarkably decreased to normal range at 2 - 6 hours after BH4 loading test. The very low DHPR activity was a final diagnostic mark for DHPR deficiency. The very low DHPR activities of 0.27 nmol/(min x 5 mm disc) (6.11% - 7.00% of normal controls) were found in two patients with DHPR deficiency in this study. (3) The incidences of PTPS deficiency and DHPR deficiency among 1392 patients with hyperphenylalaninemia were 8.41% (117/1392) and 0.18% (2/1108) respectively. About 67.23% (80/119) patients with BH4 deficiency came from the south of Yangtze liver. The 80% (8/10) provinces or cities with higher incidence of BH4 deficiency are located in eastern and southern China. The incidence of PTPS deficiency among patients with HPA and normal newborns was 10.81% (8/74) and 0.007 per thousand (8/1,121,429) respectively in Shanghai, China according to data from neonatal screening.

CONCLUSIONThe awareness of differential diagnosis for BH4 deficiency from clinic pediatricians has been increased in most provinces or cities of China in last three years, but it should be more strengthened.

Biopterin ; analogs & derivatives ; deficiency ; China ; epidemiology ; Diagnosis, Differential ; Humans ; Incidence ; Infant, Newborn ; Neonatal Screening ; Phenylketonurias ; complications ; diagnosis ; epidemiology

OBJECTIVETo explore the effect of the level of lipid peroxidation and biomechanical properties after chronic treating with tetrahydrobiopterin (BH4) in thoracic aorta of hyperlipemia (HL) rats.

METHODSHL rats were given BH4 chronically. The opening angle in the zero-stress state and the relationship between pressure and diameter (P-D) of mesenteric artery were measured by computer image 8, 16, and 24 week-old respectively.

RESULTSTreating with BH4 chronically from 8 week-old in HL rats, there was a significant increase in the zero-stress state of opening angle of thoracic aorta. The P-D curve of mesenteric artery moved upward.

CONCLUSIONTreating with BH4 prevented the structure and function of artery from abnormal changing, and attenuated lipid peroxidation in HL rats.

Animals ; Aorta, Thoracic ; metabolism ; physiopathology ; Biomechanical Phenomena ; drug effects ; Biopterin ; analogs & derivatives ; therapeutic use ; Hyperlipidemias ; drug therapy ; Lipid Peroxidation ; drug effects ; Male ; Rats ; Rats, Wistar

OBJECTIVETo explore the oxidative modification effect and its mechanism of low density lipoprotein (LDL) in hyperlipidemia (HL) rats after treating with tetrahydrobiopterin (BH4).

METHODSFifty four 8-week-old male Wistar rats were used, these 54 rats were randomly divided into control group, high fat diet group (HL group), high fat diet and injected BH4 group (HL + BH4 group), and 18 in each group. The BH4 levels of blood fats and blood serum and its metabolites, the aortic reactive oxygen species, the end product malondialdehyde (MDA) and the LDL oxidation level were all determined by killing 6 experimental rats in each group at the first 8, 16, and 24 weeks of age respectively.

RESULTSTreating with BH4 after 8 and 16 weeks, there was no significant difference in serum lipids among three groups (P > 0. 05); but ROS and MDA decreased significantly (P < 0.01); compared with control and HL groups, the BH4 level of HL + BH4 group increased a lot (P < 0.01); compared with control group, the BH4 content reduced obviously in aortic homogenate of HL group (P < 0.01), but the total petrin levels (TB = BH4 + BH2 + B) had no significant difference (P > 0.05); the serum TBARS formation increased gradually with the increase of week-ages, but compared with HL group, the serum TBARS formation of HL + BH4 group reduced significantly (P < 0. 01).

CONCLUSIONTreating with BH4 can reduced the LDL oxidation, the mechanism may be related to the correct of NOS uncoupling, the reduce of ROS generation and the decrease of LDL lipid peroxidation.

