Search

Background/Objectives: As COVID-19 can be severe, early predictive markers of both severity and onset of secondary bacterial infections are needed. This study first examined changes over time in the levels of plasma neopterin (NP) and biopterins (BPs), among others, in patients with COVID-19 and then in those with secondary bacterial infection complications. Methods: Fifty-two patients with COVID-19 admitted to two tertiary care centers were included. They were divided into a severe group (intubated + mechanical ventilation) (n = 10) and a moderate group (non-intubated + oxygen administration) (n = 42), and changes over time in plasma NP, plasma BPs, IFN-γ, lymphocyte count, CRP, and IL-6 were investigated. Four of the patients in the severe group (n = 10) developed secondary bacterial infections during treatment. Plasma NP and plasma BPs of patients with bacterial sepsis (no viral infection) (n = 25) were also examined. Results: The plasma NP, IL-6, CRP, and SOFA levels were significantly higher in the severe group, while the IFN-γ level and lymphocyte count were significantly lower. The higher plasma NP in the severe group persisted only up to 1 week after symptom onset. The plasma BPs were higher in complications of bacterial infection. Conclusions: The timing of sample collection is important for assessing severity through plasma NP, while plasma BPs may be a useful diagnostic tool for identifying the development of secondary bacterial infection in patients with COVID-19. Further investigation is needed to clarify the mechanism by which NP and BPs, which are involved in the same biosynthetic pathway, are differentially activated depending on the type of pathogen. [ABSTRACT FROM AUTHOR]

Background and aims: Biopterins, including tetrahydrobiopterin (BH4), dihydrobiopterin (BH2), and biopterin (B), were crucial enzyme cofactors in vivo. Despite their recognized clinical significance, there remain notable research gaps and controversies surrounding experimental outcomes. This study aims to clarify the biopterins-related issues, including analytical art, physiological intervals, and pathophysiological implications. Materials and methods: A novel LC-MS/MS method was developed to comprehensively profile biopterins in plasma, utilizing chemical derivatization and cold-induced phase separation. Subsequently, apparently healthy individuals were enrolled to investigate the physiological ranges. And the relationships between biopterins and biochemical indicators were analyzed to explore the pathophysiological implications. Results: The developed method was validated as reliable for detecting biopterins across the entire physiological range. Timely anti-oxidation was found to be essential for accurate assessment of biopterins. The observed overall mean ± SDs levels were 3.51 ± 0.94, 1.54 ± 0.48, 2.45 ± 0.84 and 5.05 ± 1.14 ng/mL for BH4, BH2, BH4/BH2 and total biopterins. The status of biopterins showed interesting correlations with age, gender, hyperuricemia and overweight. Conclusion: In conjunction with proper anti-oxidation, the newly developed method enables accurate determination of biopterins status in plasma. The observed physiological intervals and pathophysiological implications provide fundamental yet inspiring support for further clinical researches. [ABSTRACT FROM AUTHOR]

Naho Yamamoto,1 Saki Kasuga,1 Daijiro Kabata,2 Myu Ono,1 Sakura Ando,1 Taisuke Hashimoto,1 Shiori Fujikawa,1 Hiroko Fujitani,3 Ayumi Shintani,2 Takashi Hamazaki1 1Department of Pediatrics, Osaka Metropolitan University Graduate School of Medicine, Osaka, Japan; 2Department of Medical Statistics, Osaka Metropolitan University Graduate School of Medicine, Osaka, Japan; 3Fujitani Clinic, Osaka, JapanCorrespondence: Naho Yamamoto, Department of Pediatrics, Osaka Metropolitan University Graduate School of Medicine, Osaka, Japan, Tel +81-6-6645-3816, Fax +81-6-6636-8737, Email naho_yama@omu.ac.jpPurpose: Pteridines are metabolites of tetrahydrobiopterin (BH4), being coenzymes for nitric oxide synthase (NOS). No study has clarified the relationship among pteridines and NOS, fractional exhaled nitric oxide (FeNO) generated by pteridines, and reactive oxygen species. In this study, we administered arginine, a precursor of NO, and confirmed changes in the levels of pteridines, FeNO, and reactive oxygen species and their relationship to clarify the pathogenesis of airway inflammation in which oxidative stress is involved, such as bronchial asthma.Patients and Methods: This is a prospective, randomized open-label study. Children, aged 2 to 15 years, who were scheduled for growth hormone stimulation tests and were able to undergo a respiratory function test were recruited. They were randomly divided into two groups: arginine-administered and control groups. In the former, L-arginine hydrochloride was intravenously administered. After administration, the levels of diacron-reactive oxygen metabolites (d-ROMs), serum pteridines, serum amino acids, and fractional exhaled NO (FeNO) were measured.Results: We analyzed 15 children aged 4 to 14 years. In the arginine-administered group, there was an increase in the FeNO level and a decrease in the d-ROMs level, reaching a peak 30 min after administration, compared with the control group. In addition, there was a decrease in the serum biopterin level and an increase in the d-ROMs level, reaching peak 60 min after administration.Conclusion: The administration of L-arginine increased the NO level and decreased the d-ROMs level. Due to this, biopterin may be consumed and decreased, leading to an increase in the d-ROMs level. As a reduction in reactive oxygen species leads to the relief of inflammation, arginine and biopterin may be useful for inhibiting inflammation.Keywords: asthma, biomarker, biopterin, child, FeNO, d-ROMs

Purpose: Pteridines are metabolites of tetrahydrobiopterin (BH4), being coenzymes for nitric oxide synthase (NOS). No study has clarified the relationship among pteridines and NOS, fractional exhaled nitric oxide (FeNO) generated by pteridines, and reactive oxygen species. In this study, we administered arginine, a precursor of NO, and confirmed changes in the levels of pteridines, FeNO, and reactive oxygen species and their relationship to clarify the pathogenesis of airway inflammation in which oxidative stress is involved, such as bronchial asthma. Patients and Methods: This is a prospective, randomized open-label study. Children, aged 2 to 15 years, who were scheduled for growth hormone stimulation tests and were able to undergo a respiratory function test were recruited. They were randomly divided into two groups: arginine-administered and control groups. In the former, L-arginine hydrochloride was intravenously administered. After administration, the levels of diacron-reactive oxygen metabolites (d-ROMs), serum pteridines, serum amino acids, and fractional exhaled NO (FeNO) were measured. Results: We analyzed 15 children aged 4 to 14 years. In the arginine-administered group, there was an increase in the FeNO level and a decrease in the d-ROMs level, reaching a peak 30 min after administration, compared with the control group. In addition, there was a decrease in the serum biopterin level and an increase in the d-ROMs level, reaching peak 60 min after administration. Conclusion: The administration of L-arginine increased the NO level and decreased the d-ROMs level. Due to this, biopterin may be consumed and decreased, leading to an increase in the d-ROMs level. As a reduction in reactive oxygen species leads to the relief of inflammation, arginine and biopterin may be useful for inhibiting inflammation. [ABSTRACT FROM AUTHOR]

Depressive disorder is a severe mental condition. In addition to genetic factors, immunological-inflammatory factors, oxidative stress, and disturbances in neurotransmitter metabolism, kynurenine and serotonin pathways may play a role. The exact mechanisms, especially in depressed children and adolescents, are not fully understood. Our primary hypothesis was whether the metabolites of tryptophan degradation in children and adolescentswith depressive disordermight be influenced by omega-3 FAs compared to omega-6 FAs during a 12-week supplementation. A secondary hypothesis was to investigate whether tryptophan metabolites in children and adolescents are associated with markers of inflammatory response, oxidative stress, cortisol, and the serum omega-6/ omega-3 FA ratio. Metabolites of tryptophan degradation and pteridines, neopterin, and biopterin in urine were analyzed with an HPLC system. Surprisingly, omega-3 FAs stimulated both kynurenine (kynurenine/tryptophan ratio) and serotonin (5-hydroxytryptophan) pathways, whereas omega-6 FAs only increased the kynurenine/tryptophan ratio. Neopterin and biopterin were not different from the healthy controls. Biopterin increased after omega-3 FA supplementation. Serotonin was positively correlated with lipoperoxidation and a marker of oxidative protein damage. Of the monitored tryptophan metabolites, only 5-hydroxyindolacetic acid was positively correlated with the severity of depression, total cholesterol, and negatively with brain-derived neurotrophic factor and glutathione peroxidase. In conclusion, in children and adolescents, both supplemented FAs stimulated the kynurenine pathway (kynurenine/ tryptophan ratio) and kynurenine formation. However, the serotonin pathway (5-hydroxytryptophan) was stimulated only by omega-3 FA. Tryptophan metabolism is associated with oxidative stress, inflammation, total cholesterol, and cortisol.We are the first to point out the association between the kynurenine pathway (KYN/ TRP ratio) and the omega-6/omega-3 FA ratio. Themetabolite 5-HIAA could play a role in the pathophysiology of depressive disorder in children and adolescents. [ABSTRACT FROM AUTHOR]

