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Abstract In the progresses of prostate cancer (PCa), sarcosine levels increase not only in cells but also in urine samples, suggesting that the measurement of sarcosine in urine can benefit in the creation of non-invasive diagnostic methods for PCa. In this work, a biosensor has been developed for the quantification of sarcosine via the cross coupled chemical and electrochemical reactions with a limit of detection (LOD) of 0.4 nM (S/N = 3). The biosensor has been successfully applied to the analysis of sarcosine in urine samples, which has a great potential application in noninvasive clinical diagnosis of PCa. Graphical abstract An amperometric biosensor has been developed for the quantification of sarcosine via the detection of H 2 O 2 generated from the catalyzed oxidation of sarcosine by sarcosine oxidase (SOX) in the urine. In order to improve the sensitivity of the biosensor, the PAA are immobilized on the electrode surface. PAA is a polymer that would act as a reducing agent on the gold electrode surface and improve the sensitivity of the biosensor through the cross coupled chemical and electrochemical reactions with a limit of detection (LOD) of 0.4 nM (S/N = 3). Unlabelled Image Highlights • A simple and sensitive biosensor for the detection of sarcosine has been developed. • The cross coupled chemical and electrochemical reactions are used for signal amplification skillfully. • Sarcosine has been detected effectively in complex biological samples. • This method may have potential applications in noninvasive clinical diagnosis of PCa. [ABSTRACT FROM AUTHOR]

Abstract Background Plasma and serum are the most widely used matrices in clinical studies. However, some variability in absolute concentrations of metabolites are likely to be observed in these collection tubes matrices. Methods We analyzed 189 metabolites using the same protocol for quantitative targeted metabolomics (LC-MS/MS AbsoluteIDQ p180 Kit Biocrates) in three types of samples, serum, plasma EDTA and citrate, of 80 subjects from the Cooperative Health Research In South Tyrol cohort (40 healthy elderly and 40 healthy young). Results The concentration levels were higher in serum than citrate and EDTA, in particular for amino acids and biogenic amines. The average Pearson's correlation coefficients were however always higher than 0.7 for these two classes of metabolites. We could also demonstrate that blank EDTA vacutainer tubes contain a significant amount of sarcosine. Finally, we compared the metabolome of young people against elderly subjects and found that the highest number of metabolites significantly changing with age was detected in serum. Conclusion Serum samples provide higher sensitivity for biomarker discovery studies. Due to the presence of spurious amount of sarcosine in vacutainer EDTA tubes, plasma EDTA is not suitable for studies requiring accurate quantification of sarcosine. Highlights • Comparison of concentration levels for 189 metabolites in serum, plasma citrate and EDTA in 80 healthy subjects. • Serum samples provide higher concentration levels and citrate samples provide lower biological variation. • Blank vacutainer tubes used for the collection of plasma EDTA samples contain high amount of sarcosine. [ABSTRACT FROM AUTHOR]

[Display omitted] • A predicted sarcosine oxidase gene from T. roseum was expressed in E. coli and reconstructed with natural cofactors or analogues. • A soluble mutant TrSOX (F235V/F339L) was prepared and characterized for the first time. • The reconstruction could not significantly affect the 2D structure of TrSOXs • Halogen atoms on the position 7- or 8-site of isoalloxazine ring could promote the catalytic properties or affect the substrate selectivity. A predicted sarcosine oxidase (SOX) gene from Thermomicrobium roseum DSM 5159 was expressed, reconstructed, and characterized. Regarding sequence alignment, T. roseum sarcosine oxidase (TrSOX) showed ∼35% similarity with several bacterial SOXs and ∼25% similarity with MTOX from E. coli. Regarding structural modeling and molecular docking, TrSOX interacted with natural flavin cofactors and flavin analogues including flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) in the cofactor binding site. The TrSOX gene was built on a plasmid containing a L-rhamnose-inducible promoter and expressed in E. coli BL21 (DE3) as inclusion body. In addition, a soluble mutant (F235 V/F339 L) was identified in a random mutagenesis screening experiment. The inclusion body associated with the native gene were reconstructed with natural or modified ligands. Circular dichroism detection revealed the stable secondary structure of the enzyme, which was unaffected by the ligands. Furthermore, regarding reconstructed TrSOX with a ligand modified at the 7- or 8-position by a halogen atom, the K m , k cat , relative specificity, optimal reaction temperature, and pH adaptability were significantly different compared with refolded TrSOX-FAD, and flavin analogues affected the catalytic properties of this enzyme. [ABSTRACT FROM AUTHOR]

Layer-by-layer composite films of carbon nanotubes (CNTs) within a chitosan matrix with sarcosine oxidase enzyme and capped with Nafion have been developed and optimized as a versatile 1st generation amperometric sarcosine biosensing platform that operates successfully both as an isolated sarcosine sensor as well as a functional component within a creatinine sensor. Accurate measurement of sarcosine in urine and creatinine in blood may help with early diagnosis of diseases such as prostate cancer and renal failure, respectively. In this study, each material within the film is systematically optimized toward sarcosine sensitivity, including a critical evaluation of different CNTs effect on sensing performance. Films featuring carboxylic acid–modified single–walled carbon nanotubes and strategic enzyme doping were shown to be most effective sarcosine sensing platforms, exhibiting excellent sensitivity (~0.5 μA/mM), a linear response (≤0.75 mM), fast response time (8 s), low limits of detection (~6 μM), as well as both continuous use stability (7 days) and effective shelf life (>12 days). Operation of the sarcosine sensor was demonstrated in a sarcosine-spiked urine matrix, detecting the analyte at physiologically relevant concentrations (≥20 μM) and successfully quantifying sarcosine-spiked urine samples at 20, 40, and 90 μM ( n  = 9, 7, and 3) with high average percent recoveries (>98%) and low relative error. The sarcosine sensing platform was also adapted to a 1st generation creatinine biosensing scheme in which the sarcosine enzymatic reaction is critical to a tri-enzymatic cascade event. The creatinine sensor yielded sensitivity of ~0.6 μA/mM, similar sensing performance parameters to the sarcosine sensor, and was effectively operated in blood serum at physiologically relevant creatinine concentrations (>1 mM). The demonstrated functionality of these sensors in their respective biological fluids at physiological concentrations of the analyte species suggests potential clinical application as diagnostic tools. [ABSTRACT FROM AUTHOR]

Sarcosine is a newly discovered effective biomarker for prostate cancer. However, the low concentration of sarcosine in tissue cells, plasma or urine blocks the development of sarcosine biosensors. In this manuscript, porous zeolitic imidazolate framework-8 (ZIF8) was synthesized and was used as carriers to load nano platinum (Pt@ZIF8). The porous structure of ZIF8 helped to stabilize the nano platinum and keep its high catalytic activity. The Pt@ZIF8 modified sarcosine biosensor had good response toward sarcosine due to its unique structure and morphology. The linear range of the as prepared biosensor is from 5 to 30 μM, which is consistent with the detection demand. The prepared sarcosine biosensor is suitable to be developed as a kind of portable diagnostic facilities for prostate cancer. [ABSTRACT FROM AUTHOR]

