Epigenome-wide association study on the plasma metabolome suggests self-regulation of the glycine and serine pathway through DNA methylation.

BACKGROUND: The plasma metabolome reflects the physiological state of various biological processes and can serve as a proxy for disease risk. Plasma metabolite variation, influenced by genetic and epigenetic mechanisms, can also affect the cellular microenvironment and blood cell epigenetics. The interplay between the plasma metabolome and the blood cell epigenome remains elusive. In this study, we performed an epigenome-wide association study (EWAS) of 1183 plasma metabolites in 693 participants from the LifeLines-DEEP cohort and investigated the causal relationships in DNA methylation-metabolite associations using bidirectional Mendelian randomization and mediation analysis. RESULTS: After rigorously adjusting for potential confounders, including genetics, we identified five robust associations between two plasma metabolites (L-serine and glycine) and three CpG sites located in two independent genomic regions (cg14476101 and cg16246545 in PHGDH and cg02711608 in SLC1A5) at a false discovery rate of less than 0.05. Further analysis revealed a complex bidirectional relationship between plasma glycine/serine levels and DNA methylation. Moreover, we observed a strong mediating role of DNA methylation in the effect of glycine/serine on the expression of their metabolism/transport genes, with the proportion of the mediated effect ranging from 11.8 to 54.3%. This result was also replicated in an independent population-based cohort, the Rotterdam Study. To validate our findings, we conducted in vitro cell studies which confirmed the mediating role of DNA methylation in the regulation of PHGDH gene expression. CONCLUSIONS: Our findings reveal a potential feedback mechanism in which glycine and serine regulate gene expression through DNA methylation.

Serum dysregulation of serine and glycine metabolism as predictive biomarker for cognitive decline in frail elderly subjects.

Frailty is a common age-related clinical syndrome characterized by a decline in the function of multiple organ systems, increased vulnerability to stressors, and a huge socio-economic burden. Despite recent research efforts, the physiopathological mechanisms underlying frailty remain elusive and biomarkers able to predate its occurrence in the early stages are still lacking. Beyond its physical component, cognitive decline represents a critical domain of frailty associated with higher risk of adverse health outcomes. We measured by High-Performance Liquid Chromatography (HPLC) a pool of serum amino acids including L-glutamate, L-aspartate, glycine, and D-serine, as well as their precursors L-glutamine, L-asparagine, and L-serine in a cohort of elderly subjects encompassing the entire continuum from fitness to frailty. These amino acids are known to orchestrate excitatory and inhibitory neurotransmission, and in turn, to play a key role as intermediates of energy homeostasis and in liver, kidney, muscle, and immune system metabolism. To comprehensively assess frailty, we employed both the Edmonton Frail Scale (EFS), as a practical tool to capture the multidimensionality of frailty, and the frailty phenotype, as a measure of physical function. We found that D-serine and D-/Total serine ratio were independent predictors of EFS but not of physical frailty. Furthermore, higher levels of glycine, glycine/L-serine and D-/Total serine were associated with worse cognition and depressive symptoms in the frail group. These findings suggest that changes in peripheral glycine and serine enantiomers homeostasis may represent a novel biochemical correlate of frailty.

Clinical toxicology of acute glyphosate self-poisoning: impact of plasma concentrations of glyphosate, its metabolite and polyethoxylated tallow amine surfactants on the toxicity.

