Plasma Methionine Levels Research Articles

The Association of the Essential Amino Acids Lysine, Methionine, and Threonine with Clinical Outcomes in Patients at Nutritional Risk: Secondary Analysis of a Randomized Clinical Trial.

Lysine, methionine, and threonine are essential amino acids with vital functions for muscle and connective tissue health, metabolic balance, and the immune system. During illness, the demand for these amino acids typically increases, which puts patients at risk for deficiencies with harmful clinical consequences. In a secondary analysis of the Effect of Early Nutritional Support on Frailty, Functional Outcomes, and Recovery of Malnourished Medical Inpatients Trial (EFFORT), which compared individualized nutritional support to usual care nutrition in patients at nutritional risk, we investigated the prognostic impact of the lysine, methionine, and threonine metabolism. We had complete clinical and amino acid data in 237 patients, 58 of whom reached the primary endpoint of death at 30 days. In a model adjusted for comorbidities, sex, nutritional risk, and trial intervention, low plasma methionine levels were associated with 30-day mortality (adjusted HR 1.98 [95% CI 1.16 to 3.36], p = 0.01) and with a decline in functional status (adjusted OR 2.06 [95% CI 1.06 to 4.01], p = 0.03). The results for lysine and threonine did not show statistically significant differences regarding clinical outcomes. These findings suggest that low levels of methionine may be critical during hospitalization among patients at nutritional risk. Further studies should investigate the effect of supplementation of methionine in this patient group to improve outcomes.

A panel of four plasma amino acids is a promising biomarker for newly diagnosed bladder cancer

BackgroundMetastasis and recurrence are the main causes of death in post-operative bladder cancer (BC), emphasizing the importance of exploring early-stage diagnostic markers. Serum biomarkers constitute a promising diagnostic approach for asymptomatic stage cancer as they are non-invasive, have high accuracy and low cost. AimsTo correlate concentrations of plasma amino acids with BC progression to assess their utility as an early-stage diagnostic. MethodsNewly diagnosed BC patients (n=95) and normal controls (n=96) were recruited during the period from 1 December 2018 to 30 December 2020. General and food frequency questionnaires established their basic information and dietary intake data. Venous blood samples were collected from fasting subjects and used to detect levels of plasma amino acids by liquid chromatography-mass spectrometry. Verification was performed on the GSE13507 transcriptome gene expression matrix of BC from Gene Expression Omnibus (GEO) database. ResultsEleven amino acids have been identified as altered in the plasma of newly diagnosed BC patients compared to controls (P < 0.05). Adjusted by gender, education, smoking and other factors, plasma ornithine level (OR = 0.256, 95% CI: 0.104 ∼ 0.630) is a protective factor for BC, plasma levels of methionine (OR = 3.460, 95% CI: 1.384 ∼ 8.651), arginine (OR = 3.851, 95% CI: 1.542 ∼ 9.616), and glutamate (OR = 3.813, 95% CI: 1.543 ∼ 9.419) are all risk factors for BC. ROC analysis demonstrated that the combination of plasma ornithine, methionine, arginine and glutamate could accurately diagnose BC (AUC = 0.84, 95% CI: 0.747 ∼ 0.833). In addition, the mRNA level of arginase 1 was decreased (P < 0.05), while the inducible nitric oxide synthase was increased significantly, which may be linked with the disturbance of arginine metabolism in BC patients. Further analysis of GEO database confirmed the role of arginine metabolism. ConclusionA biomarker panel containing four amino acids may provide a feasible strategy for the early diagnosis of BC. However, further validation is required through prospective studies.

Supplementing dietary betaine during late gestation increases plasma betaine and methionine concentrations in pregnant Merino ewes but not neonatal lambs