Abstract: Objective: Augmentation of sarcosine, a natural inhibitor of the glycine transporter type I, normalizes glutamatergic neurotransmission, having beneficial impact on primary negative symptoms in schizophrenia and may also influence immune system and interleukin 6 (IL‐6) levels. Aim: Finding a relationship between initial IL‐6 serum concentrations or its changes and severity of symptoms as a result of sarcosine addition to stable antipsychotic treatment. Method: Fifity‐eight individuals with schizophrenia with predominantly negative symptoms completed a 6‐month randomized, double‐blind placebo‐controlled prospective study. Patients received 2 g of sarcosine (n = 29) or placebo (n = 30) daily per os. We measured IL‐6 levels and severity of symptoms at the beginning, after 6 weeks and 6 months. As main clinical tools, we used Positive and Negative Syndrome Scale (PANSS) and Calgary depression scale for schizophrenia (CDSS). Results: Augmentation with sarcosine had no effect on IL‐6 serum levels in all time points. We noted significant improvements in negative symptoms, general psychopathology, and total PANSS score in the sarcosine group. We found correlation of initial serum IL‐6 with severity of positive symptoms and negative association between IL‐6 levels reduction and positive symptoms reduction. Conclusions: Sarcosine does not significantly affect IL‐6 concentrations but IL‐6 may be involved in mechanisms related to the presence of positive symptoms. [ABSTRACT FROM AUTHOR]

An enzyme-coupled colorimetric assay for quantification of urinary sarcosine was developed. The proposed method is a specific reaction based on hydrogen peroxide (H2O2) formation via sarcosine oxidase (SOX). The liberated H2O2 reacts with Amplex Red in the presence of horseradish peroxidase (HRP) to produce the redfluorescent oxidation product, resorufin, which can be measured spectrophotometrically (OD570). The method was performed in the 96-well microtiter plate. Reaction conditions, such as pH and reaction time were optimized. At the optimum conditions, the limit of detection (LOD) and quantification (LOQ) were found to be 0.7 and 1 µM, respectively. A good linearity was revealed with a coefficient of 0.990. The assay showed no significant interference from ascorbic acid, glucose and bilirubin. In addition, it is extremely specific for sarcosine rather than other amino acids. The determination of sarcosine in human urine displayed high accuracy and good reproducibility. This method is promising to differentiate prostate cancer patients from healthy subjects according to urinary sarcosine level. Altogether, this study provides a rapid, simple and specific tool to determine urinary sarcosine which could be useful for prostate cancer diagnosis. [ABSTRACT FROM AUTHOR]

An improved amperometric biosensor for detection of creatinine was developed based on immobilization of nanoparticles (NPs) of creatininase (CA), creatinase (CI), and sarcosine oxidase (SOx) onto glassy carbon (GC) electrode. Transmission electron microscopy (TEM) and fourier transform infrared spectroscopy (FTIR) were employed for characterization of enzyme nanoparticles (ENPs). The GC electrode was characterized by scanning electron microscopy (SEM), cyclic voltammetry (CV) and electrochemical impedance spectra (EIS) at different stages of its amendment. The biosensor showed optimum response within 2s at pH 6.0 in 0.1 M sodium phosphate buffer and 25 °C, when operated at 1.0 V against Ag/AgCl. Biosensor exhibited wider linear range from 0.01 μM to 12 μM with a limit of detection (LOD) of 0.01 μM. The analytical recoveries of added creatinine in sera were 97.97 ± 0.1% for 0.1 mM and 98.76 ± 0.2% for 0.15 mM, within and between batch coefficients of variation (CV) were 2.06% and 3.09% respectively. A good correlation (R 2 = 0.99) was observed between sera creatinine values obtained by standard enzymic colorimetric method and the present biosensor. This biosensor measured creatinine level in sera of apparently healthy subjects and persons suffering from renal and muscular dysfunction. The ENPs electrode lost 10% of its initial activity within 240 days of its regular uses, when stored at 4 °C. [ABSTRACT FROM AUTHOR]

Novel penta- and tetra-cyclic cage-like compounds and dispiro heterocycles were obtained as a result of the microwave-assisted three-component domino reactions of heterocyclic or carbocyclic ketones, ninhydrin and sarcosine under solvent- and catalyst-free conditions. Interestingly, heterocyclic ketones afforded fused polycyclic cage-like compounds, whereas the carbocyclic ketones led to the formation of dispiro heterocycles. [ABSTRACT FROM AUTHOR]

Indicator displacement assay plays a fundamental role in the development of chemosensors. We explored here an ingenious yet effective strategy for amperometric assay of electrochemically inactive sarcosine based on an indicator displacement principle, in which 1,2-naphthoquinone-4-sulphonic acid sodium salt (NQS) was proposed as the receptor and the electroactive Ru(NH 3 ) 6 3+ cations used as an indicator. Due to the stronger binding affinity of the NQS toward sarcosine than toward Ru(NH 3 ) 6 3+ , the developed amperometric indicator displacement assay (A-IDA) exhibits high selectivity and excellent sensitivity toward sarcosine determination as well as with a lower detection limit (30.00 nM, S/N =3). [ABSTRACT FROM AUTHOR]

Sarcosine, an N -methyl- d -aspartate receptor enhancer, can improve depression-like behavior in rodent models and depression in humans. We found that a single dose of sarcosine exerted antidepressant-like effects with rapid concomitant increases in the mammalian target of rapamycin (mTOR) signaling pathway activation and enhancement of α-amino-3-hydroxy-5-methylisoxazole-4-propionate receptor (AMPAR) membrane insertion. Sarcosine may play a crucial role in developing novel therapy for depression. For a detailed understanding of sarcosine, this study examined the effects of long-term sarcosine treatment on the forced swim test (FST), mTOR signaling, and AMPAR membrane insertion in rats. The effects of long-term sarcosine treatment were examined in naive rats and rats exposed to chronic unpredictable stress (CUS). Long-term sarcosine treatment (560 mg/kg/d for 21 d) significantly ameliorated the increased immobility induced by CUS in the FST, reaffirming the potential role of sarcosine as an antidepressant for depressed patients. The same long-term treatment exhibited no such effect in naive rats despite increased mTOR activation and AMPAR membrane insertion in both groups. Our findings clearly show CUS-exposed rats are sensitive to long-term sarcosine treatment in FST and the response at the same dose is absent in naïve rats. Nevertheless, the distinct sensitivity to long-term sarcosine treatment in rats with or without CUS is not associated with the activated mTOR signaling pathway or increased AMPAR membrane insertion. Additionally, understanding the behavioral and molecular basis of distinct responses is vital important for developing personalized treatment programs to increase the probability of success when treating depression. [ABSTRACT FROM AUTHOR]

Methylphenidate, a stimulant that activates dopaminergic and noradrenergic function, is an important agent in the treatment of attention deficit hyperactivity disorder (ADHD). Sarcosine, a glycine transporter-1 inhibitor, may also play a role in treating ADHD by modulating the glutamatergic neurotransmission system through activating N-methyl-D-aspartate type glutamate receptors. This study aimed to assess the efficacy of sarcosine in treating children with ADHD. We conducted a six-week, randomized, double-blind, placebo-controlled clinical trial. The primary outcome measures were those on the Inattention, Hyperactivity/impulsivity, and oppositional defiant disorder (ODD) subscales of the Swanson, Nolan, and Pelham, version IV scale. Efficacy and safety were measured bi-weekly. A total of 116 children with ADHD were enrolled. Among them, 48 (83%) of the 58 sarcosine recipients and 44 (76%) of the 58 placebo recipients returned for the first post-treatment visit. The missing data values were imputed by the last observation carry forward method. From a multiple linear regression analysis, using the generalized estimating equation approach, and an intention to treat analysis, the efficacy of sarcosine marginally surpassed that of placebo at weeks 2, 4, and 6, with p-values=0.01, 0.026, and 0.012, respectively, although only for ODD symptoms. Treatment of ADHD by sarcosine (0.03 g/kg/day) was well tolerated. Sarcosine could possibly be a novel agent for managing ODD symptoms in the context of ADHD. However, future larger-scale studies are warranted to optimize its dosage. [ABSTRACT FROM AUTHOR]