INTRODUCTION: Common major co-formulants in glyphosate-based herbicides, polyethoxylated tallow amine surfactants, are suspected of being more toxic than glyphosate, contributing to the toxicity in humans. However, limited information exists on using polyethoxylated tallow amine concentrations to predict clinical outcomes. We investigated if plasma concentrations of glyphosate, its metabolite and polyethoxylated tallow amines can predict acute kidney injury and case fatality in glyphosate poisoning. METHODS: We enrolled 151 patients with acute glyphosate poisoning between 2010 and 2013. Plasma concentrations of glyphosate, its metabolite, aminomethylphosphonic acid, and polyethoxylated tallow amines were determined in 2020 using liquid chromatography-tandem mass spectrometry. Associations between exposure and poisoning severity were assessed. RESULTS: Plasma concentrations of glyphosate and aminomethylphosphonic acid demonstrated good and moderate performances in predicting acute kidney injury (≥2), with an area under the receiver operating characteristic curve of 0.83 (95% CI 0.69-0.97) and 0.76 (95% CI 0.59-0.94), respectively. Polyethoxylated tallow amines were detected in one-fifth of symptomatic patients, including one of four fatalities and those with unsaturated tallow moieties being good indicators of acute kidney injury (area under the receiver operating characteristic curve ≥0.7). As the number of repeating ethoxylate units in tallow moieties decreased, the odds of acute kidney injury increased. Glyphosate and aminomethylphosphonic acid concentrations were excellent predictors of case fatality (area under the receiver operating characteristic curve >0.9). DISCUSSION: The 2.7% case fatality rate with 49% acute, albeit mild, acute kidney injury following glyphosate poisoning is consistent with previously published data. A population approach using model-based metrics might better explore the relationship of exposure to severity of poisoning. CONCLUSIONS: Plasma concentrations of glyphosate and its metabolite predicted the severity of clinical toxicity in glyphosate poisoning. The co-formulated polyethoxylated tallow amine surfactants were even more strongly predictive of acute kidney injury but were only detected in a minority of patients.

Association between Childhood Overweight and Altered Concentrations of Circulating Amino Acids.

(1) Background: Dysregulated serum amino acids (AA) are known to be associated with obesity and risk of Type 2 Diabetes (T2D) in adults, and recent studies support the same notion in the pubertal age. It is, however, unknown whether childhood overweight may already display alterations of circulating AA. (2) Methods: We used liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS)-targeted metabolomics to determine plasma concentrations of AA and AA-related molecules in 36 children aged 7-12 years with normal weight or overweight. Clinical and anthropometric parameters were measured. (3) Results: Overweight in children is associated with an altered AA profile, with increased branched-chain amino acids (BCAA) and decreased glycine levels, with no clinically manifested metabolic conditions. Moreover, z-BMI was positively and negatively correlated with BCAA and glycine levels, respectively, even after adjustment for age and gender. We also found a correlation between the AA profile and clinical parameters such as lipids profile and glycemia. (4) Conclusions: A pattern of low glycine, and increased BCAA is correlated to z-BMI, total cholesterol, and triglycerides in overweight but otherwise healthy children. Our data suggest that, in childhood overweight, AA disturbances may precede other clinical parameters, thus providing an early indicator for the later development of metabolic disease.

Longitudinal Analysis of One-Carbon Metabolism-Related Metabolites in Maternal and Cord Blood of Japanese Pregnant Women.

One-carbon metabolism (OCM) is a complex and interconnected network that undergoes drastic changes during pregnancy. In this study, we investigated the longitudinal distribution of OCM-related metabolites in maternal and cord blood and explored their relationships. Additionally, we conducted cross-sectional analyses to examine the interrelationships among these metabolites. This study included 146 healthy pregnant women who participated in the Chiba Study of Mother and Child Health. Maternal blood samples were collected during early pregnancy, late pregnancy, and delivery, along with cord blood samples. We analyzed 18 OCM-related metabolites in serum using stable isotope dilution liquid chromatography/tandem mass spectrometry. We found that serum S-adenosylmethionine (SAM) concentrations in maternal blood remained stable throughout pregnancy. Conversely, S-adenosylhomocysteine (SAH) concentrations increased, and the total homocysteine/total cysteine ratio significantly increased with advancing gestational age. The betaine/dimethylglycine ratio was negatively correlated with total homocysteine in maternal blood for all sampling periods, and this correlation strengthened with advances in gestational age. Most OCM-related metabolites measured in this study showed significant positive correlations between maternal blood at delivery and cord blood. These findings suggest that maternal OCM status may impact fetal development and indicate the need for comprehensive and longitudinal evaluations of OCM during pregnancy.

Creatine homeostasis and the kidney: comparison between kidney transplant recipients and healthy controls.