Supplementing dietary betaine during pregnancy has improved vigour and post-natal survival rates to day seven in twin lambs, potentially due to improvements in the circulating metabolite profile of these animals. This study determined whether maternal betaine supplementation during late gestation would increase circulating levels of betaine, creatinine and other amino acids in ewes and their newborn lambs, leading to subsequent improvements in lamb viability and growth. Twenty-five single-bearing Merino ewes received dietary betaine at either 0 (Control, CTL), 2 (Low Betaine, LB) or 4 g/day (High Betaine, HB) from day 125 of gestation (dG 125) until parturition (150.7 ± 0.6 days). Blood samples were collected from the ewes prior to feeding (0 h), and at 2, 4, 6, 8 and 24 h post-feeding on dG 125 and dG 140 prior to feeding and from the lamb within 2 h after birth. Lamb viability measures were taken at 2, 24 and 72 h and day 7 post-partum, marking (⁓30 days of age) and weaning (⁓104 days of age). Betaine supplementation at LB and HB increased plasma betaine levels at 6, 8 and 24 h post-feeding on dG 125 (P < 0.05) and tended to increase plasma betaine on dG 140 (P = 0.085). Plasma methionine levels increased at 2 and 4 h post-feeding on dG 125 relative to baseline levels (0 h) in LB ewes, yet remained unchanged in HB ewes. On dG 125, creatinine levels were higher in CTL ewes compared with betaine-supplemented ewes at 4 and 8 h post-feeding (both P < 0.05). While plasma betaine and methionine levels were unchanged in neonatal lambs, plasma concentrations of glutamine and lysine were higher (P < 0.05) in HB lambs compared to CTL and LB lambs. HB lambs tended to be heavier at marking (P = 0.057) and were heavier at weaning compared to CTL and LB lambs (P < 0.05). This study indicated that betaine supplementation increased plasma betaine and methionine levels in ewes, but not in neonatal lambs. Further research is warranted to determine whether higher levels of betaine supplementation increase neonatal concentrations.

Dietary cystine restriction increases the proliferative capacity of the small intestine of mice.

Currently, over 88 million people are estimated to have adopted a vegan or vegetarian diet. Cysteine is a semi-essential amino acid, which availability is largely dependent on dietary intake of meat, eggs and whole grains. Vegan/vegetarian diets are therefore inherently low in cysteine. Sufficient uptake of cysteine is crucial, as it serves as substrate for protein synthesis and can be converted to taurine and glutathione. We found earlier that intermolecular cystine bridges are essential for the barrier function of the intestinal mucus layer. Therefore, we now investigate the effect of low dietary cystine on the intestine. Mice (8/group) received a high fat diet with a normal or low cystine concentration for 2 weeks. We observed no changes in plasma methionine, cysteine, taurine or glutathione levels or bile acid conjugation after 2 weeks of low cystine feeding. In the colon, dietary cystine restriction results in an increase in goblet cell numbers, and a borderline significant increase mucus layer thickness. Gut microbiome composition and expression of stem cell markers did not change on the low cystine diet. Remarkably, stem cell markers, as well as the proliferation marker Ki67, were increased upon cystine restriction in the small intestine. In line with this, gene set enrichment analysis indicated enrichment of Wnt signaling in the small intestine of mice on the low cystine diet, indicative of increased epithelial proliferation. In conclusion, 2 weeks of cystine restriction did not result in apparent systemic effects, but the low cystine diet increased the proliferative capacity specifically of the small intestine and induced the number of goblet cells in the colon.

Safety and Feasibility of the Addition of a Radiosensitizing Methionine-Restricted Diet to Radiation Therapy

Methionine is an amino acid necessary for numerous processes critical for cell growth and survival. While normal cells can tolerate methionine deficiency, most cancer cells are methionine auxotrophs, requiring dietary intake since they cannot synthesize it. In vitro, methionine deficiency causes cancer cells to undergo cell cycle arrest and cell death, and in vivo a methionine restricted diet (MRD) enhances radiosensitization without significant adverse effects. Combining a MRD and radiation therapy (RT) to treat human malignancies has never been evaluated. The hypothesis of this Phase I study was that a MRD would be safe, and feasible to administer concurrently with curative-intent RT. Eligible patients included adults with any non-skin cancer malignancy undergoing standard radiation therapy without concurrent cytotoxic chemotherapy. The MRD consisted of low-protein cereals, grains, and breads; fruits; vegetables; margarines and oils; and simple carbohydrates. A clinical dietician developed a personalized meal plan with each subject to reduce methionine consumption to 5-10 mg/kg body weight/day, while maintaining adequate protein and caloric intake. An unlimited supply of a commercially available methionine-free protein supplement was provided to minimize hunger and weight loss. The MRD extended from 2 weeks before initiation of RT, through 2 weeks beyond completion of RT. The primary endpoint for safety was the rate of grade 3 or higher acute and late toxicities per CTCAE, over 12 months follow-up. Feasibility was assessed with a biweekly quantitative plasma amino acid panel during the MRD. The target accrual was 15 subjects. Over two years, 53 patients were offered enrollment, 9 subjects enrolled, 5 completed the MRD and RT, and 4 withdrew during the MRD. The table summarizes subjects' characteristics and outcomes. There was no grade 3 or higher adverse events attributable to the MRD. Methionine plasma levels varied over the course of treatment, and while no subject achieved the target of 13 μM, two nadired at 14 μM. The trial was closed early due to slow accrual and subjects' difficulty maintaining the diet. This study suggests a MRD is safe with thoracic or abdominopelvic RT, with toxicities comparable to those expected with RT alone. However, the diet was challenging, and unacceptable to most patients with cancer.