A new method based on headspace programmed-temperature vaporizer gas chromatography with mass spectrometry has been developed and validated for the determination of amino acids (alanine, sarcosine, ethylglycine, valine, leucine, and proline) in human urine samples. Derivatization with ethyl chloroformate was employed successfully to determine the amino acids. The derivatization reaction conditions as well as the variables of the headspace sampling were optimized. The existence of a matrix effect was checked and the analytical characteristics of the method were determined. The limits of detection were 0.15-2.89 mg/L, and the limits of quantification were 0.46-8.67 mg/L. The instrumental repeatability was 1.6-11.5%. The quantification of the amino acids in six urine samples from healthy subjects was performed with the method developed with the one-point standard additions protocol, with norleucine as the internal standard. [ABSTRACT FROM AUTHOR]

The discovery of a receptor that binds prorenin and renin in human endothelial and mesangial cells highlights the possible effect of renin-independent prorenin in the resumption of meiosis in oocytes that was postulated in the 1980s.This study aimed to identify the (pro)renin receptor in the ovary and to assess the effect of prorenin on meiotic resumption. The (pro)renin receptor protein was detected in bovine cumulus-oocyte complexes, theca cells, granulosa cells, and in the corpus luteum. Abundant (pro)renin receptor messenger ribonucleic acid (mRNA) was detected in the oocytes and cumulus cells, while prorenin mRNA was identified in the cumulus cells only. Prorenin at concentrations of 10 −10 , 10 −9 , and 10 −8 M incubated with oocytes co-cultured with follicular hemisections for 15 h caused the resumption of oocyte meiosis. Aliskiren, which inhibits free renin and receptor-bound renin/prorenin, at concentrations of 10 −7 , 10 −5 , and 10 −3 M blocked this effect ( P < 0.05). To determine the involvement of angiotensin II in prorenin-induced meiosis resumption, cumulus-oocyte complexes and follicular hemisections were treated with prorenin and with angiotensin II or saralasin (angiotensin II antagonist). Prorenin induced the resumption of meiosis independently of angiotensin II. Furthermore, cumulus-oocyte complexes cultured with forskolin (200 μM) and treated with prorenin and aliskiren did not exhibit a prorenin-induced resumption of meiosis ( P < 0.05). Only the oocytes’ cyclic adenosine monophosphate levels seemed to be regulated by prorenin and/or forskolin treatment after incubation for 6 h. To the best of our knowledge, this is the first study to identify the (pro)renin receptor in ovarian cells and to demonstrate the independent role of prorenin in the resumption of oocyte meiosis in cattle. [ABSTRACT FROM AUTHOR]

Aim: Find changes in matrix metallopeptidase-9 (MMP-9) levels during augmentation of antipsychotic treatment with sarcosine and a relationship between schizophrenia symptoms severity and initial level of MMP-9. Method: Fifty-eight patients with diagnosis of schizophrenia with predominant negative symptoms participated in a six-month prospective RCT (randomized controlled trial). The patients received two grams of sarcosine (n = 28) or placebo (n = 30) daily. At the beginning, after six weeks and after six months MMP-9 levels were measured. Severity of symptomatology was assessed with the Positive and Negative Syndrome Scale (PANSS) and Calgary Depression Scale for Schizophrenia (CDSS). Results: MMP-9 serum levels were stable after six weeks and six months in both groups. We noted improvement in negative symptoms, general psychopathology and total PANSS score in sarcosine group compared to placebo; however, there was no correlations between serum MMP-9 concentrations and PANSS scores in all assessments. Initial serum MMP-9 concentrations cannot be used as an improvement predictor acquired during sarcosine augmentation. Conclusions: Our results indicate that either MMP-9 is not involved in the N-methyl-D-aspartate (NMDA)-dependent mechanism of sarcosine action in terms of clinical parameters or sarcosine induced changes in peripheral MMP-9 concentrations cannot be detected in blood assessments. [ABSTRACT FROM AUTHOR]

In the present study, we developed an acute chemically induced model of sarcosinemia in Wistar rats. Wistar rats of 7, 14 and 21 postpartum days received sarcosine intraperitoneally in doses of 0.5 mmol/Kg of body weight three time a day at intervals of 3 h. Control animals received saline solution (NaCl 0.85 g%) in the same volume (10 mL/Kg of body weight). The animals were killed after 30 min, 1, 2, 3 or 6 h after the last injection and the brain and the blood were collected for sarcosine measurement. The results showed that plasma and brain sarcosine concentrations achieved levels three to four times higher than the normal levels and decreased in a time-dependent way, achieving normal levels after 6 hours. Considering that experimental animal models are useful to investigate the pathophysiology of human disorders, our model of sarcosinemia may be useful for the research of the mechanisms of neurological dysfunction caused by high tissue sarcosine levels. [ABSTRACT FROM AUTHOR]

Sarcosine, an endogenous amino acid, is a competitive inhibitor of the type I glycine transporter and an N-methyl-D-aspartate receptor (NMDAR) coagonist. Recently, we found that sarcosine, an NMDAR enhancer, can improve depression-related behaviors in rodents and humans. This result differs from previous studies, which have reported antidepressant effects of NMDAR antagonists. The mechanisms underlying the therapeutic response of sarcosine remain unknown. This study examines the role of mammalian target of rapamycin (mTOR) signaling and a-amino-3-hydroxy-5-methylisoxazole-4-propionate receptor (AMPAR) activation, which are involved in the antidepressant-like effects of several glutamatergic system modulators. The effects of sarcosine in a forced swim test (FST) and the expression levels of phosphorylated mTOR signaling proteins were examined in the absence or presence of mTOR and AMPAR inhibitors. In addition, the influence of sarcosine on AMPAR trafficking was determined by analyzing the phosphorylation of AMPAR subunit GluR1 at the PKA site (often considered an indicator for GluR1 membrane insertion in neurons). A single injection of sarcosine exhibited antidepressant-like effects in rats in the FST and rapidly activated the mTOR signaling pathway, which were significantly blocked by mTOR inhibitor rapamycin or the AMPAR inhibitor 2,3-dihydroxy-6-nitro- 7-sulfamoyl-benzo(f)quinoxaline (NBQX) pretreatment. Moreover, NBQX pretreatment eliminated the ability of sarcosine to stimulate the phosphorylated mTOR signaling proteins. Furthermore, GluR1 phosphorylation at its PKA site was significantly increased after an acute in vivo sarcosine treatment. The results demonstrated that sarcosine exerts antidepressant-like effects by enhancing AMPAR-mTOR signaling pathway activity and facilitating AMPAR membrane insertion. [ABSTRACT FROM AUTHOR]

Although collagenolytic matrix metalloproteinases (MMPs) possess common domain organizations, there are subtle differences in their processing of collagenous triple-helical substrates. In this study, we have incorporated peptoid residues into collagen model triple-helical peptides and examined MMP activities toward these peptomeric chimeras. Several different peptoid residues were incorporated into triple-helical substrates at subsites P3, P1, P1', and P10' individually or in combination, and the effects of the peptoid residues were evaluated on the activities of full-length MMP-1, MMP-8, MMP-13, and MMP-14/MT1-MMP. Most peptomers showed little discrimination between MMPs. However, a peptomer containing N-methyl Gly (sarcosine) in the P1' subsite and N-isobutyl Gly (NLeu) in the P10' subsite was hydrolyzed efficiently only by MMP-13 [nomenclature relative to the α1(I)772-786 sequence]. Cleavage site analysis showed hydrolysis at the Gly-Gln bond, indicating a shifted binding of the triple helix compared to the parent sequence. Favorable hydrolysis by MMP-13 was not due to sequence specificity or instability of the substrate triple helix but rather was based on the specific interactions of the P7' peptoid residue with the MMP-13 hemopexin-like domain. A fluorescence resonance energy transfer triple-helical peptomer was constructed and found to be readily processed by MMP-13, not cleaved by MMP-1 and MMP-8, and weakly hydrolyzed by MT1-MMP. The influence of the triple-helical structure containing peptoid residues on the interaction between MMP subsites and individual substrate residues may provide additional information about the mechanism of collagenolysis, the understanding of collagen specificity, and the design of selective MMP probes. [ABSTRACT FROM AUTHOR]