Creatine is a natural nitrogenous organic acid that is integral to energy metabolism and crucial for proper cell functioning. The kidneys are involved in the first step of creatine production. With kidney transplantation being the gold-standard treatment for end-stage kidney disease, kidney transplant recipients (KTR) may be at risk of impaired creatine synthesis. We aimed to compare creatine homeostasis between KTR and controls. Plasma and urine concentrations of arginine, glycine, guanidinoacetate, creatine and creatinine were measured in 553 KTR and 168 healthy controls. Creatine intake was assessed using food frequency questionnaires. Iothalamate-measured GFR data were available in subsets of 157 KTR and 167 controls. KTR and controls had comparable body weight, height and creatine intake (all P > 0.05). However, the total creatine pool was 14% lower in KTR as compared to controls (651 ± 178 vs. 753 ± 239 mmol, P < 0.001). The endogenous creatine synthesis rate was 22% lower in KTR as compared to controls (7.8 ± 3.0 vs. 10.0 ± 4.1 mmol per day, P < 0.001). Despite lower GFR, the plasma guanidinoacetate and creatine concentrations were 21% and 41% lower in KTR as compared to controls (both P < 0.001). Urinary excretion of guanidinoacetate and creatine were 66% and 59% lower in KTR as compared to controls (both P < 0.001). In KTR, but not in controls, a higher measured GFR was associated with a higher endogenous creatine synthesis rate (std. beta: 0.21, 95% CI: 0.08; 0.33; P = 0.002), as well as a higher total creatine pool (std. beta: 0.22, 95% CI: 0.11; 0.33; P < 0.001). These associations were fully mediated (93% and 95%; P < 0.001) by urinary guanidinoacetate excretion which is consistent with production of the creatine precursor guanidinoacetate as rate-limiting factor. Our findings highlight that KTR have a disturbed creatine homeostasis as compared to controls. Given the direct relationship of measured GFR with endogenous creatine synthesis rate and the total creatine pool, creatine supplementation might be beneficial in KTR with low kidney function.Trial registration ID: NCT02811835.Trial registration URL: https://clinicaltrials.gov/ct2/show/NCT02811835 .

Comparison of the precursor, amino acid oxidation, and end-product methods for the evaluation of protein turnover in senior dogs.

Stable isotope methods have been used to study protein metabolism in humans; however, there application in dogs has not been frequently explored. The present study compared the methods of precursor (13C-Leucine), end-products (15N-Glycine), and amino acid oxidation (13C-Phenylalanine) to determine the whole-body protein turnover rate in senior dogs. Six dogs (12.7 ± 2.6 years age, 13.6 ± 0.6 kg bodyweight) received a dry food diet for maintenance and were subjected to all the above-mentioned methods in succession. To establish 13C and 15N kinetics, according to different methodologies blood plasma, urine, and expired air were collected using a specifically designed mask. The volume of CO2 was determined using respirometry. The study included four methods viz. 13C-Leucine, 13C-Phenylalanine evaluated with expired air, 13C-Phenylalanine evaluated with urine, and 15N-Glycine, with six dogs (repetitions) per method. Data was subjected to variance analysis and means were compared using the Tukey test (P<0.05). In addition, the agreement between the methods was evaluated using Pearson correlation and Bland-Altman statistics. Protein synthesis (3.39 ± 0.33 g.kg-0,75. d-1), breakdown (3.26 ± 0.18 g.kg-0.75.d-1), and flux estimations were similar among the four methods of study (P>0.05). However, only 13C-Leucine and 13C-Phenylalanine (expired air) presented an elevated Pearson correlation and concordance. This suggested that caution should be applied while comparing the results with the other methodologies.

The Gly82Ser polymorphism in the receptor for advanced glycation endproducts increases the risk for coronary events in the general population.

The receptor for advanced glycation endproducts (RAGE) has pro-inflammatory and pro-atherogenic effects. Low plasma levels of soluble RAGE (sRAGE), a decoy receptor for RAGE ligands, have been associated with increased risk for major adverse coronary events (MACE) in the general population. We performed a genome-wide association study to identify genetic determinants of plasma sRAGE in 4338 individuals from the cardiovascular arm of the Malmö Diet and Cancer study (MDC-CV). Further, we explored the associations between these genetic variants, incident first-time MACE and mortality in 24,640 unrelated individuals of European ancestry from the MDC cohort. The minor alleles of four single nucleotide polymorphisms (SNPs): rs2070600, rs204993, rs116653040, and rs7306778 were independently associated with lower plasma sRAGE. The minor T (vs. C) allele of rs2070600 was associated with increased risk for MACE [HR 1.13 95% CI (1.02-1.25), P = 0.016]. Neither SNP was associated with mortality. This is the largest study to demonstrate a link between a genetic sRAGE determinant and CV risk. Only rs2070600, which enhances RAGE function by inducing a Gly82Ser polymorphism in the ligand-binding domain, was associated with MACE. The lack of associations with incident MACE for the other sRAGE-lowering SNPs suggests that this functional RAGE modification is central for the observed relationship.