We have previously shown that circulating levels of tetrahydrobiopterin (H4B) function as a robust biomarker for aortic aneurysms in several independent animal models. In the present study, we examined diagnostic and predictive values of circulating H4B levels in human patients of thoracic aortic aneurysm (TAA) and abdominal aortic aneurysm (AAA) for the first time, while clinically applicable biomarkers for aortic aneurysms have never been previously available. Ninety-five patients scheduled for TAA repair surgeries and 53 control subjects were recruited at University of California Los Angeles (UCLA) Ronald Regan Medical Center, while 44 control subjects and 29 AAA patients were recruited through National Institute of Health (NIH) National Disease Research Interchange (NDRI) program. We had intriguing observations that circulating H4B levels were substantially lower in TAA and AAA patients, linearly correlated with aortic H4B levels (blood: R = 0.8071, p

Abstract Tetrahydrobiopterin deficiency in newborns with atypical hyperphenylalaninemia requires rapid and accurate diagnosis and the ability to distinguish it from the classical type to prevent early irreversible neurological damage. The study aimed to evaluate neopterin and biopterin (products of tetrahydrobiopterin recycling pathway) and amino acid profiles (used in supplementation therapy) in patients with hyperphenylalaninemia after optimizing ultra-performance liquid chromatography coupled with tandem mass spectrometry to simultaneously measure neopterin, biopterin, and amino acids in dried blood spots. The study enrolled preselected infants with classic (n = 46), atypical (n = 14) hyperphenylalaninemia, and a control group (n = 50). Result Tandem mass spectrometry detected neo/biopterin in the blood with a sensitivity and specificity of 100%. The mean neo/biopterin levels were significantly lower in the atypical cases (4 ± 1 and 3 ± 1 nmol/L) than the classic (49 ± 13 and 50 ± 12 nmol/L) and control (15.2 and 15.3 nmol/L) groups and correlated with phenylalanine and phenylalanine to tyrosine ratio (all P < 0.05). The study compared classic and atypical hyperphenylalaninemia cases with the control group. Both classic and atypical cases exhibited decreased levels of arginine, valine, and leucine compared to controls. Classic cases showed increased levels of citrulline, ornithine, and methionine, while atypical cases showed increased citrulline levels only. Comparing atypical versus classic cases, atypical cases exhibited decreased levels of citrulline, ornithine, methionine, arginine, leucine, and valine (all P < 0.05). Correlation analysis revealed negative associations between ornithine and biopterin and between arginine and neopterin in classic PKU cases. These findings highlight distinct metabolic differences between classic and atypical PKU. Conclusion The optimized method detected neo/biopterin in the blood with accuracy and precision. The characteristic pattern of neo/biopterin in the blood makes it possible to differentiate between classic and atypical hyperphenylalaninemia with a sensitivity and specificity of 100%. The amino acid profile could add value when treatment with large neutral amino acids is considered.

Tetrahydrobiopterin deficiency in newborns with atypical hyperphenylalaninemia requires rapid and accurate diagnosis and the ability to distinguish it from the classical type to prevent early irreversible neurological damage. The study aimed to evaluate neopterin and biopterin (products of tetrahydrobiopterin recycling pathway) and amino acid profiles (used in supplementation therapy) in patients with hyperphenylalaninemia after optimizing ultra-performance liquid chromatography coupled with tandem mass spectrometry to simultaneously measure neopterin, biopterin, and amino acids in dried blood spots. The study enrolled preselected infants with classic (n = 46), atypical (n = 14) hyperphenylalaninemia, and a control group (n = 50). Result Tandem mass spectrometry detected neo/biopterin in the blood with a sensitivity and specificity of 100%. The mean neo/biopterin levels were significantly lower in the atypical cases (4 ± 1 and 3 ± 1 nmol/L) than the classic (49 ± 13 and 50 ± 12 nmol/L) and control (15.2 and 15.3 nmol/L) groups and correlated with phenylalanine and phenylalanine to tyrosine ratio (all P < 0.05). The study compared classic and atypical hyperphenylalaninemia cases with the control group. Both classic and atypical cases exhibited decreased levels of arginine, valine, and leucine compared to controls. Classic cases showed increased levels of citrulline, ornithine, and methionine, while atypical cases showed increased citrulline levels only. Comparing atypical versus classic cases, atypical cases exhibited decreased levels of citrulline, ornithine, methionine, arginine, leucine, and valine (all P < 0.05). Correlation analysis revealed negative associations between ornithine and biopterin and between arginine and neopterin in classic PKU cases. These findings highlight distinct metabolic differences between classic and atypical PKU. Conclusion The optimized method detected neo/biopterin in the blood with accuracy and precision. The characteristic pattern of neo/biopterin in the blood makes it possible to differentiate between classic and atypical hyperphenylalaninemia with a sensitivity and specificity of 100%. The amino acid profile could add value when treatment with large neutral amino acids is considered. [ABSTRACT FROM AUTHOR]

Inflammation is thought to be involved in the pathogenesis of autism spectrum disorder (ASD). Pteridine metabolites are biomarkers of inflammation that increase on immune system activation. In this study, we investigated the urinary pteridine metabolites in ASD patients as a possible biomarker for immune activation and inflammation. This observational, cross-sectional, prospective study collected urine samples from 212 patients with ASD and 68 age- and sex-matched healthy individuals. Urine neopterin (NE) and biopterin (BIO) levels were measured. Patients who had chronic disorders, active infection at the time of sampling, or high C-reactive protein levels were excluded. The urine NE and BIO concentrations were determined by high-performance liquid chromatography. The ratios of both NE and BIO to creatinine (CRE) were used to standardise the measurements. The NE/CRE and NE/BIO levels were significantly higher in ASD patients than controls. Univariate and multivariate models revealed a significant increase in NE/CRE and NE/BIO in ASD patients. There was a significant relationship between the NE/BIO [average area under the curve (AUC) = 0.717; range: 0.637–0.797] and NE/CRE (average AUC = 0.756; range: 0.684–0.828) ratios, which distinguished individuals with ASD from controls. The elevated NE/CRE and NE/BIO ratios suggest that inflammation and T cell-mediated immunity are involved in the pathophysiology of autism. NE/BIO could serve as a diagnostic inflammatory marker in the pathogenesis of ASD. [ABSTRACT FROM AUTHOR]

Depressive disorder is a severe mental condition. In addition to genetic factors, immunological-inflammatory factors, oxidative stress, and disturbances in neurotransmitter metabolism, kynurenine and serotonin pathways may play a role. The exact mechanisms, especially in depressed children and adolescents, are not fully understood. Our primary hypothesis was whether the metabolites of tryptophan degradation in children and adolescents with depressive disorder might be influenced by omega-3 FAs compared to omega-6 FAs during a 12-week supplementation. A secondary hypothesis was to investigate whether tryptophan metabolites in children and adolescents are associated with markers of inflammatory response, oxidative stress, cortisol, and the serum omega-6/omega-3 FA ratio. Metabolites of tryptophan degradation and pteridines, neopterin, and biopterin in urine were analyzed with an HPLC system. Surprisingly, omega-3 FAs stimulated both kynurenine (kynurenine/tryptophan ratio) and serotonin (5-hydroxytryptophan) pathways, whereas omega-6 FAs only increased the kynurenine/tryptophan ratio. Neopterin and biopterin were not different from the healthy controls. Biopterin increased after omega-3 FA supplementation. Serotonin was positively correlated with lipoperoxidation and a marker of oxidative protein damage. Of the monitored tryptophan metabolites, only 5-hydroxyindolacetic acid was positively correlated with the severity of depression, total cholesterol, and negatively with brain-derived neurotrophic factor and glutathione peroxidase. In conclusion, in children and adolescents, both supplemented FAs stimulated the kynurenine pathway (kynurenine/tryptophan ratio) and kynurenine formation. However, the serotonin pathway (5-hydroxytryptophan) was stimulated only by omega-3 FA. Tryptophan metabolism is associated with oxidative stress, inflammation, total cholesterol, and cortisol. We are the first to point out the association between the kynurenine pathway (KYN/TRP ratio) and the omega-6/omega-3 FA ratio. The metabolite 5-HIAA could play a role in the pathophysiology of depressive disorder in children and adolescents.Clinical Trial Registrationhttps://www.isrctn.com/ISRCTN81655012, identifier ISRCTN81655012.