Cylindrical brush polymers constitutepromising polymeric drugdelivery systems (nanoDDS). Because of the densely grafted side chainssuch structures may intrinsically exhibit little protein adsorption(“stealth” effect) while providing a large number offunctional groups accessible for bioconjugation reactions. Polysarcosine(PSar) is a highly water-soluble, nonionic and nonimmunogenic polypeptoidbased on the endogenous amino acid sarcosine (N-methylglycine). Here we report on the synthesis, characterization and biocompatibilityof cylindrical brush polymers with either polysarcosine side chainsor poly-l-lysine-b-polysarcosine side chains.The latter leads to block copolypept(o)id based core–shellcylindrical brushes with a cationic poly-l-lysine (PLL) coreand a neutral polysarcosine corona. The cylindrical brush polymerswere prepared by ring-opening polymerization of the respective N-carboxyanhydrides(NCA) from a macroinitiator chain. Preliminary experiments on complexformation with siRNA demonstrate that a core–shell cylindricalbrush polymer may complex on average up to 270 RNA molecules amountingto a high loading efficiency of N+/P–= 1.1. No bridging between cylindrical brushes leading to largeraggregates is observed. In vitro studies on the silencing of the expressionof ApoB100, which is abundantly expressed in AML-12 hepatocytes, inducedby siRNA-cylindrical core–shell brush complexes showed highefficiency, leading to a knock-down efficiency of ApoB100 mRNA of70%. [ABSTRACT FROM AUTHOR]

Several studies have shown that glycine transporter 1 (GlyT1) inhibitors have anxiolytic actions. There are two types of glycine receptor: the strychnine-sensitive glycine receptor (GlyA) and the strychnine-insensitive glycine receptor (GlyB); however, which receptor is the main contributor to the anxiolytic actions of GlyT1 inhibitors is yet to be determined. Here, we clarified which glycine receptor is the main contributor to the anxiolytic effects of GlyT1 inhibitors by using maternal separation-induced ultrasonic vocalization (USV) by rat pups as an index of anxiety. We confirmed that administration of the benzodiazepine diazepam or the selective serotonin reuptake inhibitor escitaloplam, which are both clinically proven anxiolytics, or the GlyT1 inhibitor SSR504734 (2-chloro-N-[(S)-phenyl[(2S)-piperidin-2-yl] methyl]-3-trifluoromethyl benzamide), decreases USV in rat pups. In addition, we showed that another GlyT1 inhibitor, ALX5407 ((R)-N-[3-(4′-fluorophenyl)-3(4′-phenylphenoxy)propyl]sarcosine) also decreases USV in rat pups. SSR504734- or ALX5407-induced decreases in USV were dose-dependently reversed by administration of the GlyA antagonist strychnine, whereas the diazepam- or escitalopram-induced decreases in USV were not. Furthermore, GlyT1-induced decreases in USV were not reversed by administration of the GlyB antagonist L-687,414. Together, these results suggest that GlyA activation is the main contributor to the anxiolytic actions of GlyT1 inhibitors and that the anxiolytic actions of diazepam and escitalopram cannot be attributed to GlyA activation. Our findings provide new insights into the importance of the activation of GlyA in the anxiolytic effects of GlyT1 inhibitors. [ABSTRACT FROM AUTHOR]

Sarcosine was recently identified as a differential metabolite that is present in urine in elevated concentration as prostate cancer develops metastases. The aim of this study is to prepare reflectometric interference spectroscopy (RIfS) nanosensors for the detection of sarcosine. Sarcosine imprinted nanoparticles were prepared by two phase miniemulsion polymerization and characterized with transmission electron microscopy, dynamic light scattering, and atomic force microscopy measurements. The glass substrates (10 × 10 mm2) were treated with Piranha solution and incubated in polyethyleneimine solution. The sarcosine imprinted nanoparticles were attached to glass substrates by spin coating of nanoparticle solution. The recognition properties of the nanosensors were evaluated by reflectometric interferometric spectroscopy. To show selectivity of sarcosine imprinted RIfS nanosensor, real-time l-alanine detection was also performed. Sarcosine detection studies were performed from aqueous solution and urine sample. A good linearity was revealed with a correlation coefficient of 0.9622 and a detection limit of 45 nM. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:55-61, 2015 [ABSTRACT FROM AUTHOR]

Abstract: A secondary amine based ionic liquid, lithiated sarcosine-trifluoromethyl sulfonyl imide (TFSI) solution has been developed for hydrothermal synthesis of hexagonal ZnO micro rods. The ionic liquid formation was confirmed by 13C-NMR, 7Li-NMR and FTIR analysis and a possible mechanism was proposed for the formation of wurtzite hexagonal ZnO micro rods using morphological observations. The lithiated sarcosine TFSI solution has shown both capping and catalytic activities in bringing out the hexagonal ZnO micro rods. [Copyright &y& Elsevier]

Aim of Study: Endothelial cells (ECs) respond to changes in metabolic status and switch over to angiogenic phenotype. There are several metabolites known to mediate this transition; however, the effect of sarcosine that accumulates in invasive prostate cancer is not known. The objective of the study was to examine whether sarcosine influences EC function and affects angiogenesis. Materials and Methods: The effect of sarcosine was studied using different model systems including chick chorioallantoic membrane (CAM), rat aortic rings in culture, and human umbilical vein ECs (HUVECs) in culture. The statistical significance of difference was analyzed by one-way analysis of variance (ANOVA) and Student's t-test using GraphPad 5 software. Results: Increased vascularization in CAM, increased endothelial sprouting in rat aortic rings in culture, and increased expression of CD31 and E-selectin suggested a possible angiogenic effect of sarcosine. Sarcosine modulated expression of angiogenic growth factors such as vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF). In ECs in culture LY294002, an inhibitor of phosphatidylinositol-3-kinase (PI3K)/Akt pathway and rapamycin, an inhibitor of mammalian target of rapamycin (mTOR) reversed the effect of sarcosine. Further, sarcosine induced upregulation and activation of Akt in HUVECs. Conclusion: These results suggest that sarcosine modulates EC function relevant to angiogenesis through modulation of PI3K/Akt/ mTOR pathway. [ABSTRACT FROM AUTHOR]

Recently, we developed a selective and efficient method of hydrogen-deuterium exchange (HDX) at the α-carbon (α-C) of sarcosine residue (N-methylglycine) in model peptides [Bąchor et al. J. Mass Spectrom. 2014, 49, 43]. Here, we report the influence of quaternary ammonium (QA) group on HDX at the α-C of sarcosine and N-methylalanine in peptides. The obtained results suggest a significant acceleration of the HDX in sarcosine residue caused by the presence of QA. The effect depends on the distance between the sarcosine residue and QA moiety. The deuterons, introduced at α-C, are resistant to the back-exchange in acidic aqueous solution. The collision induced dissociation of the deuterium-labeled analogs of QA-tagged oligosarcosine peptides without mobile hydrogen revealed the mobilization of the hydrogens localized at α-C of sarcosine residue. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Sarcosine has been identified as a potential prostate cancer marker. To provide determination of this compound, a number of methods are developing. In this study, we optimized a method for its separation by hydrophilic interaction LC with electrochemical detection ( ED). Due to the fact that mobile phases commonly used for this type of separation altered the LODs measured by electrochemical detectors, we applied postcolumn dosing of buffer suitable for ED. The optimized conditions were mobile phase A acetonitrile, mobile phase B water in the ratio A/ B 70:30, with postcolumn addition of mobile phase C (200 mM phosphate buffer pH 9). The optimal mixing ratio was A + B/ C 1:1 with a flow rate of 0.80 mL/min (0.40 + 0.40 mL/min) and detection potential of 1000 mV. Due to the optimization of the parameters for effective separation, which had to meet the optimal parameters of ED, we reached a good resolution for separation also with a good LOD (100 nM). In addition, we successfully carried out sarcosine analysis bound on our modified paramagnetic microparticles with the ability to preconcentrate sarcosine isolated from artificial urine. [ABSTRACT FROM AUTHOR]