Rapid determination of glyphosate and glufosinate in human blood by probe electrospray ionization tandem mass spectrometry.

In forensic science, glyphosate (GLYP) and glufosinate (GLUF), a class of non-selective broad-spectrum herbicides, have been frequently encountered in many fatal poisoning and suicide cases due to their widespread availability. Therefore, it is essential to develop an effective method for detecting these compounds. Some conventional methods, such as gas chromatography-mass spectrometry (GC-MS) or liquid chromatography-mass spectrometry (LC-MS), have been reported to detect these compounds. However, these methods are not ideal for their time-consuming and non-sensitive feature. Herein, probe electrospray ionization (PESI) tandem mass spectrometry (MS/MS), a fast and sensitive technique, was applied for the determination of GLYP and GLUF in human blood, which can obtain analytical results within 0.5 min without derivatization and chromatographic separation. After protein precipitation of blood samples, the supernatant was mixed with isopropanol and ultra-pure water (1:1 v/v). Then, 8 µL of the mixture was introduced into the plastic sample plate for PESI-MS/MS analysis. The limits of detection (LODs) of the method were 0.50 µg/mL and 0.25 µg/mL for two analytes, and the limits of quantitation (LOQs) were both 1.00 µg/mL, which are higher than the concentration of reported poisoning and fatal cases. In the linear range of 1-500 µg/mL, the regression coefficients (r2) for GLYP and GLUF were over 0.99. The matrix effects ranged from 94.8 % to 119.5 %, and the biases were below 4.3 %. The recoveries ranged between 84.8 % and 107.4 %, and the biases were below 7.6 %. Meanwhile, the method was effectively utilized to detect and quantify the blood, urine, and other samples. Consequently, the results suggest that PESI-MS/MS is a straightforward, fast, and sensitive method for detecting GLUF and GLYP in forensics. In the future, PESI-MS/MS will become an indispensable technique for polar substances in grassroots units of public security where rapid detection is essential.

The correlation between vitamin D3 and arginine metabolism levels in newborns with amino acid metabolism disorders.

This study aimed to explore the correlation between vitamin D3 and arginine (Arg) metabolism indicators in newborns with amino acid metabolism disorders. Based on clinical data, 30 newborns with amino acid metabolism diseases admitted to Shijiazhuang Fourth Hospital from June 2021 to June 2022 were selected as the disease group, and 30 healthy newborns from the same period were selected as the healthy group. After enrollment, blood samples were collected to measure the levels of Arg, Glycine (Gly), and vitamin D3 levels. The levels of Arg metabolism indicators and vitamin D3 levels in the 2 groups and the correlation between vitamin D3 levels and Arg metabolism indicators in the affected group were analyzed. The Arg level in the diseased group was higher than that in the healthy group, whereas the Gly and vitamin D3 levels were lower than those in the healthy group (P < .05). There was a significant negative correlation between vitamin D3 and Arg levels in the affected group, and a significant positive correlation with Gly levels (P < .05). Newborns with amino acid metabolism disorders have abnormally high Arg levels, significantly reduced Gly levels, and significantly decreased vitamin D3 levels. The degree of decline was closely related to the levels of indicators of Arg metabolism. Vitamin D3 supplementation can improve the Arg metabolism status of newborns with amino acid metabolism disorders.

Reduced plasma glycine concentration in healthy and chronically diseased older adults: a marker of visceral adiposity?