BackgroundCerebral malaria (CM) pathogenesis remains incompletely understood. Having shown low systemic levels of tetrahydrobiopterin (BH4), an enzymatic cofactor for neurotransmitter synthesis, we hypothesized that BH4 and BH4-dependent neurotransmitters would likewise be low in cerebrospinal fluid (CSF) in CM.MethodsWe prospectively enrolled Tanzanian children with CM and children with nonmalaria central nervous system conditions (NMCs). We measured CSF levels of BH4, neopterin, and BH4-dependent neurotransmitter metabolites, 3-O-methyldopa, homovanillic acid, and 5-hydroxyindoleacetate, and we derived age-adjusted z-scores using published reference ranges.ResultsCerebrospinal fluid BH4 was elevated in CM (n = 49) compared with NMC (n = 51) (z-score 0.75 vs -0.08; P

The core symptoms of many neurological disorders have traditionally been thought to be caused by genetic variants affecting brain development and function. However, the gut microbiome, another important source of variation, can also influence specific behaviors. Thus, it is critical to unravel the contributions of host genetic variation, the microbiome, and their interactions to complex behaviors. Unexpectedly, we discovered that different maladaptive behaviors are interdependently regulated by the microbiome and host genes in the Cntnap2-/- model for neurodevelopmental disorders. The hyperactivity phenotype of Cntnap2-/- mice is caused by host genetics, whereas the social-behavior phenotype is mediated by the gut microbiome. Interestingly, specific microbial intervention selectively rescued the social deficits in Cntnap2-/- mice through upregulation of metabolites in the tetrahydrobiopterin synthesis pathway. Our findings that behavioral abnormalities could have distinct origins (host genetic versus microbial) may change the way we think about neurological disorders and how to treat them.

Background Biogenic amines and pterins analysis in cerebrospinal fluid (CSF) are reliable biomarkers for the diagnosis of inherited disorders of monoamine neurotransmitters. Objective The objectives of this study were the establishment of reference values of CSF biogenic amine metabolites in a cohort of Greek children, the detection of primary defects of biogenic amine metabolism, and the assessment of biogenic amine metabolites in children with different neurological disorders. Methods CSF biogenic amine metabolites and pterins (biopterin and neopterin) were analyzed using high-performance liquid chromatography with electrochemical and fluorescence detection. Three hundred sixty-three samples were analyzed: 60 infants and children with no history of neurological disorder, 6 with inherited disorders of monoamine neurotransmitters, and 297 with diverse neurological disorders. Results Reference values were stratified into six age groups. A strong correlation between homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5HIAA) levels with age was detected (p < 0.001). Two patients were diagnosed with a defect of the biogenic amine synthetic pathway and three with a defect of tetrahydrobiopterin cofactor production. HVA and 5HIAA abnormalities were detected within different groups of neurological disorders, but none followed a specific pattern of HVA and 5HIAA abnormalities. Conclusion In the current study, Greek reference values of biogenic amines and pterins in CSF are presented. Five new patients with inherited monoamine neurotransmitter disorders are described. Nonspecific secondary biogenic amine disturbances can be seen in patients with different neurological disorders. [ABSTRACT FROM AUTHOR]

Endothelial dysfunction result from inflammation and excessive production of reactive oxygen species as part of the surgical stress response. Remote ischemic preconditioning (RIPC) potentially exerts anti-oxidative and anti-inflammatory properties, which might stabilise the endothelial function after non-cardiac surgery. This was a single centre randomised clinical trial including 60 patients undergoing sub-acute laparoscopic cholecystectomy due to acute cholecystitis. Patients were randomised to RIPC or control. The RIPC procedure consisted of four cycles of five minutes of ischaemia and reperfusion of one upper extremity. Endothelial function was assessed as the reactive hyperaemia index (RHI) and circulating biomarkers of nitric oxide (NO) bioavailability (L-arginine, asymmetric dimethylarginine (ADMA), L-arginine/ADMA ratio, tetra- and dihydrobiopterin (BH4 and BH2), and total plasma biopterin) preoperative, 2–4 h after surgery and 24 h after surgery. RHI did not differ between the groups (p = 0.07). Neither did levels of circulating biomarkers of NO bioavailability change in response to RIPC. L-arginine and L-arginine/ADMA ratio was suppressed preoperatively and increased 24 h after surgery (p < 0.001). The BH4/BH2-ratio had a high preoperative level, decreased 2–4 h after surgery and remained low 24 h after surgery (p = 0.01). RIPC did not influence endothelial function or markers of NO bioavailability until 24 h after sub-acute laparoscopic cholecystectomy. In response to surgery, markers of NO bioavailability increased, and oxidative stress decreased. These findings support that a minimally invasive removal of the inflamed gallbladder countereffects reduced markers of NO bioavailability and increased oxidative stress caused by acute cholecystitis. [ABSTRACT FROM AUTHOR]

Butyrate, a short-chain fatty acid, is utilized by the gut epithelium as energy and it improves the gut epithelial barrier. More recently, it has been associated with beneficial effects on immune and cardiovascular homeostasis. Conversely, tumor necrosis factor alpha (TNFα) is a pro-inflammatory and pro-hypertensive cytokine. While butyrate and TNFα are both linked with hypertension, studies have not yet addressed their interaction in the colon. Here, we investigated the capacity of butyrate to modulate a host of effects of TNFα in primary rodent colonic cells in vitro. We measured ATP levels, cell viability, mitochondrial membrane potential (MMP), reactive oxygen species (ROS), mitochondrial oxidative phosphorylation, and glycolytic activity in colonocytes following exposure to either butyrate or TNFα, or both. To address the potential mechanisms, transcripts related to oxidative stress, cell fate, and cell metabolism (Pdk1, Pdk2, Pdk4, Spr, Slc16a1, Slc16a3, Ppargc1a, Cs, Lgr5, Casp3, Tnfr2, Bax, Bcl2, Sod1, Sod2, and Cat) were measured, and untargeted liquid chromatography–tandem mass spectrometry (LC-MS/MS) was employed to profile the metabolic responses of colonocytes following exposure to butyrate and TNFα. We found that both butyrate and TNFα lowered cellular ATP levels towards a quiescent cell energy phenotype, characterized by decreased oxygen consumption and extracellular acidification. Co-treatment with butyrate ameliorated TNFα-induced cytotoxicity and the reduction in cell viability. Butyrate also opposed the TNFα-mediated decrease in MMP and mitochondrial-to-intracellular calcium ratios, suggesting that butyrate may protect colonocytes against TNFα-induced cytotoxicity by decreasing mitochondrial calcium flux. The relative expression levels of pyruvate dehydrogenase kinase 4 (Pdk4) were increased via co-treatment of butyrate and TNFα, suggesting the synergistic inhibition of glycolysis. TNFα alone reduced the expression of monocarboxylate transporters slc16a1 and slc16a3, suggesting effects of TNFα on butyrate uptake into colonocytes. Of the 185 metabolites that were detected with LC-MS, the TNFα-induced increase in biopterin produced the only significant change, suggesting an alteration in mitochondrial biogenesis in colonocytes. Considering the reports of elevated colonic TNFα and reduced butyrate metabolism in many conditions, including in hypertension, the present work sheds light on cellular interactions between TNFα and butyrate in colonocytes that may be important in understanding conditions of the colon. [ABSTRACT FROM AUTHOR]

Background: Tetrahydrobiopterin is a coenzyme of phenylalanine hydroxylase and other enzymes essential for the synthesis of tyrosine and other neurotransmitters. Despite proper nutritional regulation of blood phenylalanine levels, tetrahydrobiopterin deficiency results in progressive neurologic illness. Objective: The study aims to optimize a mass spec-dependent assay for quantitative measurement of biopterin and neopterin as distinctive markers of the atypical phenylketonuria due to tetrahydrobiopterin deficiency. Patients and methods: The study enrolled 46 patients with typical hyperphenylalaninaemia, 14 atypical cases, and 50 healthy children as a control group. Quantitative measurements of biopterin, neopterin, phenylalanine, and tyrosine were performed by ultra-performance liquid chromatography-mass spectrometry in the dried blood spots of patients and control samples. Results: Validation of the analytical protocol was performed at a biopterin and neopterin concentration range from 0 to 100 nmol/l. The regression coefficients for the linearity of the calibration curves exceeded 0.98. The lower limit of detection of biopterin and neopterin ranged from 1.5 to 2.5 nmol/l. The intra-day and inter-day precision and accuracy ranged from 96% - 105%, and from 97% - 110%. The mean recoveries were 105 ± 7% for biopterin and 106 ± 9% for neopterin. The short and long-term stability of the stored samples was seven days at room temperature and 12 weeks at -20 °C. Biopterin and neopterin were significantly higher in the classic group compared to the atypical and control group (p < 0.05), with their level being in the order classic > control > atypical group. Phenylalanine levels had a significant positive correlation with biopterin and neopterin levels in classic phenylketonuria (p < 0.05). Conclusions: The performance of the developed assay for biopterin and neopterin in the dried blood spot by ultraperformance liquid chromatography-mass spectrometry was accurate and precise thus, provide a legitimate diagnostic tool for cases of atypical phenylketonuria. [ABSTRACT FROM AUTHOR]

The aim of the study was to determine whether biopterin is present in significantly lower quantities in urine samples of patients with autism spectrum disorder (ASD) compared to healthy individuals.