The effect of N-methylation on sequence scrambling in the fragmentation of b5 ions has been investigated by studying a variety of peptides containing sarcosine (N-methylglycine). The product ion mass spectra for the b5 ions derived from Sar-A-A-A-Y-A and Sar-A-A-Y-A-A show only minor signals for non-direct sequence ions the major fragmentation reactions occurring from the unrearranged structures. This is in contrast to the b5 ions where the Sar residue is replaced by Ala and sequence scrambling occurs. The b5 ion derived from Y-Sar-A-A-A-A shows a product ion mass spectrum essentially identical to the spectrum of the b5 ion derived from Sar-A-A-A-Y-A, indicating that in the former case macrocyclization has occurred but the macrocyclic form shows a strong preference to reopen to put the Sar residue in the N-terminal position. Similar results were obtained in the comparison of b5 ions derived from A-Sar-A-A-Y-A and Sar-A-A-Y-A-A. The product ion mass spectra of the MH+ ions of Y-Sar-A-A-A-A and A-Sar-A-A-Y-A show substantial signals for non-direct sequence ions indicating that fragmentation of the MH+ ions channels extensively through the respective b5 ions and further fragmentation of these species. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Sarcosine, a potential biomarker of prostate cancer, has drawn great attention in recent years. However, controversial research keeps arising about its role as a biomarker that might come from the two isomers (α-alanine and β-alanine) of sarcosine due to their same molecular weight and similar properties, which could interfere with the accurate detection of sarcosine. In this study, a simple and sensitive method was developed for the detection of sarcosine and the two isomers by LC with ion-trap MS through a novel derivatization reagent N, N′-dicyclohexylcarbodiimide. N, N′-Dicyclohexylcarbodiimide is usually considered as a condensation reagent, however, it was directly used as a derivatization reagent through a rearrangement side reaction in this study. The proposed method not only improved the chromatographic retention behavior of sarcosine and its two isomers, which was a benefit to their separation, but also dramatically enhanced the detection sensitivity of sarcosine, which was more favorable for real sample analysis. The factors affecting the productivity of the derivatization reaction, such as reaction time and amount of derivatization reagent, were systematically optimized. The method shows good linearity ( R2 > 0.99), sensitivity with LODs of sarcosine as low as 1 ng/mL, and repeatability with the RSD < 6.07%. The developed method was applied to the analysis of urine. [ABSTRACT FROM AUTHOR]

Metabolomic profiling has identified, sarcosine, a derivative of the amino acid glycine, as an important metabolite involved in the etiology or natural history of prostate cancer. We examined the association between serum sarcosine levels and risk of prostate cancer in 1122 cases (813 non-aggressive and 309 aggressive) and 1112 controls in the Prostate, Lung, Colorectal and Ovarian Cancer Screening Trial. Sarcosine was quantified using high-throughput liquid chromatography–mass spectrometry. A significantly increased risk of prostate cancer was observed with increasing levels of sarcosine (odds ratio [OR] for the highest quartile of exposure [Q4] versus the lowest quartile [Q1] = 1.30, 95% confidence interval [CI]: 1.02, 1.65; P-trend 0.03). When stratified by disease aggressiveness, we observed a stronger association for non-aggressive cases (OR for Q4 versus Q1 = 1.44, 95% CI: 1.11, 1.88; P-trend 0.006) but no association for aggressive prostate cancer (OR for Q4 versus Q1 = 1.03, 95% CI: 0.73, 1.47; P-trend 0.89). Although not statistically significant, temporal analyses showed a stronger association between sarcosine and prostate cancer for serum collected closer to diagnosis, suggesting that sarcosine may be an early biomarker of disease. Interestingly, the association between sarcosine and prostate cancer risk was stronger among men with diabetes (OR = 2.66, 95% CI: 1.04, 6.84) compared with those without reported diabetes (OR = 1.23, 95% CI: 0.95–1.59, P-interaction = 0.01). This study found that elevated levels of serum sarcosine are associated with an increased prostate cancer risk and evidence to suggest that sarcosine may be an early biomarker for this disease. [ABSTRACT FROM PUBLISHER]

Abstract: Rats treated with three daily urocortin 1 (UCN) injections into the basolateral amygdala (BLA; i.e., UCN/BLA-primed rats) develop prolonged anxiety-associated behavior and vulnerability to panic-like physiological responses (i.e., tachycardia, hypertension and tachypnea) following intravenous infusions of 0.5M sodium lactate (NaLac, an ordinarily mild interoceptive stressor). In these UCN-primed rats, the osmosensitive subfornical organ (SFO) may be a potential site that detects increases in plasma NaLac and mobilizes panic pathways since inhibiting the SFO blocks panic following NaLac in this model. Furthermore, since SFO neurons synthesize angiotensin II (A-II), we hypothesized that the SFO projects to the BLA and releases A-II to mobilizing panic responses in UCN/BLA-primed rats following NaLac infusions. To test this hypothesis, rats received daily bilateral injections of UCN or vehicle into the BLA daily for 3days. Five to seven days following the intra-BLA injections, we microinjected either the nonspecific A-II type 1 (AT1r) and 2 (AT2r) receptor antagonist saralasin, or the AT2r-selective antagonist PD123319 into the BLA prior to the NaLac challenge. The UCN/BLA-primed rats pre-injected with saralasin, but not PD123319 or vehicle, had reduced NaLac-induced anxiety-associated behavior and panic-associated tachycardia and tachypnea responses. We then confirmed the presence of AT1rs in the BLA using immunohistochemistry which, combined with the previous data, suggest that A-II's panicogenic effects in the BLA is AT1r dependent. Surprisingly, the SFO had almost no neurons that directly innervate the BLA, which suggests an indirect pathway for relaying the NaLac signal. Overall these results are the first to implicate A-II and AT1rs as putative neurotransmitter-receptors in NaLac induced panic-like responses in UCN/BLA-primed rats. [Copyright &y& Elsevier]

Misfolding of the α-helical membrane protein peripheral myelin protein 22 (PMP22) has been implicated in the pathogenesis of the common neurodegenerative disease known as Charcot-Marie-Tooth disease (CMTD) and also several other related peripheral neuropathies. Emerging evidence suggests that the propensity of PMP22 to misfold in the cell may be due to an intrinsic lack of conformational stability. Therefore, quantitative studies of the conformational equilibrium of PMP22 are needed to gain insight into the molecular basis of CMTD. In this work, we have investigated the folding and unfolding of wild type (WT) human PMP22 in mixed micelles. Both kinetic and thermodynamic measurements demonstrate that die denaturation of PMP22 by n-lauroyl sarcosine (LS) in dodecylphosphocholine (DPC) micelles is reversible. Assessment of the conformational equilibrium indicates that a significant fraction of unfolded PMP22 persists even in the absence of the denaturing detergent However, we find the stability of PMP22 is increased by glycerol, which facilitates quantitation of thermodynamic parameters. To our knowledge, this work represents the first report of reversible unfolding of a eukaryotic multispan membrane protein. The results indicate that WT PMP22 possesses minimal conformational stability in micelles, which parallels its poor folding efficiency in the endoplasmic reticulum. Folding equilibrium measurements for PMP22 in micelles may provide an approach to assess the effects of cellular metabolites or potential therapeutic agents on its stability. Furthermore, these results pave the way for future investigation of the effects of pathogenic mutations on the conformational equilibrium of PMP22. [ABSTRACT FROM AUTHOR]