BACKGROUND: Previous studies have shown that a reduced plasma concentration of the amino acid glycine (Gly) is associated with intra-abdominal obesity, but the mechanism remains unclear. OBJECTIVES: This study aimed to investigate whether lower plasma Gly concentrations in older adults are independently associated with (visceral) adiposity, age, sex, presence of chronic disease, and glucose intolerance, and whether they are caused by a reduced Gly whole-body production (WBP) and/or increased Gly disposal capacity. METHODS: We studied 102 older adults (47 males/55 females, 68.5 ± standard deviation 6.4 y) without comorbidities and 125 older adults with chronic obstructive pulmonary disease (COPD) (58 males/67 females, 69.7 ± 8.6 y). We assessed body composition and visceral adipose tissue (VAT) by dual-energy x-ray absorptiometry and muscle function by dynamometry. We measured postabsorptive plasma amino acid profile and glucose, followed by pulse administration of stable isotope-labeled Gly ([2,2-2H2]), and blood sampling was performed to measure the WBP of Gly. Results are expressed as means and 95% confidence intervals (CIs). RESULTS: We found a lower plasma Gly concentration in healthy males and males with COPD than in females (Healthy: 211; 95% CI: 193,230 compared with 248; 95% CI: 225,271; COPD: 200; 95% CI: 186,215 compared with 262: 95% CI: 241, 283; P < 0.0001, respectively), with no difference between healthy and COPD groups. A negative relationship was found between unadjusted plasma Gly and VAT mass (R2: 0.16; slope: -1.7; 95% CI: -2.4, -1.2; P < 0.0021), but not with total body fat or fasting glucose. The strong association between lower plasma Gly and increased VAT mass in older adults was independent of age, sex, body weight, lean mass or body mass index, and the presence of COPD. Inclusion of these covariates increased the R2 to 0.783. We found no relation between the VAT and WBP of Gly (P = 0.35) or Gly clearance (P = 0.187) when lean mass was considered. CONCLUSIONS: Reduced plasma Gly in older adults can be considered a marker of visceral adiposity, independent of sex, age, body composition, presence of chronic disease, and whole-body Gly production or clearance. This study was registered on clinicaltrials.gov as NCT01787682, NCT02082418, NCT02157844, NCT02770092, NCT02780219, NCT03796455, and NCT04461236.

Coingestion of Collagen With Whey Protein Prevents Postexercise Decline in Plasma Glycine Availability in Recreationally Active Men.

Whey protein ingestion during recovery from exercise increases myofibrillar but not muscle connective protein synthesis rates. It has been speculated that whey protein does not provide sufficient glycine to maximize postexercise muscle connective protein synthesis rates. In the present study, we assessed the impact of coingesting different amounts of collagen with whey protein as a nutritional strategy to increase plasma glycine availability during recovery from exercise. In a randomized, double-blind, crossover design, 14 recreationally active men (age: 26 ± 5 years; body mass index: 23.8 ± 2.1 kg·m-2) ingested in total 30 g protein, provided as whey protein with 0 g (WHEY), 5 g (WC05); 10 g (WC10), and 15 g (WC15) of collagen protein immediately after a single bout of resistance exercise. Blood samples were collected frequently over 6 hr of postexercise recovery to assess postprandial plasma amino acid kinetics and availability. Protein ingestion strongly increased plasma amino acid concentrations (p < .001) with no differences in plasma total amino acid availability between treatments (p > .05). The postprandial rise in plasma leucine and essential amino acid availability was greater in WHEY compared with the WC10 and WC15 treatments (p < .05). Plasma glycine and nonessential amino acid concentrations declined following whey protein ingestion but increased following collagen coingestion (p < .05). Postprandial plasma glycine availability averaged -8.9 ± 5.8, 9.2 ± 3.7, 23.1 ± 6.5, and 39.8 ± 11.0 mmol·360 min/L in WHEY, WC05, WC10, and WC15, respectively (incremental area under curve values, p < .05). Coingestion of a small amount of collagen (5 g) with whey protein (25 g) is sufficient to prevent the decline in plasma glycine availability during recovery from lower body resistance-type exercise in recreationally active men.

Interplay Between Plasma Glycine and Branched-Chain Amino Acids Contributes to the Development of Hypertension and Coronary Heart Disease.