[Figure: see text]. [ABSTRACT FROM AUTHOR]

Neopterin (NP), biopterin (BP) and monapterin (MP) exist in saliva. The physiological role of salivary NP as well as the pathophysiological role of increased NP in the immune-activated state has been unclear. Saliva is a characteristic specimen different from other body fluids. In this study, we analysed salivary NP and related pterin compounds, BP and MP and revealed some of its feature. High-performance liquid chromatography (HPLC) analysis of saliva and plasma obtained from 26 volunteers revealed that salivary NP existed mostly in its fully oxidized form. The results suggested that salivary NP as well as BP would mostly originate from the oral cavity, perhaps the salivary glands, and that salivary NP levels might not reflect those in the plasma. We also found that a gender difference existed in correlations between concentrations of salivary total concentrations of NP (tNP) and BP (tBP). HPLC analysis of saliva obtained from 5 volunteers revealed that the concentrations of salivary tNP as well as tBP fluctuated in an irregular fashion in various individuals. MP, a diastereomer of NP, might have come from oral cavity NP itself or its precursor. These results indicated that the nature of salivary NP might be different from that of NP in the blood or urine. [ABSTRACT FROM AUTHOR]

Tetrahydrobiopterin (BH4) is the natural cofactor of several enzymes widely distributed among eukaryotes, including aromatic amino acid hydroxylases (AAAHs), nitric oxide synthases (NOSs), and alkylglycerol monooxygenase (AGMO). We show here that the nematode Caenorhabditis elegans, which has three AAAH genes and one AGMO gene, contains BH4 and has genes that function in BH4 synthesis and regeneration. Knockout mutants for putative BH4 synthetic enzyme genes lack the predicted enzymatic activities, synthesize no BH4, and have indistinguishable behavioral and neurotransmitter phenotypes, including serotonin and dopamine deficiency. The BH4 regeneration enzymes are not required for steady-state levels of biogenic amines, but become rate limiting in conditions of reduced BH4 synthesis. BH4-deficient mutants also have a fragile cuticle and are generally hypersensitive to exogenous agents, a phenotype that is not due to AAAH deficiency, but rather to dysfunction in the lipid metabolic enzyme AGMO, which is expressed in the epidermis. Loss of AGMO or BH4 synthesis also specifically alters the sensitivity of C. elegans to bacterial pathogens, revealing a cuticular function for AGMO-dependent lipid metabolism in host-pathogen interactions.

CRISPR-Cas transcriptional programming in bacteria is an emerging tool to regulate gene expression for metabolic pathway engineering. Here we implement CRISPR-Cas transcriptional activation (CRISPRa) in P. putida using a system previously developed in E. coli. We provide a methodology to transfer CRISPRa to a new host by first optimizing expression levels for the CRISPRa system components, and then applying rules for effective CRISPRa based on a systematic characterization of promoter features. Using this optimized system, we regulate biosynthesis in the biopterin and mevalonate pathways. We demonstrate that multiple genes can be activated simultaneously by targeting multiple promoters or by targeting a single promoter in a multi-gene operon. This work will enable new metabolic engineering strategies in P. putida and pave the way for CRISPR-Cas transcriptional programming in other bacterial species. [Display omitted] • Bacterial CRISPR activation (CRISPRa) was successfully ported to P. putida. • Multi-gene programs with simultaneous CRISPRa and CRISPRi can be implemented. • Systematic analysis shows that rules for effective E. coli CRISPRa apply in P. putida. • CRISPRa can activate endogenous promoters. • Inducible CRISPRa can be obtained by expressing dCas9 from inducible promoters. [ABSTRACT FROM AUTHOR]

This article explains the mechanisms underlying choices of pharmacotherapy for hypoxic-ischemic insults of both preterm and term babies. Some preclinical data are strong enough that clinical trials are now underway. Challenges remain in deciding the best combination therapies for each age and insult.

Purpose: Pteridines are metabolites of tetrahydrobiopterin (BH 4 ), being coenzymes for nitric oxide synthase (NOS). No study has clarified the relationship among pteridines and NOS, fractional exhaled nitric oxide (FeNO) generated by pteridines, and reactive oxygen species. In this study, we administered arginine, a precursor of NO, and confirmed changes in the levels of pteridines, FeNO, and reactive oxygen species and their relationship to clarify the pathogenesis of airway inflammation in which oxidative stress is involved, such as bronchial asthma., Patients and Methods: This is a prospective, randomized open-label study. Children, aged 2 to 15 years, who were scheduled for growth hormone stimulation tests and were able to undergo a respiratory function test were recruited. They were randomly divided into two groups: arginine-administered and control groups. In the former, L-arginine hydrochloride was intravenously administered. After administration, the levels of diacron-reactive oxygen metabolites (d-ROMs), serum pteridines, serum amino acids, and fractional exhaled NO (FeNO) were measured., Results: We analyzed 15 children aged 4 to 14 years. In the arginine-administered group, there was an increase in the FeNO level and a decrease in the d-ROMs level, reaching a peak 30 min after administration, compared with the control group. In addition, there was a decrease in the serum biopterin level and an increase in the d-ROMs level, reaching peak 60 min after administration., Conclusion: The administration of L-arginine increased the NO level and decreased the d-ROMs level. Due to this, biopterin may be consumed and decreased, leading to an increase in the d-ROMs level. As a reduction in reactive oxygen species leads to the relief of inflammation, arginine and biopterin may be useful for inhibiting inflammation., Competing Interests: Prof. Dr. Ayumi Shintani reports grants and/or personal fees from Kyowa Kirin, AstraZeneca K.K., Asahi Kasei Pharma Corporation, Chugai Pharmaceutical Co, Daiichi Sankyo Co, Merck Biopharma, Pfizer Japan Inc, Takeda Pharmaceutical Co, Shionogi Pharma, and Janssen Pharmaceutical K.K, outside the submitted work. The authors report no other conflicts of interest in this work., (© 2024 Yamamoto et al.)

Alkylglycerol mono-oxygenase (EC 1.14.16.5) forms a third, distinct, class among tetrahydrobiopterin-dependent enzymes in addition to aromatic amino acid hydroxylases and nitric oxide synthases. Its protein sequence contains the fatty acid hydroxylase motif, a signature indicative of a di-iron centre, which contains eight conserved histidine residues. Membrane enzymes containing this motif, including alkylglycerol mono-oxygenase, are especially labile and so far have not been purified to homogeneity in active form. To obtain a first insight into structure-function relationships of this enzyme, we performed site-directed mutagenesis of 26 selected amino acid residues and expressed wild-type and mutant proteins containing a C-terminal Myc tag together with fatty aldehyde dehydrogenase in Chinese-hamster ovary cells. Among all of the acidic residues within the eight-histidine motif, only mutation of Glu137 to alanine led to an 18-fold increase in the Michaelis-Menten constant for tetrahydrobiopterin, suggesting a role in tetrahydrobiopterin interaction. A ninth additional histidine residue essential for activity was also identified. Nine membrane domains were predicted by four programs: ESKW, TMHMM, MEMSAT and Phobius. Prediction of a part of the structure using the Rosetta membrane ab initio method led to a plausible suggestion for a structure of the catalytic site of alkylglycerol mono-oxygenase.