Effects of dietary methionine supplementation from different sources on growth performance and meat quality of barrows and gilts

Methionine is indispensable for growth and meat formation in pigs. However, it is still unclear that increasing dietary sulphur-containing amino acid (SAA) levels using different methionine sources affects the growth performance and meat quality of barrows and gilts. To investigate this, 144 pigs (half barrows and half gilts) were fed the control (100% SAA, CON), DL-Methionine (125% SAA, DL-Met)-supplemented, or OH-Methionine (125% SAA, OH-Met)-supplemented diets during the 11–110 kg period. The results showed that plasma methionine levels varied among treatments during the experimental phase, with increased plasma methionine levels observed following increased SAA consumption during the 25–45 kg period. In contrast, pigs fed the DL-Met diet had lower plasma methionine levels than those fed the CON diet (95–110 kg). Additionally, gilts fed the DL-Met or OH-Met diets showed decreased drip loss in longissimus lumborum muscle (LM) compared to CON-fed gilts. OH-Met-fed gilts had higher pH45min values than those fed the CON or DL-Met diets, whereas OH-Met-fed barrows had higher L45min values than those fed the CON or DL-Met diets. Moreover, increased consumption of SAA, regardless of the methionine source, tended to decrease the shear force of the LM in pigs. In conclusion, this study indicates that increasing dietary levels of SAA (+25%) appeared to improve the meat quality of gilts by decreasing drip loss and increasing meat tenderness.

Oligosaccharides and diet-related dilated cardiomyopathy in beagles.

In 2018 the US Food and Drug Administration reported a potential link between grain-free, legume-containing dog foods and the development of canine dilated cardiomyopathy in atypical breeds. One hypothesis was that high oligosaccharide content in legumes reduced bioavailability of taurine, an amino acid with some previous links to canine dilated cardiomyopathy. To address this hypothesis, in the present study, 8 Beagle dogs consumed four diets: a husbandry commercial dental diet, and three test diets formulated with either 30% rice (control), 30% pea (grain-free) or 30% rice with the addition of 1% raffinose (the predominant oligosaccharide found in peas). The study was conducted in a randomized, crossover design with 5 week feeding periods. Measurement of basic health parameters (weight, body condition score, complete blood cell count, chemistry panel), plasma amino acids, cardiac biomarkers (plasma N-terminal pro-brain natriuretic peptide (NT-proBNP) and cardiac-specific troponin I), fecal bile acids and echocardiographic exams were completed pre-study after feeding the husbandry diet as well as after each test feeding period. Echocardiography showed 9-11% reduction in ejection fraction and 17- 20% greater left ventricular end systolic volume with the husbandry diet compared to both grain-containing test diets. Concentrations of plasma NT-proBNP were 1.3-2 times greater after the husbandry diet compared to the grain-based diet, with the oligosaccharide and pea-based diets showing intermediate levels. Plasma taurine levels were unchanged across diets, while plasma methionine levels were highest and cysteine/cystine levels were lowest after dogs ate the husbandry diet. Results indicate that raffinose in the diet is sufficient, but not required to see an increase NT-proBNP, but did not induce any changes in cardiac function after 5 weeks of feeding. Whether this could progress to reduction in cardiac function with longer term feeding is uncertain. A reduced cardiac function along with the greatest increase in NT-proBNP was observed after feeding the husbandry diet that contained the highest amount of insoluble fiber but did not contain legumes or oligosaccharide. Further research into the impact of insoluble fiber in the dental diet is needed to support these novel observations.

Evaluation of the clinical, biochemical, and genetic presentation of neonatal and adult-onset 5,10-methylene tetrahydrofolate reductase (MTHFR) deficiency in patients from Pakistan.