BACKGROUND: Higher levels of plasma glycine are linked to a reduced risk, while increased levels of total branched-chain amino acids (tBCAAs) are associated with a higher risk of essential hypertension and coronary heart disease (CHD). As these metabolic components are interconnected, analyzing the tBCAAs/glycine ratio may help to understand their interplay in the pathogenesis of cardiovascular disease. METHODS: The Cox regression approach was combined with the development of novel genetic tools for assessments of associations between plasma metabolomic data (glycine, tBCAAs, and tBCAAs/glycine ratio) from the UK Biobank and the development of hypertension and CHD. Genome-wide association study was performed on 186 523 White UK Biobank participants to identify new independent genetic instruments for the 2-sample Mendelian randomization analyses. P-gain statistic >10 identified instruments associated with tBCAAs/glycine ratio significantly stronger compared with individual amino acids. Outcomes of genome-wide association study on hypertension and CHD were derived from the UK Biobank (nonoverlapping sample), FinnGen, and CARDIoGRAMplusC4D. RESULTS: The tBCAAs/glycine ratio was prospectively associated with a higher risk of developing hypertension and CHD (hazard ratio quintile Q5 versus Q1, 1.196 [95% CI, 1.109-1.289] and 1.226 [95% CI, 1.160-1.296], respectively). Mendelian randomization analysis demonstrated that tBCAAs/glycine ratio (P-gain >10) was a risk factor for hypertension (meta-analyzed inverse-variance weighted causal estimate 0.45 log odds ratio/SD (95% CI, 0.26-0.64) and CHD (0.48 [95% CI, 0.29-0.67]) with an absolute effect significantly larger compared with the effect of glycine (-0.06 [95% CI, -0.1 to -0.03] and -0.08 [95% CI, -0.11 to -0.05], respectively) or tBCAAs (0.22 [95% CI, 0.09-0.34] and 0.12 [95% CI, 0.01-0.24], respectively). CONCLUSIONS: The total BCAAs/glycine ratio is a key element of the metabolic signature contributing to hypertension and CHD, which may reflect biological pathways shared by glycine and tBCAAs.

Simultaneous determination of diquat, paraquat, glufosinate, and glyphosate in plasma by liquid chromatography/tandem mass spectrometry: from method development to clinical application.

Diquat (DQ), paraquat (PQ), glufosinate (GLU), and glyphosate (GLYP) are commonly used herbicides that have been confirmed to be toxic to humans. Rapid and accurate measurements of these toxicants in clinical practice are beneficial for the correct diagnosis and timely treatment of herbicide-poisoned patients. The present study aimed to establish an efficient, convenient, and reliable method to achieve the simultaneous quantification of DQ, PQ, GLU, and GLYP in human plasma using liquid chromatography-tandem mass spectrometry (LC-MS/MS) without using derivatization or ion-pairing reagents. DQ, PQ, GLU, and GLYP were extracted by the rapid protein precipitation and liquid-liquid extraction method and then separated and detected by LC-MS/MS. Subsequently, linearity, limit of detection (LOD), limit of quantification (LOQ), precision, accuracy, extraction recovery, matrix effect, dilution integrity, and stability were evaluated to validate the method based on the FDA criteria. Finally, the validated method was applied to real plasma samples collected from 166 Chinese patients with herbicide poisoning. The results showed satisfactory linearity with low LOD (1 ng/mL for DQ and PQ, 5 ng/mL for GLU, and 10 ng/mL for GLYP, respectively) and low LOQ (5 ng/mL for DQ and PQ, 25 ng/mL for GLU and GLYP, respectively). In addition, the precision, accuracy, extraction recovery, and stability of the method were acceptable. The matrix effect was not observed in the analyzed samples. Moreover, the developed method was successfully applied to determine the target compounds in real plasma samples. These data provided reliable evidence for the application of this LC-MS/MS method for clinical poisoning detection.

Glycine supplementation can partially restore oxidative stress-associated glutathione deficiency in ageing cats.