In the present study, the possible activation of cellular immunity in SCD patients was investigated. As immune activation parameters, neopterin concentrations and kynurenine/tryptophan ratio for tryptophan degradation in 35 pediatric patients with sickle cell disease (31 HbSS and 4 HbSß) were determined. Our results have shown that neopterin levels (both urinary and serum) are increased in pediatric patients with sickle cell disease. The increase in neopterin concentration was accompanied by significantly increased biopterin, kynurenine concentration and kynurenine/tryptophan ratio. The mechanism of immune activation and the effects of inflammatory mediators in sickle cell disease are poorly understood, especially in terms of cell-mediated immunity. Further in-vivo and in-vitro studies are required to illuminate the association between neopterin levels and neutrophil activation in sickle cell disease. [ABSTRACT FROM AUTHOR]

Conjugated and unconjugated pteridines and their derivatives are important cofactors in cellular metabolism. Hence, the amount of unconjugated pteridines in biological fluids has been found to be modified as a result of several disorders. It is necessary to note that while for the control of pteridines in urine samples there are numerous reference data, the literature referred to for the analysis of these analytes in serum/plasma is scarce. In biological fluids, pteridines can exist in different oxidation states, and these compounds can be classified into two groups according to: (a) oxidized or aromatic pteridines and (b) reduced pteridines. Oxidized pteridines yield a strong fluorescence signal, whereas reduced pteridines present a low quantum yield of fluorescence. In order to enable the analysis of the reduced forms, several preoxidation procedures to generate aromatic rings have been established. Also, stabilization of the reduced forms by the addition of reducing agents has been widely reported. The objective of this paper is to show possibilities and different approaches in the analysis of pteridines in serum samples. We have mainly focused on the description of the current situation in the application of high-performance chromatography methods with fluorimetric detection.

The analysis of pterins in the cerebrospinal fluid (CSF) is mandatory for the etiologic diagnosis of inborn errors of dopamine and serotonin metabolism. The success of the available therapeutic strategies for preventing the ongoing brain dysfunction is tightly dependent of the early diagnosis of these neurotransmitter disorders. Previous methods of pterins determination in the CSF have in common at least one reversed phase HPLC step coupled to electrochemical or fluorescence detection (FD). They differ in the oxidation procedure of the reduced forms of pterins into their oxidized fluorescent counterparts. Most of the methods using the FD include at least one offline chemical oxidation procedure and cannot allow the direct quantification of tetrahydrobiopterin (BH4). A recent method proposed a single step simultaneous quantification of all forms of pterins including BH4 by HPLC coupled to FD after post-column coulometric oxidation. Nowadays, recent advances in mass spectrometry (MS), notably in term of sensitivity, allow the direct unambiguous determination of all forms of pterins in the CSF by LC-MS/MS.

Urinary biopterin (Bio) and neopterin (Neo) are important markers for clinical diagnosis of hyperphenylalaninemia. Herein, we developed a high‐throughput analysis method based on electrospray ionization mass spectrometry (ESI‐MS) with polymer tips for the rapid quantitative detection of Bio and Neo in clinical urine samples. Different polymer tips were investigated. It is found that the best detection sensitivity was achieved with hydrophobic polymer tip, ie, polyethylene tips. The high‐throughput polymer tip‐ESI‐MS method allowed a rapid analysis speed at ~40 seconds per sample. The limits of quantification (LOQ) (S/N ≥ 10) for the detection of Bio and Neo were improved to be 5.0 ng/mL. Acceptable relative standard deviation (RSD) values for Neo and Bio were measured to be 12.2% and 13.4% for direct measurement of Bio and Neo in raw urine samples, respectively. Furthermore, Bio and Neo were directly quantified from 18 clinical urine samples by presented method. The ratios of urinary Bio‐to‐Neo were analyzed for diagnosis of hyperphenylalaninemia. The results demonstrated that the present polymer tip‐ESI‐MS method is a promising strategy for the rapid analysis of clinical samples. [ABSTRACT FROM AUTHOR]

Abstract Background Pteridines are metabolites of tetrahydrobiopterin, which serves as co-enzyme of nitric oxide synthase. We sought to investigate the usefulness of pteridines as biomarkers for childhood asthma control. Methods We conducted a single-center prospective cohort study involving 168 asthmatic children aged 4–17 years who visited the periodical asthma checkup program. Serum neopterin and biopterin levels were measured as pteridines at each visit along with measurement of FeNO, respiratory function tests, nasal eosinophil test, blood eosinophil count, and IgE level. We calculated coefficients for relation between pteridines and asthma control, which was assessed by questionnaires (JPAC: Japanese Pediatric Asthma Control Program). Results A total of 168 participants aged 10.3 ± 3.39 years (mean ± SD) with asthma were recruited. The participants in this study contained 58 patients (34.5%) of complete-controlled based on JPAC, 132 patients (76.0%) of well-controlled group based on GINA. FeNO and serum neopterin level did not correlate with following period's JPAC scores. In contrast, serum biopterin level significantly correlated with following period's JPAC total score (Coefficients 0.398; 95% CI 0.164 to 0.632; p value 0.001) and frequency of wheezing during exercise (Coefficients 0.272; 95% CI 0.217 to 0.328; p value < 0.001). Conclusions We found serum biopterin effected the following period's control status of asthmatic children, thus monitoring biopterin level will be a useful for management of asthma to adjust treatment. Graphical abstract Image 1 [ABSTRACT FROM AUTHOR]

Cigarette smoking (CS) is a well-established risk factor for cardiovascular disease (CVD). Endothelial dysfunction (ED) with loss of nitric oxide (NO) production is a central mechanism leading to the advent of CVD. Despite many prior studies of this major health problem, the exact mechanism by which CS induces ED is not well understood. This study examines the mechanism by which CS induces ED with altered endothelial NO synthase (eNOS) function in aortic endothelial cells (AECs). Exposure of AECs to cigarette smoke extract (CSE) resulted in a marked decrease in NO production with concomitant increase in superoxide (O 2 .- ) generation and accumulation of 4-hydroxy-2-nonenal protein adducts. CSE exposure led to depletion of the essential eNOS cofactor tetrahydrobiopterin (BH 4 ) as well as total biopterin levels and decreased the expression level of guanosine triphosphate cyclohydrolase (GTPCH), the rate limiting enzyme in BH 4 biosynthesis. Moreover, exposure of AECs to CSE increased the level of ubiquitinated proteins and increased 26 S proteasomal activity in a concentration-dependent manner. Pre-treatment with MG132, a 26 S proteasome inhibitor, partially prevented CSE-induced loss of BH 4, total biopterin, GTPCH, and increased NO production following CSE exposure, indicating a role of the ubiquitin-proteasome system in CSE-induced eNOS dysfunction. In conclusion, CSE-induced eNOS dysfunction and uncoupling occurs due to BH 4 depletion with BH 4 de novo synthesis limited by diminished GTPCH expression. [ABSTRACT FROM AUTHOR]

Objective: Tetrahydrobiopterin (BH4) is the essential cofactor of endothelial nitric oxide synthase (eNOS) and intracellular levels of BH4 is regulated by oxidative stress. The aim of this paper was to describe the influence of exogenous endothelin-1 on intracellular BH4 and its oxidation products dihydrobiopterin (BH2) and biopterin (B) in a wide range of vascular tissue. Design: Segments of internal mammary artery (IMA) and human saphenous vein (SV) from 41 patients undergoing elective surgery were incubated in ET-1 (0.1 μM). Aorta and lung from transgenic mice overexpressing ET-1 in the endothelium (ET-TG) were analysed with regards to intracellular biopterin levels. Human umbilical vein endothelial cells (HUVEC) were incubated in ET-1 (0.1 μM) and intracellular biopterin levels were analysed. From 6 healthy women undergoing caesarean section, subcutaneous fat was harvested and the resistance arteries in these biopsies were tested for ET-mediated endothelial dysfunction. Results: In HUVEC, exogenous ET-1 (0.1 μM) did not significantly change intracellular BH4, 1.54 ± 1.7 vs 1.68 ± 1.8 pmol/mg protein; p = .8. In IMA and SV, exogenous ET-1(0.1 μM) did not change intracellular BH4 n = 10, p = .4. In aorta from wild type vs ET-TG mice there was no significant difference in intracellular BH4 between the groups: 1.3 ± 0.49 vs 1.23 ± 0.3 pmol/mg protein; p = .6. In resistance arteries (n = 6) BH4 together with DTE (an antioxidant) was not able to prevent ET-mediated endothelial dysfunction. Conclusion: ET-1 did not significantly alter intracellular tetrahydrobiopterin levels in IMA, SV, HUVEC or aorta from ET-TG mice. These findings are important for future research in ET-1 mediated superoxide production and endothelial dysfunction. [ABSTRACT FROM AUTHOR]