Abstract: Sarcosine (N-methylglycine) is an intermediate in glycine degradation and can also be synthesised from glycine in mammals. Sarcosine metabolism in Haemonchus contortus and Teladorsagia circumcincta differed from that of mammals in that creatinase activity was present and sarcosine was demethylated only by sarcosine oxidase (SOX) and not by sarcosine dehydrogenase (SDH). The mean SOX activity was 30nmolmin−1 mg−1 protein in homogenates of L3 and adult worms of both parasites and the apparent K m for sarcosine was 1.1mM. Addition of 2mM Cd2+ inhibited activity by 30%. There was no SDH activity with either NAD+ or NADP+ as co-factor. Mean creatinase activity in L3 T. circumcincta and adult worms of both species was 31±6nmolmin−1 mg−1 protein, but was undetectable in L3 H. contortus. Activity was inhibited by up to 70% by Cu2+, Fe2+, Fe3+ and Zn2+. Possessing creatinase would allow host creatine to be incorporated into amino acids by the parasites. [Copyright &y& Elsevier]

Heterobimetallic complexes of Zn(II) and Sn(IV) with sarcosine have been synthesized at room temperature under stirring conditions by the reaction of sarcosine and zinc acetate in 2 : 1 molar ratio followed by the stepwise addition of CS2 and organotin(IV) halides, where R =Me, n-Bu, and Ph. The complexes were characterized by elemental analysis, FT-IR and NMR (¹H, 13C) spectroscopy. IR data showed that the ligand acts in a bidentate manner. NMR data revealed the four coordinate geometry in solution state. In vitro antimicrobial activities data showed that complexes (3) and (4) were effective against bacterial and fungal strains with few exceptions. [ABSTRACT FROM AUTHOR]

Sarcosinemia is an autosomal recessive metabolic trait manifested by relatively high concentrations of sarcosine in blood and urine. Sarcosine is a key intermediate in 1-carbon metabolism and under normal circumstances is converted to glycine by the enzyme sarcosine dehydrogenase. We encountered six families from two different descents (French and Arab), each with at least one individual with elevated levels of sarcosine in blood and urine. Using the 'candidate gene approach' we sequenced the gene encoding sarcosine dehydrogenase ( SARDH), which plays an important role in the conversion of sarcosine to glycine, and found four different mutations (P287L, V71F, R723X, R514X) in three patients. In an additional patient, we found a uniparental disomy in the region of SARDH gene. In two other patients, we did not find any mutations in this gene. We have shown for the first time that mutations in the SARDH gene are associated with sarcosinemia. In addition, our results indicate that other genes are most probably involved in the pathogenesis of this condition. [ABSTRACT FROM AUTHOR]

Abstract: Conventional IgG-ELISA methods for diagnosing cat scratch disease (CSD) caused by Bartonella hensela are still poor in sensitivity and specificity, which generally employ bacterial whole-cell proteins or N-lauroyl-sarcosine–insoluble proteins as the antigen. By Western blot analysis, we found that sarcosine-soluble fraction of proteins (SSP) showed highly specific reaction to immunofluorescence assay (IFA)–positive sera obtained from CSD patients compared with the above antigens. Clinical utility of the new ELISA employing SSP was evaluated using sera from 118 patients with clinically suspected CSD (sera positive by IFA: titers ≥1:256, n = 46; negative: titers <128, n = 72) and 88 sera from healthy individuals. Sensitivity and specificity of distinguishing IFA-positive patients from healthy individuals were 95.7% and 97.7%, respectively. Fifteen discordant results were observed (13 ELISA(+)/IFA(−); 2 ELISA(−)/IFA(+)). However, all 15 sera reacted with SSP by Western blot analysis, indicating superiority of the new ELISA over IFA. The ELISA employing SSP greatly improved the accuracy of diagnosing CSD. [Copyright &y& Elsevier]

SARCOSIN, also named Krp1, has been identified as a protein exclusively expressed in striated muscle tissue. Here we report on the role of SARCOSIN in skeletal muscle development and differentiation. We demonstrate, by means of whole-mount in situ hybridization, that Sarcosin mRNA is expressed in the myotome part of the mature somites in mouse embryos from embryonic day 9.5 onwards. Sarcosin is not expressed in the developing heart at these embryonic stages, and in adult tissues the mRNA expression levels are five times lower in the heart than in skeletal muscle. SARCOSIN protein partially co-localizes with the M-band protein myomesin and between and below laterally fusing myofibrils in adult skeletal muscle tissue. RNA interference mediated knock-down of SARCOSIN in the C2C12 myoblast cell line appeared to be stimulatory in the early phase of differentiation, but inhibitory at a later phase of differentiation. [ABSTRACT FROM AUTHOR]

A supramolecular approach for the specific detection of sarcosine, recently linked to the occurrence of aggressive prostate cancer forms, has been developed. A hybrid active surface was prepared by the covalent anchoring on Si substrates of a tetraphosphonate cavitand as supramolecular receptor and it was proven able to recognize sarcosine from its nonmethylated precursor, glycine, in water and urine. The entire complexation process has been investigated in the solid state, in solution, and at the solid-liquid interface to determine and weight all the factors responsible of the observed specificity. The final outcome is a Si-based active surface capable of binding exclusively sarcosine. The complete selectivity of the cavitand-decorated surface under these stringent conditions represents a critical step forward in the use of these materials for the specific detection of sarcosine and related metabolites in biological fluids. [ABSTRACT FROM AUTHOR]

recent study showed that sarcosine may be potentially useful for the diagnosis and prognosis of prostate cancer (PCa). The aim of this study was to validate diagnostic value of sarcosine for PCa, to evaluate urine metabolomic profiles in patients with PCa in comparison of non-cancerous control, and to further explore the other potential metabolic biomarkers for PCa. Isotope dilution gas chromatography/mass spectrometry (ID GC/MS) metabolomic approach was applied to evaluate sarcosine using [methyl-D]-sarcosine as an internal standard. Microwave-assisted derivatization (MAD) together with GC/MS was utilized to obtain the urinary metabolomic information in 20 PCa patients compared with eight patients with benign prostate hypertrophy and 20 healthy men. Acquired metabolomic data were analyzed using a two-sample t test. Diagnostic models for PCa were constructed using principal component analysis and were assessed with receiver-operating characteristic curves. Results showed that the urinary sarcosine level has no statistical difference between the PCa group and the control group. In addition, nine metabolomic markers between the PCa group and the healthy male group were selected, which constructed a diagnostic model with a high area under the curve value of 0.9425. We conclude that although urinary sarcosine value has limited potential in the diagnostic algorithm of PCa, urinary metabolomic panel based on GC/MS assay following MAD may potentially become a diagnostic tool for PCa. [ABSTRACT FROM AUTHOR]

The present study reports a differential scanning calorimetry (DSC) study of the amino acids sarcosine [CAS Registry No. 107-97-1], α-alanine (dl) [CAS Registry No. 302-72-7], β-alanine [CAS Registry No. 107-95-9], N-benzyl-α-alanine (dl) [CAS Registry No. 40297-69-6], and N-benzyl-β-alanine [CAS Registry No. 5426-62-0] in the temperature interval from T= 268 K to their respective melting/decomposition temperatures. Temperatures and heat capacities as a function of temperature of all solids and the enthalpies and entropies of fusion of two of the amino acids, sarcosine and N-benzyl-β-alanine, are reported. [ABSTRACT FROM AUTHOR]