Intracellular levels of glutathione, the major mammalian antioxidant, are reported to decline with age in several species. To understand whether ageing affects circulating glutathione levels in cats, blood was sampled from two age groups, < 3 years and > 9 years. Further, to determine whether dietary supplementation with glutathione precursor glycine (GLY) affects glutathione concentrations in senior cats (> 8 years), a series of free GLY inclusion level dry diets were fed. Subsequently, a 16-week GLY feeding study was conducted in senior cats (> 7 years), measuring glutathione, and markers of oxidative stress. Whole blood and erythrocyte total, oxidised and reduced glutathione levels were significantly decreased in senior cats, compared with their younger counterparts (P ≤ 0·02). The inclusion level study identified 1·5 % free GLY for the subsequent dry diet feeding study. Significant increases in erythrocyte total and reduced glutathione were observed between senior cats fed supplemented and control diets at 4 weeks (P ≤ 0·03; maximum difference of 1·23 µM). Oxidative stress markers were also significantly different between groups at 8 (P = 0·004; difference of 0·68 nG/ml in 8-hydroxy-2'-deoxyguanosine) and 12 weeks (P ≤ 0·049; maximum difference of 0·62 nG/mG Cr in F2-isoprostane PGF2α). Senior cats have lower circulating glutathione levels compared with younger cats. Feeding senior cats a complete and balanced dry diet supplemented with 1·5 % free GLY for 12 weeks elevated initial erythrocyte glutathione and altered markers of oxidative stress. Dietary supplementation with free GLY provides a potential opportunity to restore age-associated reduction in glutathione in cats.

Relationship between indirect genetic effects for growth, environmental enrichment, coping style and sex with the serum metabolome profile of pigs.

Including Indirect Genetic Effects (IGE) in breeding programs to reduce aggression in group housed animals has been proposed. However, the effect of selection for IGE for growth on animal metabolism and physiology is unknown. The purpose of this study was twofold: (1) To investigate the effects of this new breeding method along with two housing (barren and straw), coping style (high and low resisters) and sex (female and castrated males) options on the metabolome profile of pigs. (2) To identify and map biological processes associated with a regrouping test at 9 weeks of age. We used Nuclear Magnetic Resonance to quantify 49 serum metabolites at week 8, 9 and 22. Also, we quantified 3 catecholamines (tyramine, epinephrine, phenylethylamine) and serotonin and three water soluble vitamins (B2, B5 and B7). Overall, no significant differences were observed between negative and positive IGE animals. The magnitude of change (delta) of many metabolites as a response to the regrouping test was significantly affected by IGE, especially that of the amino acids (P < 0.05), being greater in positive IGE pigs. The regrouping test was associated with alteration in glycine, serine and threonine metabolism. In conclusion positive and negative IGE animals respond differently to the regrouping test.

SIT1 transporter as a potential novel target in treatment of COVID-19.

Studies published earlier this year demonstrated the association of the solute carrier SLC6A20 gene with the risk and severity of COVID-19. The SLC6A20 protein product (Sodium-dependent Imino Transporter 1 (SIT1)) is involved in the transport of amino acids, including glycine. Here we summarized the results of recent studies demonstrating the interaction of SIT1 with the ACE2 receptor for SARS-CoV-2 as well as an observed association of SLC6A20 with the risk and traits of Type 2 diabetes (T2D). Recently, it was also proposed that SLC6A20 represents the novel regulator of glycine levels and that glycine has beneficial effects against the proinflammatory cytokine secretion induced by SARS-CoV-2 infection. Ivermectin, as a partial agonist of glycine-gated chloride channels, was also recently suggested to interfere with the COVID-19 cytokine storm by inducing the activation of glycine receptors. Furthermore, plasma glycine levels are found to be decreased in diabetic patients. Thus, further clinical trials are warranted to confirm the potential favorable effects of targeting the SIT1 transporter and glycine levels in the treatment of COVID-19, particularly for the severe case of disease associated with hyperglycemia, inflammation, and T2D. These findings suggest that SIT1 may potentially represent one of the missing pieces in the complex puzzle observed between these two pandemic diseases and the potential novel target for their efficient treatment.

Determination of the segmental hepatic clearance rate of 99mTc-mebrofenin and its application in the functional assessment of future liver remnant after liver resection.