Tetrahydrobiopterin (BH 4 ) is an essential cofactor for dopamine, serotonin and nitric oxide synthesis. Deficits of plasma total biopterin (a measure of BH 4 ) have been described in schizophrenia and schizoaffective disorder. GCH1 encodes the first and rate-limiting enzyme in BH 4 synthesis. Peripheral GCH1 expression is lower in first episode psychosis patients versus controls, and we hypothesized that a GCH1 promoter polymorphism associated with psychiatric illness, contributes to regulation of both GCH1 expression and BH 4 levels. We tested this hypothesis in 120 subjects (85 patients with schizophrenia or schizoaffective disorder and 35 controls): Patients with the rs10137071 A allele had significantly lower plasma biopterin than GG patients and controls. In additional samples we assessed the relationship between genotype and diagnosis (schizophrenia or control) on GCH1 expression in the prefrontal cortex ( n = 67 ) and peripheral leukocytes ( n = 53 ). We found a significant linear relationship between GCH1 and study group in the CNS and periphery, with A allele patients having lower expression. Finally, in antipsychotic naïve patients ( n = 13 ) we tested for an effect of medication on GCH1 : Expression rose significantly after the onset of medication, primarily in A allele patients. These data suggest the potential for personalized genetic approaches to ameliorating BH 4 deficits in schizophrenia-spectrum disorders. [ABSTRACT FROM AUTHOR]

Determining the accurate age of malaria vectors is crucial to measure the risk of malaria transmission. A group of fluorescent chemicals derived from a pyrimidine-pyrazine ring structure known as pteridines from the head, thorax and whole body of adult female Anopheles stephensi were identified and evaluated as a tool for chronological and physiological age determination of malaria vectors. The female mosquitoes were collected from an insectary colony at an interval of every 5 days, up to 30 days, and the pteridines of head, thorax and whole body were detected fluorometrically by high-pressure liquid chromatography (HPLC) using excitation and emission wavelengths of 365 and 455 nm, respectively. In addition, alteration of the pteridines compounds was compared between blood and sugar fed mosquito groups. Although four pteridines including pterin-6-carboxylic acid, biopterin, xanthopterin and isoxanthopterin were detected, some of them were absent in the head or thorax of mosquitoes. Levels of all four pteridines were similarly decreased in a linear manner throughout 30 days. No significant difference in alteration of pteridine compounds was observed between the two groups of blood or sugar fed mosquitoes. This result indicates that diet has a little effect on pteridines alteration. Age determination based on pteridines, as an age-grading technique, could be used for field collected mosquitoes, which have either sugar or blood meal. In addition, analyzing total pteridine fluorescence from only whole body could be a convenient method to estimate the age. [ABSTRACT FROM AUTHOR]

In cardiovascular and neurovascular diseases, an increase in oxidative stress and endothelial dysfunction has been reported. There is a reduction in tetrahydrobiopterin (BH 4 ), which is a cofactor for the endothelial nitric oxide synthase (eNOS), resulting in eNOS uncoupling. Studies of the enhancement of BH 4 availability have reported mixed results for improvement in endothelial dysfunction. Our understanding of the complex interactions of eNOS uncoupling, oxidative stress and BH 4 availability is not complete and a quantitative understanding of these interactions is required. In the present study, we developed a computational model for eNOS uncoupling that considers the temporal changes in biopterin ratio in the oxidative stress conditions. Using the model, we studied the effects of cellular oxidative stress (Q supcell ) representing the non-eNOS based oxidative stress sources and BH 4 synthesis (Q BH4 ) on eNOS NO production and biopterin ratio (BH 4 /total biopterins (TBP)). Model results showed that oxidative stress levels from 0.01 to 1 nM·s −1 did not affect eNOS NO production and eNOS remained in coupled state. When the Q supcell increased above 1 nM·s −1 , the eNOS coupling and NO production transitioned to an oscillatory state. Oxidative stress levels dynamically changed the biopterin ratio. When Q supcell increased from 1 to 100 nM·s −1 , the endothelial cell NO production, TBP levels and biopterin ratio reduced significantly from 26.5 to 2 nM·s −1 , 3.75 to 0.002 μM and 0.99 to 0.25, respectively. For an increase in BH 4 synthesis, the improvement in NO production rate and BH 4 levels were dependent on the extent of cellular oxidative stress. However, a 10-fold increase in Q BH4 at higher oxidative stresses did not restore the NO-production rate and the biopterin ratio. Our mechanistic analysis reveals that a combination of enhancing tetrahydrobiopterin level with a reduction in cellular oxidative stress may result in significant improvement in endothelial dysfunction. [ABSTRACT FROM AUTHOR]

End-stage renal disease (ESRD) is the inability of the kidneys to remove waste products from the blood. The most important factors causing ESRD that require hemodialysis are diabetes and hypertension. There are limited numbers of studies to evaluate tetrahydrobiopterin pathway in these patients. The aim of the study was to evaluate tetrahydrobiopterin pathway by measuring its important components, biopterin to creatinine concentrations and dihydropteridine reductase activities in diabetes and hypertension patients treated with/without hemodialysis. The patients undergoing hemodialysis were classified as diabetic nephropathy (n=21), hypertensive nephropathy (n=20) and others (n=30), while the controls consisted of healthy subjects (n=21), diabetic subjects (n=23) and hypertensive subjects (n=22) without any renal disorder. It was found that urinary biopterin to creatinine concentrations significantly increased in kidney failure patients undergoing hemodialysis compared to the healthy control group (p

The tetrahydrobiopterin (BH4) cofactor is essential for the activity of various enzymes, including phenylalanine (Phe) hydroxylase. In phenylketonuria (PKU) patients, who are chronically exposed to high Phe levels, high urinary excretion of BH4 metabolites neopterin and biopterin is observed. The aim of this longitudinal study was to investigate consistence and variability of the urinary excretion of pterins (neopterin and biopterin) in PKU patients in relation to age and concomitant blood Phe and tyrosine levels. The study was based on the result of 274 pterin examinations (3-13 exams per subject) performed in 47 PKU patients (aged 6 days to 37 years). Multivariate analysis showed that urinary biopterin and neopterin excretion was affected by age and concomitant blood Phe concentration. The influence of blood Phe on both biopterin and neopterin levels was greater in patients younger than 4 months. Later on, interindividual variability was higher than intraindividual variability for both biopterin and neopterin.Conclusion: Common metabolic (blood Phe levels) and individual (age) factors implicated in the assessment of PKU outcome account only marginally and transiently for the variability of neopterin and biopterin excretion in PKU patients. Other unknown homeostatic factors may probably affect the individual response to chronically elevated Phe levels. What is Known: • In PKU patients, a high urinary excretion of biopterin and neopterin is found. • Biopterin and neopterin excretion is influenced by age and phenylalanine levels. W hat is New: • Blood phenylalanine concentration is the major determinant on pterin excretion in PKU patients in the first months of life. • In older PKU patients, the influence of phenylalanine on pterin excretion is less prominent. [ABSTRACT FROM AUTHOR]

Guanosine 50-triphosphate (GTP) cyclohydrolase-I (GCYH-I) catalyzes the first step in folic acid biosynthesis in bacteria and plants, biopterin biosynthesis in mammals, and the biosynthesis of 7-deazaguanosine-modified tRNA nucleosides in bacteria and archaea. The type IB GCYH (GCYH-IB) is a prokaryotic-specific enzyme found in many pathogens. GCYH-IB is structurally distinct from the canonical type IA GCYH involved in biopterin biosynthesis in humans and animals, and thus is of interest as a potential antibacterial drug target. We report kinetic and inhibition data of Neisseria gonorrhoeae GCYH-IB and two high-resolution crystal structures of the enzyme; one in complex with the reaction intermediate analog and competitive inhibitor 8-oxoguanosine 50-triphosphate (8-oxo-GTP), and one with a tris(hydroxymethyl)aminomethane molecule bound in the active site and mimicking another reaction intermediate. Comparison with the type IA enzyme bound to 8-oxo-GTP (guanosine 50-triphosphate) reveals an inverted mode of binding of the inhibitor ribosyl moiety and, together with site-directed mutagenesis data, shows that the two enzymes utilize different strategies for catalysis. Notably, the inhibitor interacts with a conserved active-site Cys149, and this residue is S-nitrosylated in the structures. This is the first structural characterization of a biologically S-nitrosylated bacterial protein. Mutagenesis and biochemical analyses demonstrate that Cys149 is essential for the cyclohydrolase reaction, and S-nitrosylation maintains enzyme activity, suggesting a potential role of the S-nitrosothiol in catalysis. [ABSTRACT FROM AUTHOR]