While sarcosine was recently identified as a potential urine biomarker for prostate cancer, further studies have cast doubt on its utility to diagnose this condition. The inconsistent results may be due to the fact that alanine and sarcosine coelute on an HPLC reversed-phase column and the mass spectrometer cannot differentiate between the two isomers, since the same parent/product ions are generally used to measure them. In this study, we developed a high-throughput liquid chromatography-mass spectrometry (LC-MS) method that resolves sarcosine from alanine isomers, allowing its accurate quantification in human serum and urine. Assay reproducibility was determined using the coefficient of variation (CV) and intraclass correlation coefficient (ICC) in serum aliquots from 10 subjects and urine aliquots from 20 subjects across multiple analytic runs. Paired serum/urine samples from 42 subjects were used to evaluate sarcosine serum/urine correlation. Both urine and serum assays gave high sensitivity (limit of quantitation of 5 ng/mL) and reproducibility (serum assay, intra- and interassay CVs < 3% and ICCs > 99%; urine assay, intra-assay CV = 7.7% and ICC = 98.2% and interassay CV = 12.3% and ICC = 94.2%). In conclusion, this high-throughput LC-MS method is able to resolve sarcosine from a- and β-alanine and is useful for quantifying sarcosine in serum and urine samples. [ABSTRACT FROM AUTHOR]

The 1,3-dipolar cycloaddition of an azomethine ylide generated by a decarboxylative route from sarcosine and isatin to 7-arylmethylidene-3-aryl-3,4-dihydro-2 H-thiazolo[3,2- a][1,3,5]triazin-6(7H)-ones afforded novel dispiro[oxindole-pyrrolidine]-thiazolo[3,2- a][1,3,5]triazines in moderate yields. The structures of the products were determined and characterized thoroughly by NMR, MS, IR, and elemental analysis. The results of experiment indicated that this 1,3-dipolar cycloaddition proceeded with high stereoselectivity and regioselectivity. J. Heterocyclic Chem., (2011). [ABSTRACT FROM AUTHOR]

The type I transmembrane protein with epidermal growth factor and two follistatin motifs 2 (TMEFF2) is expressed in brain and prostate and overexpressed in prostate cancer, but its role in this disease is unclear. Several studies have suggested that TMEFF2 plays a role in suppressing the growth and invasive potential of human cancer cells, whereas others suggest that the shed portion of TMEFF2, which lacks the cytoplasmic region, has a growth-promoting activity. Here we show that TMEFF2 has a dual mode of action. Ectopic expression of wild-type full-length TMEFF2 inhibits soft agar colony formation, cellular invasion, and migration and increases cellular sensitivity to apoptosis. However, expression of the ectodomain portion of TMEFF2 increases cell proliferation. Using affinity chromatography and mass spectrometry, we identify sarcosine dehydrogenase (SARDH), the enzyme that converts sarcosine to glycine, as a TMEFF2-interacting protein. Co-immunoprecipitation and immunofluorescence analysis confirms the interaction of SARDH with full-length TMEFF2. The ectodomain does not bind to SARDH. Moreover, expression of the full-length TMEFF2 but not the ectodomain results in a decreased level of sarcosine in the cells. These results suggest that the tumor suppressor activity of TMEFF2 requires the cytoplasmic/transmembrane portion of the protein and correlates with its ability to bind to SARDH and to modulate the level of sarcosine. [ABSTRACT FROM AUTHOR]

Glycine N-methyltransferase (GNMT) is a major hepatic enzyme that converts S-adenosylmethionine to S-adenosylhomocysteine while generating sarcosine from glycine, hence it can regulate mediating methyl group availability in mammalian cells. GNMT is also a major hepatic folate binding protein that binds to, and, subsequently, may be inhibited by 5-methyltetrafolate. GNMT is commonly diminished in human hepatoma; yet its role in cellular folate metabolism, in tumorigenesis and antifolate therapies, is not understood completely. In the present study, we investigated the impacts of GNMT expression on cell growth, folate status, methylfolate-dependent reactions and antifolate cytotoxicity. GNMT-diminished hepatoma cell lines transfected with GNMT were cultured under folate abundance or restriction. Folate-dependent homocysteine remethylation fluxes were investigated using stable isotopic tracers and gas chromatography/mass spectrometry. Folate status was compared between wild-type (WT), GNMT transgenic (GNMTtg) and GNMT knockout (GNMTko ) mice. In the cell model, GNMT expression increased folate concentration, induced folate-dependent homocysteine remethylation, and reduced antifolate methotrexate cytotoxicity. In the mouse models, GNMTtg had increased hepatic folate significantly, whereas GNMTko had reduced folate. Liver folate levels correlated well with GNMT expressions (r = 0.53, P = 0.002); and methionine synthase expression was reduced significantly in GNMTko, demonstrating impaired methylfolate-dependent metabolism by GNMT deletion. In conclusion, we demonstrated novel findings that restoring GNMT assists methylfolate-dependent reactions and ameliorates the consequences of folate depletion. GNMT expression in vivo improves folate retention and bioavailability in the liver. Studies on how GNMT expression impacts the distribution of different folate cofactors and the regulation of specific folate dependent reactions are underway. [ABSTRACT FROM AUTHOR]

Purpose: Sarcosine in prostate cancer tissue samples was recently reported to be increased during prostate cancer progression to metastasis and suggested to be a key metabolite of cancer cell invasion and aggressiveness. We reevaluated sarcosine in prostate cancer tissue samples as a potential indicator of tumor aggressiveness, and as a predictor of recurrence-free survival. Materials and Methods: Sarcosine in matched samples of malignant and nonmalignant tissue from 92 patients with prostate cancer after radical prostatectomy was measured in the framework of a global metabolite profiling study of prostate cancer by gas chromatography/mass spectrometry. We related results to age, prostate volume, tumor stage, Gleason score, preoperative prostate specific antigen and biochemical recurrence, defined as a persistent prostate specific antigen increase of greater than 0.2 ng/ml. Nonparametric statistical tests, ROC curves and Kaplan-Meier analyses were done. Results: Median sarcosine content in tissue was about 7% higher in matched malignant vs nonmalignant samples, which was significantly. Sarcosine values were not associated with tumor stage (pT2 vs pT3), tumor grade (Gleason score less than 7 vs 7 or greater) or biochemical recurrence. The lack of metastatic tissue samples was a study limitation. Conclusions: Sarcosine in prostate cancer tissue samples cannot be considered a suitable predictor of tumor aggressiveness or biochemical recurrence. [ABSTRACT FROM AUTHOR]

The current prostate cancer (PCa) diagnosis, based on the blood prostate-specific antigen (PSA) level measurement, is not a precise science. The widely used PSA biomarker for PCa has poor sensitivity and specificity and often leads to false-negative and false-positive test results. Recently, sarcosine, proline, kynurenine, uradil, and glycerol 3-phosphate were found in higher concentrations in metastatic prostate cancer urine samples. By measuring all five of these metabolites, doctors maybe better able to diagnose prostate cancer with high accuracy. However, there is no method reported for simultaneous detection of these compounds in urine samples. In this study, a novel method was developed to separate and quantify six urinary metabolites including creatinine in urine samples by using liquid chromatography/tandem mass spectrometmy. Chromatographic separations of the analytes were carried out using a phenyl-hexyl column with 0.1% formic acid in water and acetonitrile, respectively, under a gradient program. The six metabolites were detected in the multiple reaction monitoring modes with the ESIpositive mode. The linear range of the analytes was from 0.003 to 40 μmol/L The limit of detection was from 0.05 to 4 nmoI/I, and the limit of quantification ranged from 3 to 20 nmoI/L The factors affecting the separation and quantification of the six metabolites, such as mobile-phase and MS conditions, were also investigated. The technique developed in this study is simple, fast, sensitive, and selective. It can be used for quantifying these six metabolites in urine samples for potential early cancer screening. [ABSTRACT FROM AUTHOR]