99mTc-mebrofenin hepatobiliary scintigraphy with SPECT/CT (HBS-M) has become an important quantitative method to evaluate global liver function and future liver remnant (FLR) function in patients who are candidates for resective liver surgery. The purpose of this work was to describe the method in the prediction of post-surgical liver failure. The overall liver function and that of the FLR are obtained by analysis of the initial dynamic phase of the scan. Liver volume to be preserved is expressed as a percentage of the total liver volume measured in both CT sections. HBS-M is able to accurately gauge regional liver function abnormalities that could be represented as normal liver tissue parenchyma in the CT study. This technique can provide very valuable prognostic information for the estimation of the postoperative risk of liver failure in all patients who are candidates for resective liver surgery.

Metabolomic Detection Between Pancreatic Cancer and Liver Metastasis Nude Mouse Models Constructed by Using the PANC1-KAI1/CD82 Cell Line.

Background: Pancreatic cancer (PC) has a poor prognosis and is prone to liver metastasis. The KAI1/CD82 gene inhibits PC metastasis. This study aimed to explore differential metabolites and enrich the pathways in serum samples between PC and liver metastasis nude mouse models stably expressing KAI1/CD82. Methods: KAI1/CD82-PLV-EF1α-MCS-IRES-Puro vector and PANC1 cell line stably expressing KAI1/CD82 were constructed for the first time. This cell line was used to construct 3 PC nude mouse models and 3 liver metastasis nude mouse models. The different metabolites and Kyoto encyclopedia of genes and genomes (KEGG) and human metabolome database (HMDB) enrichment pathways were analyzed using the serum samples of the 2 groups of nude mouse models on the basis of untargeted ultra-performance liquid chromatography-tandem mass spectrometry platform. Results: KAI1/CD82-PLV-EF1α-MCS-IRES-Puro vector and PANC1 cell line stably expressing KAI1/CD82 were constructed successfully, and all nude mouse models survived and developed cancers. Among the 1233 metabolites detected, 18 metabolites (9 upregulated and 9 downregulated) showed differences. In agreement with the literature data, the most significant differences between both groups were found in the levels of bile acids (taurocholic acid, chenodeoxycholic acid), glycine, prostaglandin E2, vitamin D, guanosine monophosphate, and inosine. Bile recreation, primary bile acid biosynthesis, and purine metabolism KEGG pathways and a series of HMDB pathways (P < .05) contained differential metabolites that may be associated with liver metastasis from PC. However, the importance of these metabolites on PC liver metastases remains to be elucidated. Conclusions: Our findings suggested that the metabolomic approach may be a useful method to detect potential biomarkers in PC.

Association of Therapeutic Dose of Valproic Acid and Plasma Glycine Levels in Epileptic Patients.

OBJECTIVES: To determine the frequency of hyperglycinemia in epileptic patients taking valproic acid (VPA); and the correlation between therapeutic dose of valproic acid and plasma glycine levels in epileptic patients. STUDY DESIGN: Observational, cross-sectional study. PLACE AND DURATION OF STUDY: Department of Chemical Pathology and Endocrinology, Armed Forces Institute of Pathology Rawalpindi, in collaboration with Combined Military Hospital, Rawalpindi, from August 2020 to January 2021. METHODOLOGY: Plasma glycine levels were analysed on ion exchange chromatography (IEC)-based instrument, Biochrome 30+ of epileptic patients undergoing treatment with anti-epileptic agents. Therapeutic doses of valproic acid were taken as serum trough levels of valproic acid and analysed on chemiluminescence-based Abbott Architect Plus i1000 SR. Mann-Whitney U-test was applied to compare plasma glycine levels in epileptic patients on valproic acid and those on multiple anti-epileptic agents. Spearman's correlation was used to correlate plasma glycine levels in epileptic patients with trough levels of valproic acid, duration of treatment and frequency of fits/year. RESULTS: A total of 77 participants, upto 15 years of age, were enrolled. Plasma glycine levels were significantly raised (p <0.001) in those epileptics who were on valproic acid (monodrug therapy), in comparison with those on multiple anti-epileptic agents. There were significant positive correlations between glycine levels and trough valproic acid levels (r = 0.830), duration of treatment (r = 0.525) and frequency of seizures (r = 0.326). CONCLUSION: Epileptic patients treated with valproic acid (VPA) had raised plasma glycine levels, that increased with therapeutic dose of valproic acid and duration of treatment and was associated with increased frequency of fits in those patients. Key Words: Epilepsy, Seizure, Glycine, Valproic acid.