Background (6R)-5,6,7,8-tetrahydrobiopterin (BH4) deficiencies are rare inherited defects of synthesis or regeneration of BH4. Due to the resulting hyperphenylalaninemia (HPA), some of them are detected by newborn screening and require the assessment of the pattern of neopterin (Neo) and biopterin (Bio) excretion in urine to be confirmed. Aim of present study was to develop a method for the measurement of these diagnostic biomarkers in dried blood spot (DBS). Methods After DBS extraction, samples were filtered and injected into the UPLC column coupled with a tandem mass spectrometer working in positive electrospray ionization. Results The chromatographic separation was accomplished in 6 min. The LoQ was 0.57 and 1.45 nmol/l of blood for Neo and Bio respectively and the response was linear over the range 0–100 nmol/l of blood. The within- and between-day imprecision was < 6.4 and 10.8% respectively. Reference ranges for newborns, infants and children/adult were established. The method was tested in 11 patients affected by BH4 defects. Conclusions The assessment of Neo and Bio in DBS is reliable and sensitive and may be proposed as a second tier test for the newborns with hyperphenylalaninemia (HPA) as well as a new potential diagnostic tool for symptomatic subjects with BH4 deficiencies. [ABSTRACT FROM AUTHOR]

The nitric oxide synthases ( NOS) catalyze a two-step oxidation of l-arginine (Arg) to generate NO. In the first step, O2 activation involves one electron being provided to the heme by an enzyme-bound 6 R-tetrahydro- l-biopterin cofactor (H4B), and the H4B radical must be reduced back to H4B in order for NOS to continue catalysis. Although an NADPH-derived electron is used to reduce the H4B radical, how this occurs is unknown. We hypothesized that the NOS flavoprotein domain might reduce the H4B radical by utilizing the NOS heme porphyrin as a conduit to deliver the electron. This model predicts that factors influencing NOS heme reduction should also influence the extent and rate of H4B radical reduction in kind. To test this, we utilized single catalytic turnover and stop-freeze methods, along with electron paramagnetic resonance spectroscopy, to measure the rate and extent of reduction of the 5-methyl-H4B radical formed in neuronal NOS (n NOS) during Arg hydroxylation. We used several n NOS variants that supported either a slower or faster than normal rate of ferric heme reduction. We found that the rates and extents of n NOS heme reduction correlated well with the rates and extents of 5-methyl-H4B radical reduction among the various n NOS enzymes. This supports a model where the heme porphyrin transfers an electron from the NOS flavoprotein to the H4B radical formed during catalysis, revealing that the heme plays a dual role in catalyzing O2 activation or electron transfer at distinct points in the reaction cycle. [ABSTRACT FROM AUTHOR]

Neopterin, as a non-conjugated pteridine, is synthesized from guanosine triphosphate and its production is upregulated upon the activation of cellular immune response. Alterations of pteridines in body fluids are known to correlate well with existing diseases and stages, prognosis, clinical outcomes and survival data. It is of advantage to have a pteridine database of healthy volunteers to determine normal values. Thereby, especially in children there is no detailed study on pteridine levels. The aim of this study is to initiate the establishment of pteridine database of healthy children in our country. Serum neopterin levels were analysed by enzyme-linked immunosorbent assay. Urinary neopterin and biopterin levels and serum kynurenine, tryptophan levels and kynurenine/tryptophan ratio as an estimate of tryptophan breakdown were assessed with high-pressure liquid chromatography in serum and urine samples of 55 children aged between 3 months and 10 years. The results were evaluated within the subgroups of different ages and sex. Pteridine pathway components were found to be higher in children compared to adults. Higher levels of pteridine pathway components observed within the first years of life may reflect the rapid maturation of the immune system, and environmental adaptation and/or insufficiency of defence systems. On the other hand, it may also relate to a higher frequency of infections not (yet) manifested clinically.

In Helicobacter pylori seropositive individuals, the subsequent risk of non-cardia gastric cancer development is frequently attributed to the infection agent. Although the link between inflammation and gastric cancer is formally recognized, the molecular basis of evading mechanisms of Helicobacter pylori from host immune system remains unclear.

Biopterin as a stable metabolite is produced by oxidation of 5,6,7,8-tetrahydrobiopterin (BH4). It is known that many diseases may cause changes in BH4 concentration and/or dihydropteridine reductase (DHPR) enzyme activity. There is only a limited number of studies correlating DHPR activity in malignancies. The main goal of the present study was to evaluate alterations in the DHPR activity and biopterin levels in patients with breast and thyroid cancers. The breast cancer patients (n=24) were compared to patients with benign breast diseases (n=19) and controls (n=30). In addition; the patients with thyroid cancer (n=17) were compared to patients with benign thyroid diseases (n=42) and the control group. We did not observe any significant difference between the benign disorders and the malignancies. Biopterin concentrations in patients with benign and malign thyroid and breast diseases were lower than the controls (all p

The aim of the presented study was to assess the effect of a single administration of Fe3+-dextran on immune cell counts and pterin biomolecule production as novel sensors of the piglets' immune system activation, and to determine concentrations of cortisol, a traditional hormonal biosensor of the stress response. Pterins (neopterin and biopterin) in the piglets' blood serum were analyzed by separation using reversed-phase HPLC. A single dose of Fe3+-dextran produced a special stress situation in the piglets' organism which manifested itself by an increased production of neopterin (p < 0.05) and biopterin (p < 0.01) in the experimental piglets. Changes in cortisol concentrations and leukocyte counts were influenced by handling stress and were not specifically correlated to iron dextran application. Iron concentrations in the internal environment of the experimental piglets’ group were higher by an order of magnitude compared with the controls, and the highest serum concentrations of iron (p < 0.01) were reached 24 h following Fe3+-dextran administration. The data presented offer a new perspective on the evaluation of stress situations in the animal organism and, not least importantly, extends the rather modest current list of references on the role of pterins in livestock animals.

The profiles of unconjugated pteridines such as neopterin and biopterin and of tryptophan metabolism change in many pathologic conditions such as rheumatoid arthritis, malignancies and autoimmune diseases and are related to the release of pro-inflammatory cytokines. However, studies on potential effects of drugs are very limited. This study investigates whether meloxicam, a non-steroidal anti-inflammatory drug, may cause any alteration of the pteridine pathway and tryptophan metabolism in 22 patients who admitted to the Physical and Rehabilitation Medicine outpatient clinic in the Hacettepe University Hospital and suffering from various musculoskeletal pain conditions without systemic inflammation. Urine concentrations of neopterin, biopterin and xanthopterin, and serum concentrations of neopterin, tryptophan, and kynurenine, and the activity of dihydropteridine reductase were measured at the beginning of the therapy and after a 15 days treatment with meloxicam. Average urine concentrations (± S.E.M.) of pterins before therapy (neopterin: 263 ± 15.6 μmol/mol creatinine, biopterin: 72.4 ± 13.4 μmol/mol creatinine, and xanthopterin: 675 ± 93.0 μmol/mol creatinine) were not changed during meloxicam therapy (neopterin: 230 ± 30.7 μmol/mol creatinine = 88% of baseline, biopterin 115 ± 26.5 μmol /mol creatinine = 159% of baseline, and xanthopterin 684 ± 88.0 μmol/mol creatinine = 101% of baseline; all differences not significant). The activity of dihydropteridine reductase was 6.02 ± 0.21 nmol/cytochrome C reduced/min/5 mm disc before and 6.28 ± 0.18 nmol/cytochrome C reduced/min/5 mm after treatment (= 104% of baseline; p = 0.088). Serum concentrations of neopterin 6.04 ± 0.33 nmol/L before treatment increased to 6.89 ± 0.48 nmol/L after treatment (= 114% of baseline; p

