Effect of supplementing lysins and methionine to low-protein diets on growth performance, hepatic antioxidant capacity, immune status, and glycolytic activity of tibetan sheep.

Reducing the levels of dietary protein is an effective nutritional approach in lowering feed cost and nitrogen emissions in ruminants. The purpose of this study was to evaluate the effects of dietary Lys/Met ratio in a low protein diet (10%, dry matter basis) on the growth performance and hepatic function (antioxidant capacity, immune status, and glycolytic activity) in Tibetan lambs. Ninety two-month-old rams with an average weight of 15.37 ± 0.92 kg were randomly assigned to LP-L (dietary Lys/Met = 1:1), LP-M (dietary Lys/Met = 2:1) and LP-H (dietary Lys/Met = 3:1) treatments. The trial was conducted over 100 d, including 10 d of adaption to the diets. Hepatic phenotypes, antioxidant capacity, immune status, glycolytic activity and gene expression profiling was detected after the conclusion of the feeding trials. The results showed that the body weight was higher in the LP-L group when compared to those on the LP-M group (P < 0.05). In addition, the activities of the catalase (CAT) and glutathione peroxidase (GSH-Px) in the LP-L group were significantly increased compared with the LP-M group (P < 0.05), while the malondialdehyde (MDA) levels in LP-H group were significantly decreased (P < 0.05). Compared with LP-H group, both hepatic glycogen (P < 0.01) and lactate dehydrogenase (LDH) (P < 0.05) were significantly elevated in LP-L group. For the LP-L group, the hepatocytes were arranged radially with the central vein in the center, and hepatic plates exhibited tight arrangement. Transcriptome analysis identified 29, 179, and 129 differentially expressed genes (DEGs) between the LP-M vs. LP-L, LP-H vs. LP-M, and LP-H vs. LP-L groups, respectively (Q-values < 0.05 and |log2Fold Change| > 1). Gene Ontology (GO) and correlation analyses showed that in the LP-L group, core genes (C1QA and JUNB) enriched in oxidoreductase activity were positively correlated with antioxidant indicators, while the MYO9A core gene enriched in the immune response was positively associated with immune indicators, and core genes enriched in molecular function (PDK3 and PDP2) were positively correlated with glycolysis indicators. In summary, low-protein diet with a low Lys/Met ratio (1:1) could reduce the hepatic oxidative stress and improve the glycolytic activity by regulating the expression of related genes of Tibetan sheep.

Effects of supplementing coated methionine in a high plant-protein diet on growth, antioxidant capacity, digestive enzymes activity and expression of TOR signaling pathway associated genes in gibel carp, Carassius auratus gibelio.

This study explored the impacts of supplementation of different levels of coated methionine (Met) in a high-plant protein diet on growth, blood biochemistry, antioxidant capacity, digestive enzymes activity and expression of genes related to TOR signaling pathway in gibel carp (Carassius auratus gibeilo). A high-plant protein diet was formulated and used as a basal diet and supplemented with five different levels of coated Met at 0.15, 0.30, 0.45, 0.60 and 0.75%, corresponding to final analyzed Met levels of 0.34, 0.49, 0.64, 0.76, 0.92 and 1.06%. Three replicate groups of fish (initial mean weight, 11.37 ± 0.02 g) (20 fish per replicate) were fed the test diets over a 10-week feeding period. The results indicated that with the increase of coated Met level, the final weight, weight gain (WG) and specific growth rate initially boosted and then suppressed, peaking at 0.76% Met level (P< 0.05). Increasing dietary Met level led to significantly increased muscle crude protein content (P< 0.05) and reduced serum alanine aminotransferase activity (P< 0.05). Using appropriate dietary Met level led to reduced malondialdehyde concentration in hepatopancreas (P< 0.05), improved superoxide dismutase activity (P< 0.05), and enhanced intestinal amylase and protease activities (P< 0.05). The expression levels of genes associated with muscle protein synthesis such as insulin-like growth factor-1, protein kinase B, target of rapamycin and eukaryotic initiation factor 4E binding protein-1 mRNA were significantly regulated, peaking at Met level of 0.76% (P< 0.05). In conclusion, supplementing optimal level of coated Met improved on fish growth, antioxidant capacity, and the expression of TOR pathway related genes in muscle. The optimal dietary Met level was determined to be 0.71% of the diet based on quadratic regression analysis of WG.