To study the biochemical, clinical and molecular characteristics of 5,10- methylenetetrahydrofolate reductase (MTHFR) deficiency in Pakistani patients from a single center. Medical charts, urine organic acid chromatograms, plasma methionine and Hcys levels, and molecular testing results of MTHFR gene of patients presenting at the Biochemical Genetics Clinic, AKUH from 2016 to 2022 were reviewed. Neonatal MTHFR deficiency was found in five patients. The median (IQR) age of symptom onset and diagnosis were 18 (8.5-22) and 26 (16.5-31) days. The median lag between symptom onset and diagnosis was 8 (4.5-12.5) days. The median age of treatment initiation and duration of treatment were 26 (16.5-49) and 32 (25.5-54) days. The most common clinical features were lethargy, poor feeding, and seizures. The MTHFR gene sequencing revealed homozygous variants p.K510K, p.R567*, and p.R157W. Renal insufficiency manifesting as elevated serum creatinine and responding to betaine therapy was noted in one patient. This has not been previously reported in neonatal MTHFR deficiency and may reflect engagement of alternate pathways of remethylation. Adult onset MTHFR deficiency was found in six patients, with a heterogeneous neurological presentation. The median lag between symptoms onset and diagnosis was 7 (3-11) years. MTHFR gene sequencing revealed homozygous variant p.A195V in five patients from one family and p.G261V in the other. Two of the five reported variants are novel that include p.R157W and p.G261V. Eleven patients of this rare disorder from a single center indicate the need for clinical awareness and appropriate biochemical evaluation to ensure optimal outcomes.

Safety and feasibility of the addition of a radiosensitizing methionine-restricted diet to radiation therapy.

e15087 Background: Methionine is an amino acid necessary for cell growth and survival. Normal cells can tolerate methionine deficiency, but most cancer cells are methionine auxotrophs, requiring dietary intake since they cannot synthesize it. In vitro, methionine deficiency causes cancer cells to undergo cell cycle arrest and cell death, and in vivo a methionine restricted diet (MRD) enhances radiosensitization without significant adverse effects. Combining a MRD and radiation therapy (RT) for human malignancies has never been evaluated. The hypothesis of this Phase I study was that a MRD would be safe, and feasible to administer concurrently with curative-intent RT. Methods: Eligible patients included adults with any non-skin cancer malignancy undergoing standard RT without concurrent cytotoxic chemotherapy. The MRD consisted of low-protein cereals, grains, and breads; fruits; vegetables; margarines and oils; and simple carbohydrates. A clinical dietician developed a personalized meal plan with each subject to reduce methionine consumption to 5-10 mg/kg body weight/day, while maintaining adequate protein and caloric intake. An unlimited supply of a commercially available methionine-free protein supplement was provided to minimize hunger and weight loss. The MRD extended from 2 weeks before initiation of RT, through 2 weeks beyond completion of RT. The primary endpoint for safety was the rate of grade 3 or higher acute and late toxicities per CTCAE, over 12 months follow-up. Feasibility was assessed with a biweekly quantitative plasma amino acid panel during the MRD. The target accrual was 15 subjects. Results: Over two years, 53 patients were offered enrollment, 9 subjects enrolled, 5 completed the MRD and RT, and 4 withdrew during the MRD. The table summarizes subjects’ characteristics and outcomes. There was no grade 3 or higher adverse events attributable to the MRD. Methionine plasma levels varied over the course of treatment, and while no subject achieved the target of 13 μM, two nadired at 14 μM. The trial was closed early due to slow accrual and subjects’ difficulty maintaining the diet. Clinical trial information: NCT03574194 .Conclusions: This study suggests a MRD is safe with thoracic or abdominopelvic RT, with toxicities comparable to those expected with RT alone. However, the diet was challenging, and unacceptable to most patients with cancer. [Table: see text]

Pharmacokinetic/pharmacodynamic model of a methionine starvation based anti-cancer drug.