Animals ; Biopterin ; analogs & derivatives ; therapeutic use ; Hyperlipidemias ; blood ; drug therapy ; Lipid Peroxidation ; Lipids ; blood ; Lipoproteins, LDL ; metabolism ; Male ; Malondialdehyde ; metabolism ; Rats ; Rats, Wistar ; Reactive Oxygen Species ; metabolism

AIMTo explore the effect of remodeling and biomechanical properties after chronic treating with tetrahydrobiopterin (BH4) in spontaneously hypertensive rats.

METHODSThe spontaneously hypertensive rat(SHR) were given with BH4 chronically. The opening angle in the zero-stress state , wall-to-lumen area ratios (W/L) of thoracic aorta and the relationship between pressure and diameter (P-D) of mesenteric artery were measured by computer image analysis in 4, 16, and 26 week-old respectively.

RESULTSTreating with BH4 chronically from 4 weeks-old in SHR, there was a significant decrease in morphometric parameters of the thoracic aorta and an increase in the zero-stress state of opening angle of elastic artery. The P-D curve of mesenteric artery moved upward.

CONCLUSIONTreating with BH4 prevented the structure and function of artery from abnormal changing, including to attenuate the resistant vascular hypertrophy and recover the vascular elasticity and expansibility.

Animals ; Arteries ; physiopathology ; Arteriolosclerosis ; prevention & control ; Biomechanical Phenomena ; Biopterin ; analogs & derivatives ; therapeutic use ; Hypertension ; drug therapy ; physiopathology ; Male ; Nitric Oxide Synthase ; Random Allocation ; Rats ; Rats, Inbred SHR

OBJECTIVETo analyze characteristics of different hyperphenylalaninemia (HPA) and to discuss the clinical difference between southern and northern Chinese patients with tetrahydrobiopterin (BH4) responsive phenylalanine hydroxylase (PAH) deficiency.

METHODS(1)BH4 (20 mg/kg) loading test was performed in all 108 HPA patients. These patients, 63 males and 45 females, were at a mean age of 7.05 months. A combined phenylalanine (Phe) and BH4 loading test was carried out in the patients who had a basic blood Phe concentration less than 600 micromol/L. The urine pterine profile analysis and the dihydropteridine reductase (DHPR) activity in dry blood filter spot were analyzed simultaneously. (2)BH4 responsive patients were divided to southern and northern groups by their parent's native place and geographic boundary determined by Changjiang River. The change of Phe concentration after BH4 loading test was compared between the two groups.

RESULTS(1)Among the 108 HPA cases, 36 patients (33.3%) were BH4 responsive PAH deficiency, 49 (45.4%) were non-BH4 no responsive phenylketonuria (PKU)and 23(21.3%)were BH4 deficiency (BH4D). The Phe concentration of patients with BH4 responsive PAH deficiency decreased by 49.24% and 65.35% at 8 h and 24 h after oral BH4, 23 in southern group and 13 in northern group among 36 patients. (2)The mean Phe concentration at 24 h after loading test in southern and northern groups were (217.02+/-189.03) micromol/L and 458.75+/-342.54 micromol/L respectively (P<0.05), although the decrease percent of plasma Phe concentration at 2 h, 4 h, 8 h, 24 h was no distinct difference between southern and northern groups (P>0.05).

CONCLUSIONMost of mild and moderate HPA patients affected by PAH deficiency show plasma Phe concentration decrease >30% in 24 h after oral BH4 20 mg/kg, few are classic PKU. The responsiveness to BH4 is no difference between southern and northern Chinese patients with BH4 responsive PAH deficiency according to the decrease percent of plasma Phe concentration, although the Phe concentration is lower in southern patients than that in northern patients.