Relative biological efficacy of methionine hydroxy analogue-free acid compared to dl-methionine in the broiler chickens.

BACKGROUND: In the broiler's diets based on corn-soya bean meal, methionine (Met) and cystine (Cys), known as sulphur amino acids (SAAs), are the first limiting indispensable amino acids because of their limited presence, which are supplemented with different synthetic sources. Evaluation of the biological effectiveness of these sources can be important in their correct replacement, especially in the starter and growth diets. OBJECTIVES: The current study was done to assess the relative biological efficacy (RBE) of liquid Met hydroxy analogue-free acid (MHA-FA) in comparison with dl-Met (dl-Met) based on broiler performance traits at different levels of digestible SAA in the 1-11 (starter) and 11-25 (grower) days of age periods. METHODS: Two experiments were developed with treatments consisting of a basal diet without Met addition that met the nutrient and energy requirements of broilers with the exception of SAAs (Met + Cys) and five increasing Met doses for both sources (dl-Met and/or MHA-FA), resulting in digestible SAA concentrations from 0.62% to 1.02% of diet in the starter period (Trial 1) and 0.59% to 0.94% of diet in the grower period (Trial 2). The multi-linear regression model and slope ratio method were employed to calculate the RBE of MHA-FA compared with dl-Met for measured variables. RESULTS: In both experiments, the results obtained during the starter and grower periods with the different Met supplementations show significant growth responses to digestible SAAs levels. By increasing dietary dl-Met and/or MHA-FA levels, the growth performance traits and immune responses were improved (quadratic; p < 0.05). The RBE of MHA-FA compared to dl-Met on an equimolar basis was estimated 66%-89% (59%-79% on a weight-to-weight basis). CONCLUSIONS: It is concluded that the RBE of MHA-FA in comparison with dl-Met depends on broiler chicken age and what attribute is being evaluated.

Targeting Methionine Addiction Combined With Low-dose Irinotecan Arrested Colon Cancer in Contrast to High-dose Irinotecan Alone, Which Was Ineffective, in a Nude-mouse Model.

BACKGROUND/AIM: Colorectal cancer (CRC) is the third-leading cause of death in the world. Although the prognosis has improved due to improvement of chemotherapy, metastatic CRC is still a recalcitrant disease, with a 5-year survival of only 13%. Irinotecan (IRN) is used as first-line chemotherapy for patients with unresectable CRC. However, there are severe side effects, such as neutropenia and diarrhea, which are dose-limiting. We have previously shown that methionine restriction (MR), effected by recombinant methioninase (rMETase), lowered the effective dose of IRN of colon-cancer cells in vitro. The aim of the present study was to evaluate the efficacy of the combination of low-dose IRN and MR on colon-cancer in nude mice. MATERIALS AND METHODS: HCT-116 colon-cancer cells were cultured and subcutaneously injected into the flank of nude mice. After the tumor size reached approximately 100 mm3, 18 mice were randomized into three groups; Group 1: untreated control on a normal diet; Group 2: high-dose IRN on a normal diet (2 mg/kg, i.p.); Group 3: low-dose IRN (1 mg/kg i.p.) on MR effected by a methionine-depleted diet. RESULTS: There was no significant difference between the control mice and the mice treated with high-dose IRN, without MR. However, low-dose IRN combined with MR was significantly more effective than the control and arrested colon-cancer growth (p=0.03). Body weight loss was reversible in the mice treated by low-dose IRN combined with MR. CONCLUSION: The combination of low-dose IRN and MR acted synergistically in arresting HCT-116 colon-cancer grown in nude mice. The present study indicates the MR has the potential to reduce the effective dose of IRN in the clinic.

Methionine Dependence of Hair Maintenance in C57BL/6 Mice.