A new therapeutic approach against cancer is developed by the firm Erytech. This approach is based on starved cancer cells of an amino acid essential to their growth (the L-methionine). The depletion of plasma methionine level can be induced by an enzyme, the methionine-γ-lyase. The new therapeutic formulation is a suspension of erythrocytes encapsulating the activated enzyme. Our work reproduces a preclinical trial of a new anti-cancer drug with a mathematical model and numerical simulations in order to replace animal experiments and to have a deeper insight on the underlying processes. With a combination of a pharmacokinetic/pharmacodynamic model for the enzyme, substrate, and co-factor with a hybrid model for tumor, we develop a "global model" that can be calibrated to simulate different human cancer cell lines. The hybrid model includes a system of ordinary differential equations for the intracellular concentrations, partial differential equations for the concentrations of nutrients and drugs in the extracellular matrix, and individual based model for cancer cells. This model describes cell motion, division, differentiation, and death determined by the intracellular concentrations. The models are developed on the basis of experiments in mice carried out by Erytech. Parameters of the pharmacokinetics model were determined by fitting a part of experimental data on the concentration of methionine in blood. Remaining experimental protocols effectuated by Erytech were used to validate the model. The validated PK model allowed the investigation of pharmacodynamics of cell populations. Numerical simulations with the global model show cell synchronization and proliferation arrest due to treatment similar to the available experiments. Thus, computer modeling confirms a possible effect of treatment based on the decrease of methionine concentration. The main goal of the study is the development of an integrated pharmacokinetic/pharmacodynamic model for encapsulated methioninase and of a mathematical model of tumor growth/regression in order to determine the kinetics of L-methionine depletion after co-administration of Erymet product and Pyridoxine.

Plasma Methionine and Clinical Severity in Nitrous Oxide Consumption.

In the last few years, there has been an increase in the recreational use of nitrous oxide (N2O), which can lead to neurological symptoms such as sensory or motor disorders. The literature links these symptoms to a functional inactivation of vitamin B12 by oxidation of its cobalt ion, which prevents the vitamin B12 from acting as a cofactor for methionine synthase. Thus, demyelination related to methionine deficiency could be responsible for the neurological disorders associated with N2O consumption, including the combined sclerosis of the spinal cord. We aimed to study the correlation between the plasma methionine levels and clinical severity observed in N2O users. We retrospectively collected clinical and biological data from 93 patients who chronically consumed N2O. The patients were divided into four groups based of the severity of their clinical symptoms (based on their Peripheral Neuropathy Disability (PND) score). The plasma amino acids measurement, including methionine, were performed systematically by liquid chromatography coupled with mass spectrometry. Plasma methionine is significantly correlated with the clinical severity (Spearman coefficient: −0.42; p-value < 10−5), however, the average methionine level in the four groups is within the physiological values (N: 16−23 µmol/L). There is a significant inverse correlation between plasma methionine and homocysteine (Spearman coefficient: −0.57; p-value < 10−9), which confirms the action of nitrous oxide on the methionine synthase. A decrease in plasma methionine cannot be imputed as the only mechanism involved in the pathophysiology of the neurological disorders in nitrous oxide consumption. In addition, there are few therapeutic indications for the use of methionine. Thus, we should be careful concerning the potential use of methionine in nitrous oxide consumption. As a consequence, other pathophysiological mechanisms probably need to be identified in order to find potential therapeutic targets.

Branched-Chain and Aromatic Amino Acids Related to Visceral Adipose Tissue Impact Metabolic Health Risk Markers.

Visceral (VAT) and subcutaneous adipose tissue (SAT) function as endocrine organs capable of influencing metabolic health across adiposity levels. We aimed to investigate whether metabolites associated with VAT and SAT impact metabolic health through metabolite concentrations. Analyses are based on 1790 participants from the population-based Rhineland Study. We assessed plasma levels of methionine (Met), branched-chain amino acids (BCAA), aromatic amino acids (AAA), and their metabolic downstream metabolites with liquid chromatography-mass spectrometry. VAT and SAT volumes were assessed by magnetic resonance imaging (MRI). Metabolically healthy and unhealthy phenotypes were defined using Wildman criteria. Metabolically unhealthy participants had higher concentrations of BCAA than metabolically healthy participants (P < 0.001). In metabolically unhealthy participants, VAT volumes were significantly associated with levels of L-isoleucine, L-leucine, indole-3-lactic acid, and indole-3-propionic acid (in log SD units: β = 0.16, P = 0.003; β = 0.12, P = 0.038; β = 0.11, P = 0.035 and β = -0.16, P = 0.010, respectively). Higher concentrations of certain BCAA and AAA-downstream metabolites significantly increased the odds of cardiometabolic risk markers. The relation between VAT volume and cardiometabolic risk markers was mediated by BCAA (indirect effects 3.7%-11%, P = 0.02 to < 0.0001), while the effect of VAT on systemic inflammation was mediated through higher kynurenine concentrations (indirect effect 6.4%, P < 0.0001). Larger volumes of VAT in metabolically unhealthy individuals are associated with altered concentrations of circulating BCAA and AAA-downstream metabolites, increasing the odds of cardiometabolic risk markers. This suggests that these metabolites are involved in the mechanisms that underlie the relationship of abdominal VAT with metabolic health.