Biopterin ; analogs & derivatives ; pharmacology ; therapeutic use ; Child, Preschool ; China ; Dihydropteridine Reductase ; blood ; Female ; Humans ; Infant ; Infant, Newborn ; Male ; Phenylalanine ; blood ; Phenylketonurias ; blood ; drug therapy ; Time Factors

PURPOSE: To investigate the effect of insulin on the production of nitric oxide (NO) in the trabecular meshwork (TM) cells and the enzymatic synthetic pathway of tetrahydrobiopterin (BH4) synthesis. METHODS: Primarily cultured human TM cells were exposed to 1, 10, and 100 microgram/ml of insulin and 0, 1, 10, 100 and 1000 nM dexamethasone for 3 days. To evaluate the enzymatic pathway of BH4 synthesis, 10 micrometer dexamethasone, 5 mM diaminopyrimidinone, 100 micrometer ascorbic acid, 100 micrometer sepiapterin, or 10 micrometer methotrexate were also co-administered respectively. Cellular survival and NO production were measured with MTT and Griess assay. RESULTS: Insulin enhanced NO production in a dose-dependent manner significantly (p<0.05) without affecting cell viability, whereas dexamethasone inhibited NO production. With co-exposure of insulin, diaminopyrimidinone and sepiapterin inhibited insulin-induced NO production. Ascorbic acid increased NO production independent of insulin and methotrexate did not affect to the action of insulin in NO production. CONCLUSIONS: Insulin increases NO production in TM cells via de novo synthetic pathway for BH4 synthesis. Insulin could be involved in the regulation of trabecular outflow by enhancing NO production in TM cells.
Trabecular Meshwork/cytology/*drug effects/*metabolism/physiology ; Nitric Oxide/*biosynthesis ; Insulin/administration & dosage/*pharmacology ; Humans ; Dose-Response Relationship, Drug ; Cells, Cultured ; Cell Survival/drug effects ; Biopterin/*analogs & derivatives/biosynthesis

OBJECTIVESTo assess the incidence of tetrahydrobiopterin (BH4) deficiency among patients with hyperphenylalaninemia (HPA) in southern Chinese and evaluate clinical outcome and gene mutations in tetrahydrobiopterin deficient patients.

METHODSUrinary neopterin (N) and biopterin (B) was analyzed in 87 patients with hyperphenylalaninemia by high-performance liquid chromatography. Further combined loading tests with phenylalanine (Phe) (100 mg/kg) and tetrahydrobiopterin (BH4) (7.5 mg/kg) were performed in suspected patients with abnormal urinary pterin profiles. Gene mutation analysis was performed for patients with BH4 deficiency and their parents. BH4 deficient patients were treated with BH4 and neurotransmitter precursors after diagnosis. Blood phenylalanine levels, clinical symptoms and mental development were followed up.

RESULTSEleven patients were diagnosed as having BH4 deficiency caused by 6-pyruvoyl tetrahydropterin synthase (PTPS) deficiency. The incidence of tetrahydrobiopterin (BH4) deficiency among patients with hyperphenylalaninemia (HPA) in southern Chinese was 10%. Combined loading tests with phenylalanine and oral BH4 were done in 4 of 11 patients and their phenylalanine levels were decreased to normal 4 - 6h after BH4 administration. Four different mutations (P87S, N52S, D96N and G144R) in the PTPS gene were detected in 5 families. Five PTPS-deficient patients were treated with synthetic BH4, neurotransmitter precursors (L-dopa plus carbidopa, and 5-hydroxytryptophan). They had satisfactory physical and mental development after treatment. One patient with partial PTPS deficiency had normal growth and mental development without treatment.

CONCLUSIONSOur results emphasize that screening for BH4 deficiency should be carried out in all patients with hyperphenylalaninemia in order to minimize the misdiagnosis. Patients with BH4 deficiency should be treated early with BH4 and a combination of neurotransmitter precursors.

Biopterin ; administration & dosage ; analogs & derivatives ; deficiency ; urine ; China ; DNA Mutational Analysis ; DNA, Complementary ; chemistry ; genetics ; Follow-Up Studies ; Genetic Testing ; Humans ; Mutation, Missense ; Neopterin ; urine ; Phenylketonurias ; blood ; enzymology ; genetics ; Phosphorus-Oxygen Lyases ; genetics ; metabolism