BACKGROUND/AIM: Hair-follicle keratinocytes contain high levels of cysteine, which is derived from methionine, rapidly proliferate, and form the hair shaft. The high proliferation rate of hair-follicle keratinocytes resembles that of aggressive cancer cells. In the present study, we determined the effect of a methionine-deficient diet on hair loss (alopecia) in mice with or without homocysteine supplementation. MATERIALS AND METHODS: Mice were fed a normal rodent diet (2020X, ENVIGO) (Group 1); a methionine-choline-deficient diet (TD.90262, ENVIGO) (Group 2); a methionine-choline-deficient diet with a 10 mg/kg/day supply of homocysteine administered by intra-peritoneal (i.p.) injection for 2 weeks (Group 3). In Group 2, mice were fed a methionine-choline-deficient diet for an additional 2 weeks but with 10 mg/kg/day of i.p. l-homocysteine and the mice were observed for two additional weeks. Subsequently, the mice were fed a standard diet that included methionine. Hair loss was monitored by photography. RESULTS: After 14 days, hair loss was observed in Group 2 mice on a methionine-restricted diet but not in Group 3 mice on the methionine-restricted diet which received i.p. homocysteine. In Group 2, at 2 weeks after methionine restriction, hair loss was not rescued by homocysteine supplementation. However, after restoration of methionine in the diet, hair growth resumed. Thus, after 2 weeks of methionine restriction, only methionine restored hair loss, not homocysteine. CONCLUSION: Hair maintenance requires methionine in the diet. Future experiments will determine the effects of methionine restriction on hair-follicle stem cells.

Extensive Shrinkage and Long-term Stable Disease in a Teenage Female Patient With High-grade Glioma Treated With Temozolomide and Radiation in Combination With Oral Recombinant Methioninase and a Low-methionine Diet.

BACKGROUND/AIM: Gliomas are the most common and recalcitrant malignant primary brain tumors. All cancer types are addicted to methionine, which is a fundamental and general hallmark of cancer known as the Hoffman effect. Particularly glioma cells exhibit methionine addiction. Because of methionine addiction, [11C]-methionine positron emission tomography (MET-PET) is widely used for glioma imaging in clinical practice, which can monitor the extent of methionine addiction. Methionine restriction including recombinant methioninase (rMETase) and a low-methionine diet, has shown high efficacy in preclinical models of gliomas, especially in combination with chemotherapy. The aim of the present study was to determine the efficacy of methionine restriction with oral rMETase (o-rMETase) and a low-methionine diet, combined with radiation and temozolomide (TMZ), on a teenage female patient with high-grade glioma. CASE REPORT: A 16-year-old girl was diagnosed with high-grade glioma. Magnetic resonance imaging (MRI) showed a left temporal-lobe tumor with compression to the left lateral ventricle and narrowing of sulci in the left temporal lobe. After the start of methionine restriction with o-rMETase and a low-methionine diet, along with TMZ combined with radiotherapy, the tumor size shrunk at least 60%, with improvement in the left lateral ventricle and sulci. The patient's condition remains stable for 19 months without severe adverse effects. CONCLUSION: Methionine restriction consisting of o-rMETase and a low-methionine diet, in combination with radiation and TMZ as first-line chemotherapy, were highly effective in a patient with high-grade glioma.

Different methionine to cysteine supplementation ratios altered bone quality of broilers with or without Eimeria challenge assessed by dual energy X-ray absorptiometry and microtomography.

Despite the acknowledged significance of nutrition in bone development, effects of methionine (Met) and cysteine (Cys) on bone quality remain under-researched, particularly during Eimeria challenge. We investigated the effects of different supplemental Met to Cys ratios (MCR) on bone quality of broilers under Eimeria challenge. A total of 720 fourteen-day old Cobb500 broilers were allocated into a 5 × 2 factorial arrangement. Five diets with Met and Cys supplemented at MCR of 100:0, 75:25, 50:50, 25:75, and 0:100 were fed to the birds with or without Eimeria challenge. Body composition was measured by dual energy x-ray absorptiometry, and the femur bone characteristics were assessed by microtomography. Data were analyzed by two-way ANOVA and orthogonal polynomial contrast. The results reaffirmed the detrimental effects of Eimeria challenge on bone quality. On 9 d post inoculation (DPI), significant interaction effects were found for whole body bone mineral content (BMC), lean tissue weight, and body weight (P < 0.05); in the nonchallenged group (NCG), these parameters linearly decreased as MCR decreased (P < 0.05). In the challenged group (CG), body weight and lean tissue weight were unaffected by MCR, and BMC linearly increased as MCR decreased (P < 0.05). For the cortical bone of femoral metaphysis on 6 DPI, bone mineral density (BMD) linearly increased as MCR decreased (P < 0.05). Bone volume to tissue volume ratio (BV/TV) in the CG linearly increased as MCR decreased (P < 0.05). On 9 DPI, BMC and TV linearly increased as MCR decreased (P < 0.05) in the NCG. BMD and BV/TV changed quadratically as MCR decreased (P < 0.05). For the trabecular bone of femoral metaphysis on 9 DPI, BV/TV, and trabecular number linearly increased as MCR decreased (P < 0.05) in the NCG. For the femoral diaphysis, BV, TV, BMC on 6 DPI, and BMD on 9 DPI linearly increased as MCR decreased (P < 0.05). In conclusion, this study showed that both Eimeria challenge and varying supplemental MCR could influence bone quality of broilers.