Identification of three novel pathogenic mutations in cystathionine beta-synthase gene of Pakistani intellectually disabled patients.

Classical homocystinuria (HCU) is an autosomal recessive inborn error of metabolism, which is caused by the cystathionine-β-synthase (CBS: encoded by CBS) deficiency. Symptoms of untreated classical HCU patients include intellectual disability (ID), ectopia lentis and long limbs, along with elevated plasma methionine, and homocysteine. A total of 429 ID patients (age range: 1.6-23 years) were sampled from Northern areas of Punjab, Pakistan. Biochemical and genetic analyses were performed to find classical HCU disease in ID patients. Biochemically, nine patients from seven unrelated families were identified with high levels of plasma methionine and homocysteine. Targeted exonic analysis of CBS confirmed seven causative homozygous mutations; of which three were novel missense mutations (c.451G>T; p.Gly151Trp, c.975G>C; p.Lys325Asn and c.1039+1G>T splicing), and four were recurrent variants (c.451+1G>A; IVS4+1 splicing, c.770C>T; p.Thr257Met, c.808_810del GAG; p.Glu270del and c.752T>C; p.Leu251Pro). Treatment of patients was initiated without further delay with pyridoxine, folic acid, cobalamin, and betaine as well as dietary protein restriction. The immediate impact was noticed in behavioral improvement, decreased irritability, improved black hair color, and socialization. Overall, health outcomes in this disorder depend on the age and symptomatology at the time of treatment initiation. With personalized treatment and care, such patients can reach their full potential of living as healthy a life as possible. This screening study is one of the pioneering initiatives in Pakistan which would help to minimize the burden of such treatable inborn errors of metabolism in the intellectually disabled patients.

119 Awardee Talk: A Matter of the Heart: The Use of Pulses in Canine and Feline Nutrition

Abstract In the pet food industry, the popularity of plant-based protein sources has risen as consumer demand for grain-free and novel protein sources increase. However, minimal research has been conducted as regards use of pulses and their effects on acceptability and digestibility in canine and feline nutrition. In addition, market shift towards grain-free and pulse-rich diets has presented concerns about the suitability of pulses as viable protein sources. In July 2018, the FDA published a press release discussing a potential link between diets containing legumes and the development of dilated cardiomyopathy (DCM) in dogs. As such, part of our research program has focused on the 1) characterization of the macronutrient composition and protein quality of plant-based protein sources, 2) evaluation of the effects of plant-based protein sources in extruded canine and feline diets on macronutrient digestibility, and modulation of fecal microbiota and metabolites, and 3) determination of the longitudinal effects of high pulse inclusion in diets on taurine status in dogs. Pulse ingredients contained approximately 20% crude protein, and had methionine or tryptophan as the first-limiting amino acids using the precision-fed cecectomized rooster assay. When incorporated in canine and feline diets, pulses did not result in negative effects on macronutrient digestibility or fecal scores. However, shifts in fecal microbial and metabolites were observed in animals fed pulse-based in contrast with animal-based protein diets. The inclusion of 45% green lentils in extruded diets did not cause decreased plasma or whole blood taurine or methionine levels. Additionally, excess taurine was not depleted from these animals in the urine or feces. Taurine status was considered normal in these dogs, despite changes in fecal bile acid concentrations and microbiota. Although we are currently carrying out further research projects investigating potential association of pulse rich diets and canine DCM. Current research findings from our laboratory suggest the inclusion of pulses may not be the causative factor. The use of plant-based proteins as ingredients in canine and feline diets not only meet consumer demand but also provide a valuable, safe, and nutritionally adequate alternative to traditional protein sources.