Pteridines, such as neopterin, biopterin, and tetrahydrobiopterin (BH 4), may be involved in depression pathophysiology owing to their links to immune-inflammatory response, oxidative and nitrosative stress, and monoaminergic transmission. Nonetheless, studies assessing pteridines in depression are inconsistent. We conducted a systematic review and meta-analysis of observational studies comparing blood pteridine concentrations between subjects with depression and healthy controls (HCs). We searched Embase, MEDLINE, and PsycInfo for articles indexed through November 2021. Study quality was appraised, evaluating age and gender comparability between groups, sample representativeness, and methods to assess depression. Random-effects meta-analyses were carried out, generating pooled standardized mean differences (SMDs). Heterogeneity across studies was estimated using the I2 statistic. Twenty-four studies, involving 3075 subjects, were included. Individuals with depression showed blood neopterin concentrations higher than HCs (k = 19; SMD = 0.36; p < 0.001) with moderate heterogeneity across studies (I2 = 58.2%). No moderating role of age, gender, or type of blood sample was found. Sensitivity analyses showed no impact of inconsistency and quality of studies on findings. Neopterin concentrations were higher among individuals with major depressive disorder compared to HCs (SMD = 0.44; p < 0.001). This held true also when considering only drug-free subjects (SMD = 0.68; p = 0.003). No differences in biopterin concentrations were found between subjects with depression and HCs (k = 5; SMD = –0.35; p = 0.086), though this result was limited by inconsistency of findings (I2 = 77.9%) and quality of studies. Finally, no sufficient data were available for a meta-analysis on BH 4. As a whole, our work partly supports the hypothesis of an imbalance of pteridine metabolism in depression. • We conducted a meta-analysis on blood pteridine concentrations in depression. • Blood neopterin concentrations were higher in depression. • Drug-free participants with depression showed a greater increase. • No differences in blood biopterin concentrations were estimated. • Our meta-analysis partly supports imbalances of pteridine metabolism in depression. [ABSTRACT FROM AUTHOR]

Tetrahydrobiopterin (BH4) is an essential cofactor of nitric oxide synthase (NOS) and aromatic amino acid hydroxylases. BH4 and 7,8-dihydrobiopterin (BH2) are metabolically interchangeable at the expense of NADPH. Exogenously administered BH4 can be metabolized by the body, similar to vitamins. At present, synthetic BH4 is used as an orphan drug for patients with inherited diseases requiring BH4 supplementation. BH4 supplementation has also drawn attention as a means of treating certain cardiovascular symptoms, however, its application in human patients remains limited. Here, we tracked biopterin (BP) distribution in blood, bile, urine, liver, kidney and brain after BH4 administration (5 mg/kg rat, i.v.) with or without prior treatment with probenecid, a potent inhibitor of uptake transporters particularly including organic anion transporter families such as OTA1 and OAT3. The rapid excretion of BP in urine was driven by elevated blood concentrations and its elimination reached about 90% within 120 min. In the very early period, BP was taken up by the liver and kidney and gradually released back to the blood. BH4 administration caused a considerable decrease in the BH4% in blood BP as an inevitable compensatory process. Probenecid treatment slowed down the decrease in blood BP and simultaneously inhibited its initial rapid excretion in the kidney. At the same time, the BH4% was further lowered, suggesting that the probenecid-sensitive BP uptake played a crucial role in BH2 scavenging in vivo. This suggested that the overproduced BH2 was taken up by organs by means of the probenecid-sensitive process, and was then scavenged by counter-conversion to BH4 via the BH4 salvage pathway. Taken together, BH4 administration was effective at raising BP levels in organs over the course of hours but with extremely low efficiency. Since a high BH2 relative to BH4 causes NOS dysfunction, the lowering of the BH4% must be avoided in practice, otherwise the desired effect of the supplementation in ameliorating NOS dysfunction would be spoiled. [ABSTRACT FROM AUTHOR]

Background: Xerostomia is defined as dry mouth resulting from a change in the amount or composition of saliva and is often a major oral health complication associated with diabetes mellitus (DM). Studies have shown that xerostomia is more common in females at the onset of DM. Evidence suggests that nitric oxide (NO) plays a critical role in healthy salivary gland function. However, the specific mechanisms by which NO regulates salivary gland function at the onset of DM have yet to be determined. This study has two aims: 1) to determine whether protein expression or dimerization of NO synthase enzymes (neuronal [nNOS] and endothelial [eNOS]) are altered in the onset of diabetic xerostomia; and 2) to determine whether the changes in nNOS/eNOS protein expression or dimerization are correlated with changes in NO cofactor tetrahydrobiopterin (BH4) biosynthetic enzymes (guanosine triphosphate cyclohydrolase-1 and dihydrofolate reductase).Methods: Functional and Western blot studies were performed in streptozotocin-induced and control Sprague-Dawley female rats with DM (type 1 [t1DM]) using standardized protocols. Confirmation of xerostomia was determined by increased water intake and decreased salivary flow rate.Results: The results showed that in female rats with DM, salivary hypofunction is correlated with decreased submandibular and parotid gland sizes. The results also show a decrease in NOS and BH4 biosynthetic enzyme in submandibular glands.Conclusions: These results indicate that a decrease in submandibular NO-BH4 protein expression may provide insight pertaining to mechanisms for the development of hyposalivation in DM-induced xerostomia. Furthermore, understanding the role of the NO-BH4 pathway may give insight into possible treatment options for the patient with DM experiencing xerostomia. [ABSTRACT FROM AUTHOR]

Tetrahydrobiopterin (BH 4 ) represents a potential strategy for the treatment of cardiac remodeling, fibrosis and/or diastolic dysfunction. The effects of oral treatment with BH 4 (Sapropterin™ or Kuvan™) are however dose-limiting with high dose negating functional improvements. Cardiomyocyte-specific overexpression of GTP cyclohydrolase I (mGCH) increases BH 4 several-fold in the heart. Using this model, we aimed to establish the cardiomyocyte-specific responses to high levels of BH 4 . Quantification of BH 4 and BH 2 in mGCH transgenic hearts showed age-based variations in BH 4 :BH 2 ratios. Hearts of mice (<6 months) have lower BH 4 :BH 2 ratios than hearts of older mice while both GTPCH activity and tissue ascorbate levels were higher in hearts of young than older mice. No evident changes in nitric oxide (NO) production assessed by nitrite and endogenous iron–nitrosyl complexes were detected in any of the age groups. Increased BH 4 production in cardiomyocytes resulted in a significant loss of mitochondrial function. Diminished oxygen consumption and reserve capacity was verified in mitochondria isolated from hearts of 12-month old compared to 3-month old mice, even though at 12 months an improved BH 4 :BH 2 ratio is established. Accumulation of 4-hydroxynonenal (4-HNE) and decreased glutathione levels were found in the mGCH hearts and isolated mitochondria. Taken together, our results indicate that the ratio of BH 4 :BH 2 does not predict changes in neither NO levels nor cellular redox state in the heart. The BH 4 oxidation essentially limits the capacity of cardiomyocytes to reduce oxidant stress. Cardiomyocyte with chronically high levels of BH 4 show a significant decline in redox state and mitochondrial function. [ABSTRACT FROM AUTHOR]

Five previously undescribed biopterin glycosides, microcystbiopterin A–E, were isolated from the extracts of two bloom materials of Microcystis spp. collected from a fishpond (IL-337) and Lake Kinneret (IL-347), Israel. The structure of the pterins was established by interpretation of their UV, CD, 1D and 2D NMR spectra and HR mass measurements. Microcystbiopterin D is the first heptose containing pterin glycoside to be reported in the literature. Their antimicrobial and cytotoxic properties were evaluated but all were found not active in both assays. [ABSTRACT FROM AUTHOR]

Cerebrospinal fluid (CSF) neopterin been previously reported in various diseases. In this study CSF neopterin, biopterin, and nitrite/nitrate (NOx) Contents were measured and the correlation between them in child patients with various kinds of neurological diseases were investigated. Changes in the CSF neopterin levels in patients with bacterial meningitis were similar to those previously reported for those with bacterial meningitis; on the 2th hospital day they were significantly higher than on admission, and on the 6th hospital day they were tapered. The CSF biopterin levels and CSF NOx content in patients with bacterial meningitis on admission were significantly higher than those with other categories and were decreased gradually. Although patients with high levels of CSF neopterin tended to have high CSF biopterin levels in any categories, there was no significant correlation between CSF neopterin and biopterin levels. The CSF biopterin and NOx levels in patients with convulsions were higher than those with aseptic meningitis. Since the neuro-protective or anticonvulsant role for NO was previously reported, high CSF biopterin and NOx levels in patients having epilepsy or febrile convulsions may be regarded as one of the endogenous mechanisms for recovery from an overexcitatory brain in patients with convulsive diseases.