Abstract: Memory reconsolidation is a dynamic process in which a previously consolidated memory becomes labile following reactivation by a reminder. In a previous study in the crab Chasmagnathus memory model, we showed that a water-shortage episode, via angiotensin modulation during reconsolidation, could reveal a memory that otherwise remains unexpressed: weakly trained animals cannot reveal long-term memory (LTM) except when an episode of noticeable ethological meaning, water deprivation, is contingent upon reconsolidation. However, these results are at variance with two of our previous interpretations: weak training protocols do not build LTM and angiotensin II modulates the strength of the information storing process. A parsimonious hypothesis is that in Chasmagnathus angiotensins regulate LTM expression, but not LTM storage. Here, we tested three predictions of this hypothesis. First, the well-known retrograde amnesic effect of the angiotensin II antagonist saralasin is not due to interference on memory storage, but to modulation of memory expression. Second, the recovery of the LTM memory expression of the apparently amnesic retrograde effect produced by saralasin, through the water-shortage episode contingent upon reconsolidation, must be reconsolidation specific. Consequently, summation-like effects and retrieval deficits cannot explain these results because of the parametric conditions of reconsolidation. Third, weak training protocols build an unexpressed LTM that requires mRNA transcription and translation, a diagnostic characteristic of LTM. Results show that angiotensin modulates LTM expression but not LTM memory storage in the crab Chasmagnathus. The results lead us to suggest that, in Chasmagnathus, LTM expression – the process of gaining appreciable control over behavior of the reactivated trace in the retrieval session – may be considered a distinct attribute of its long-term storage. This strategy, a positive modulation during reconsolidation, is proposed to distinguish between memories that can be reactivated, labilized and are not expressed, and memories that are not stored long term, obliterated or altered in other retrieval mechanisms. [Copyright &y& Elsevier]

We characterized the crystal structures of heterotetrameric sarcosine oxidase (SO) from Corynebacterium sp. U-96 complexed with methylthioacetate (MTA), pyrrole 2-carboxylate (PCA) and sulphite, and of sarcosine-reduced SO. SO comprises α-, β-, γ- and δ-subunits; FAD and FMN cofactors; and a large internal cavity. MTA and PCA are sandwiched between the re-face of the FAD isoalloxazine ring and the β-subunit C-terminal residues. Reduction of flavin cofactors shifts the β-subunit Ala1 towards the α-subunit Met55, forming a surface cavity at the oxygen-channel vestibule and rendering the β-subunit C-terminal residues mobile. We identified three channels connecting the cavity and the enzyme surface. Two of them exist in the inter-subunit space between α and β-subunits, and the substrate sarcosine seems to enter the active site through either of these channels and reaches the re-side of the FAD isoalloxazine ring by traversing the mobile β-subunit C-terminal residues. The third channel goes through the α-subunit and has a folinic acid-binding site, where the iminium intermediate is converted to Gly and either formaldehyde or, 5,10-methylenetetrahydrofolate. Oxygen molecules are probably located on the surface cavity and diffuse to the FMN isoalloxazine ring; the H2O2 formed exits via the oxygen channel. [ABSTRACT FROM PUBLISHER]

Abstract: Background: Sarcosine in urine was recently suggested to be a promising tool in prostate cancer (PCa) diagnostics. Objective: To reevaluate sarcosine as a potential biomarker for early PCa detection and for prediction of tumour aggressiveness. Design, setting, and participants: Sarcosine was measured in urine samples from 106 PCa patients and 33 patients with no evidence of malignancy (NEM), confirmed by 8–12 core prostate biopsies, after standardised digital rectal examination, as well as from 12 healthy men and women. The results were related to the clinicopathologic data on prostate volume, tumour stage, Gleason score, and prostate specific antigen (PSA). Measurements: Sarcosine in urine was determined by gas chromatography-mass spectrometry using a commercial amino acid assay and was normalised to urine creatinine. Nonparametric statistical tests and receiver operating characteristics (ROC) analyses were performed to assess the diagnostic performance. Results and limitations: The median sarcosine-creatinine ratio in urine was 13% lower in PCa than in NEM patients. Sarcosine values were not associated with tumour stage (pT2 vs pT3) or grade (Gleason score <7 vs ≥7). ROC analyses proved that the discrimination between PCa and NEM patients was not improved by sarcosine in comparison with total PSA, but it was significantly worse than the percent free PSA. The higher proportion of PCa than NEM patients can be considered a limitation of this study. Conclusions: Sarcosine in urine after rectal digital examination cannot be considered as a suitable marker to differentiate between patients with and without PCa. [Copyright &y& Elsevier]

Mice lacking metabotropic glutamate receptors 5 (mGluR5) exhibit reduced glutamatergic function and behavioral abnormalities, including deficits in prepulse inhibition (PPI) of the startle response that may be relevant to schizophrenia. Thus, these mice are an animal model that may be used for preclinical evaluation of potentially new classes of antipsychotic compounds. Recent clinical studies have suggested several compounds that modulate glutamatergic transmission through distinct mechanisms, such as potentiation of the N-methyl- d-aspartate (NMDA) receptor glycine site, activation of group II mGluR, and activation of glutamate-cysteine antiporters, as being efficacious in the treatment of schizophrenia. The aim of this work is to evaluate the effects of sarcosine (a selective inhibitor of the glycine transporter 1 [GlyT1]), LY379268 (a group II mGluR agonist), and N-acetylcysteine (a cysteine prodrug that indirectly activates cystine-glutamate antiporters to increase glutamate levels in the extrasynaptic space) on PPI deficits in mGluR5 knockout mice. Sarcosine and N-acetylcysteine, but not LY379268, ameliorated PPI deficits in mGluR5 knockout mice. The ability of N-acetylcysteine to restore PPI deficits was not blocked by the group II mGluR antagonist LY341495, indicating that the effects of N-acetylcysteine were not attributable to activation of group II mGluRs by glutamate. These findings provide evidence that the interactions between mGluR5 and NMDA receptors are involved in the regulation of PPI and suggest that activation of glutamate receptors, other than group II receptors, by increased endogenous glutamate transmission, may ameliorate the behavioral abnormalities associated with mGluR5 deficiency. [ABSTRACT FROM AUTHOR]

Recent evidence indicates that enhancing N-methyl-D-aspartate (NMDA) neurotransmission with the treatment of NMDA/glycine site agonists, such as D-serine, or a glycine transporter-1 (GlyT-1) antagonist, N-methylglycine (sarcosine), can improve symptoms of schizophrenia. To compare these two novel approaches, 60 patients with chronic schizophrenia were enrolled into a 6-wk double-blind, placebocontrolled trial of add-on treatments at the reported effective dosages (2 g/d). Clinical assessments were conducted every other week. Treatment grouprtreatment duration interaction analysis by multiple linear regression showed that sarcosine was superior to placebo at all four outcome measures of Positive and Negative Syndrome Scale (PANSS) total (p=0.005), Scale for the Assessment of Negative Symptoms (SANS) (p=0.021), Quality of Life (QOL) (p=0.025), and Global Assessment of Functioning (GAF) (p=0.042). However, D-serine did not differ significantly from placebo in any measure. Sarcosine treatment was better than D-serine in effect sizes for all outcome measures. Sarcosine also surpassed placebo in most of the measures of five PANSS factors and five SANS subscales. All treatments were well tolerated. These findings suggest that the GlyT-1 inhibitor is more efficacious than the NMDA/glycine site agonist in treatment for schizophrenia, including life quality and global function, at the dosages tested. [ABSTRACT FROM AUTHOR]