Effects of rumen-protected methionine supplementation on the performance of high production dairy cows in the tropics

Increasing methionine availability in dairy cow diets during the first third of lactation may enhance their performance and health. The aim of this study was to determine the effect of supplementing rumen-protected methionine (Smartamine® M, SM) in a lactation diet with protein and energy levels calculated according to the literature. Seventy-six multiparous Holstein cows (39.1 ± 6.8 kg of milk/d and 65 ± 28 DIM) were assigned to 1 of 2 dietary treatments (38/treatment) according to a randomized complete block design with a 2-wk (covariate) and 10-wk experimental period. Treatments were a basal diet (CON; 3.77 Lys:1Met); and CON + 23 g SM (2.97 Lys:1 Met). Individual milk samples were taken every 2 weeks to determine milk composition. Blood was collected from 24 cows on d+30 d to measure plasma AA levels. Body weight and body condition score (BCS) were measured at the beginning and the end of the experiment. The SM diet promoted higher milk yield (41.7 vs. 40.1 kg/d; P = 0.03). Energy-corrected milk yield (41.0 vs. 38.0 kg/d), milk protein yield (1.30 vs. 1.18 kg/d), milk protein (3.14% vs. 2.97%) and casein (2.39% vs. 2.28%) were also different (P < 0.01) as well as milk fat yield (1.42 vs. 1.29 kg/d; P = 0.02). A trend (P = 0.06) for higher milk fat % (3.41% vs. 3.21%) was observed. Both diets resulted in similar body weight, but CON-fed cows tended (P = 0.08) to have higher BCS. Higher plasma methionine levels were determined with SM compared with CON (29.6 vs. 18.4 μM; P < 0.01), but lysine and histidine were not different. Dietary supplementation of RPM improved productive performance by increasing milk yield and milk components yields, suggesting better dietary AA utilization when Met levels are adjusted in Lys-adequate lactation diets.

High levels of blood glutamic acid and ornithine in children with intellectual disability

Objectives: Aminoacidopathies are inborn errors of metabolism (IEMs) that cause intellectual disability in children. Luckily, aminoacidopathies are potentially treatable, if diagnosed earlier in life. The focus of this study was the screening of aminoacidopathies in a cohort of patients suspected for IEMs. Methods: Blood samples from healthy (IQ > 90; n = 391) and intellectually disabled (IQ < 70; n = 409) children (suspected for IEMs) were collected from different areas of Northern Punjab, Pakistan. An analytical HPLC assay was used for the screening of plasma amino acids. Results: All the samples (n = 800) were analyzed on HPLC and forty-three out of 409 patient samples showed abnormal amino acid profiles mainly in the levels of glutamic acid, ornithine and methionine. Plasma concentration (Mean ± SD ng/mL) were significantly high in 40 patients for glutamic acid (patients: 165 ± 38 vs. controls: 57 ± 8, p < 0.00001) and ornithine (patients: 3177 ± 937 vs. controls: 1361 ± 91, p < 0.0001). Moreover, 3 patients showed abnormally high (53.3 ± 8.6 ng/mL) plasma levels of methionine. Conclusion: In conclusion, biochemical analysis of samples from such patients at the metabolites level could reveal the underlying diseases which could be confirmed through advanced biochemical and genetic analyses. Thus, treatment to some of such patients could be offered. Thus burden of intellectual disability caused by such rare metabolic diseases could be reduced from the target populations.

Adenosine kinase deficiency: Three new cases and diagnostic value of hypermethioninemia

Adenosine kinase (ADK) deficiency is characterized by liver disease, dysmorphic features, epilepsy and developmental delay. This defect disrupts the adenosine/AMP futile cycle and interferes with the upstream methionine cycle. We report the clinical, histological and biochemical courses of three ADK children carrying two new mutations and presenting with neonatal cholestasis and neurological disorders. One of them died of liver failure whereas the other two recovered from their liver damage. As the phenotype was consistent with a mitochondrial disorder, we studied liver mitochondrial respiratory chain activities in two patients and revealed a combined defect of several complexes. In addition, we retrospectively analyzed methionine plasma concentration, a hallmark of ADK deficiency, in a cohort of children and showed that methionine level in patients with ADK deficiency was strongly increased compared with patients with other liver diseases. ADK deficiency is a cause of neonatal or early infantile liver disease that may mimic primary mitochondrial disorders. In this context, an elevation of methionine plasma levels over twice the upper limit should not be considered as a nonspecific finding. ADK deficiency induced-liver dysfunction is most often transient, but could be life-threatening.

Characterization of macrophage phenotype, redox, and purinergic response upon chronic treatment with methionine and methionine sulfoxide in mice.

Hypermethioninemia is a disorder characterized by high plasma levels of methionine (Met) and its metabolites such as methionine sulfoxide (MetO). Studies have reported associated inflammatory complications, but the mechanisms involved in the pathophysiology of hypermethioninemia are still uncertain. The present study aims to evaluate the effect of chronic administration of Met and/or MetO on phenotypic characteristics of macrophages, in addition to oxidative stress, purinergic system, and inflammatory mediators in macrophages. In this study, Swiss male mice were subcutaneously injected with Met and MetO at concentrations of 0.35-1.2g/kg body weight and 0.09-0.3g/kg body weight, respectively, from the 10th-38th day post-birth, while the control group was treated with saline solution. The results revealed that Met and/or MetO induce an M1/classical activation phenotype associated with increased levels of tumor necrosis factoralphaand nitrite, and reduced arginase activity. It was also found that Met and/or MetO alter the activity of antioxidant enzymes superoxide dismutase, catalase, and glutathione peroxidase, as well as the levels of thiol and reactive oxygen species in macrophages. The chronic administration of Met and/or MetO also promotes alteration in the hydrolysis of ATP and ADP, as indicated by the increased activity of ectonucleotidases. These results demonstrate that chronic administration of Met and/or MetO promotes activated pro-inflammatory profile by inducing M1/classical macrophage polarization. Thus, the changes in redox status and purinergic system upon chronic Met and/or MetO exposure may contribute towards better understanding of the alterations consistent with hypermethioninemic patients.

Cystathionine beta synthase deficiency and brain edema associated with methionine excess under betaine supplementation: Four new cases and a review of the evidence

CBS deficient individuals undergoing betaine supplementation without sufficient dietary methionine restriction can develop severe hypermethioninemia and brain edema. Brain edema has also been observed in individuals with severe hypermethioninemia without concomitant betaine supplementation. We systematically evaluated reports from 11 published and 4 unpublished patients with CBS deficiency and from additional four cases of encephalopathy in association with elevated methionine. We conclude that, while betaine supplementation does greatly exacerbate methionine accumulation, the primary agent causing brain edema is methionine rather than betaine. Clinical signs of increased intracranial pressure have not been seen in patients with plasma methionine levels below 559 μmol/L but occurred in one patient whose levels did not knowingly exceed 972 μmol/L at the time of manifestation. While levels below 500 μmol/L can be deemed safe it appears that brain edema can develop with plasma methionine levels close to 1000 μmol/L. Patients with CBS deficiency on betaine supplementation need to be regularly monitored for concordance with their dietary plan and for plasma methionine concentrations. Recurrent methionine levels above 500 μmol/L should alert clinicians to check for clinical signs and symptoms of brain edema and review dietary methionine intake. Levels approaching 1000 μmol/L do increase the risk of complications and levels exceeding 1000 μmol/L, despite best dietetic efforts, should be acutely addressed by reducing the prescribed betaine dose.

Clinical Studies of Methionine-Restricted Diets for Cancer Patients.

Methionine (MET) restriction (MR) has been shown to arrest cancer growth and sensitizes tumors to chemotherapy. MR total parenteral nutrition (MR TPN) with a chemotherapy-containing amino acid solution ("AO-90") (lacking both MET and L-cysteine[CYS]) showed synergistic effects with 5-fluorouracil (5-FU) in tumor-bearing rats and in a Phase I clinical trial with gastrointestinal tract cancers compared to 5-FU in a MET-containing TPN. All gastric cancer patients underwent gastrectomy. Resected tumors in the AO-90 group showed significant reduction of cancer histologically, while almost no effect was seen in the control group. A Phase II clinical trial of dietary MR combined with cystemustine treatment for melanoma or glioma was carried out. Twenty-two patients (20 with metastatic melanoma and 2 with recurrent gloma) received a median of four cycles of the combination of a 1-day MR diet with cystemustine (60mg/m2) every 2weeks. This combination was well tolerated (toxicity and nutritional status). The median disease-free survival was 1.8months and the median survival was 4.6months, with two long-duration stabilizations. MET depletion in plasma was 40%. In another study, eight patients with a variety of metastatic solid tumors were enrolled in a Phase I clinical trial of a commercially available MR medical food. Participants remained on the experimental diet for an average of 17.3weeks. Plasma methionine levels fell from 21.6 to 9μm within 2weeks, a 58% decline. The only side effect was weight loss of approximately 0.5kg per week. A feasibility study combining dietary MR with a FOLFOX regimen in patients with metastatic colorectal cancer was carried out. The plasma MET concentration was reduced by dietary MR by 58% on the first day of the MR diet. Among the four patients evaluable for response, three experienced a partial response and one patient had disease stabilization. The results of the above-described clinical trials indicate the clinical potential of MR.