Higher dietary methyl donor micronutrient consumption is associated with higher muscle strength in adults: a cross-sectional study.

Methyl donor micronutrients might affect muscle strength via DNA methylation. We aimed to evaluate the combined relationship of dietary methyl donor micronutrients containing betaine, choline, methionine, vitamin B12, vitamin B6 and folate on muscle strength. This cross-sectional study was conducted on 267 subjects including 113 men and 154 women. Dietary intake of micronutrients was assessed utilising a validated 168-item semi-quantitative FFQ, and methyl donor micronutrient score (MDMS) was calculated. The muscle strength of the participants was measured using a digital handgrip dynamometer. The association was determined using linear regression analysis. The mean age of participants was 36·8 ± 13·2 years. After taking into account potential confounding variables, there was no significant association between dietary methyl donor micronutrient score (MDMS) and the mean left-hand muscle strength (ß: 0·07, se: 0·05, P = 0·07); however, the changes were significant in the mean right-hand muscle strength (ß: 0·09, se: 0·04, P = 0·03). There was also a significant positive relationship between mean muscle strength and methyl donors' intake after fully adjusting for potential confounders (ß: 0·08, se: 0·04, P = 0·04). In conclusion, our findings revealed that higher dietary methyl donor micronutrient consumption is associated with enhanced muscle strength. As a result, advice on a higher intake of methyl donor-rich foods including grains, nuts, dairy products and seafood might be recommended by dietitians as a general guideline to adhere to. Additional prospective studies are needed to confirm the findings.

Prognostic value of 11C-methionine volume-based PET parameters in IDH wild type glioblastoma.

PURPOSE: 11C-Methionine (11C-MET) PET prognostication of isocitrate dehydrogenase (IDH) wild type glioblastomas is inadequate as conventional parameters such as standardized uptake value (SUV) do not adequately reflect tumor heterogeneity. We retrospectively evaluated whether volume-based parameters such as metabolic tumor volume (MTV) and total lesion methionine metabolism (TLMM) outperformed SUV for survival correlation in patients with IDH wild type glioblastomas. METHODS: Thirteen IDH wild type glioblastoma patients underwent preoperative 11C-MET PET. Both SUV-based parameters and volume-based parameters were calculated for each lesion. Kaplan-Meier curves with log-rank testing and Cox regression analysis were used for correlation between PET parameters and overall survival. RESULTS: Median overall survival for the entire cohort was 393 days. MTV (HR 1.136, p = 0.007) and TLMM (HR 1.022, p = 0.030) were inversely correlated with overall survival. SUV-based 11C-MET PET parameters did not show a correlation with survival. In a paired analysis with other clinical parameters including age and radiotherapy dose, MTV and TLMM were found to be independent factors. CONCLUSIONS: MTV and TLMM, and not SUV, significantly correlate with overall survival in patients with IDH wild type glioblastomas. The incorporation of volume-based 11C-MET PET parameters may lead to a better outcome prediction for this heterogeneous patient population.

Betaine and related compounds: Chemistry, metabolism and role in mitigating heat stress in poultry.

Betaine can operate as an osmolyte and a methyl donor. Betaine is an osmolyte and a methyl donor. Betaine is likewise a zwitterion with osmotic capabilities that can help an animal cope with osmotic stress. Previous investigations have suggested that betaine has various impacts, albeit these studies do not consistently provide the same results. Dietary betaine has received a lot of attention owing to its osmoprotectant, methionine-sparing and antioxidant properties. Betaine is extensively assessed concerning performance and body composition. The tolerance to high temperatures, flock livability, and breast meat output is among the factors frequently mentioned in the literature as being altered by betaine. Betaine, a multi-nutritional agent, may help poultry resist heat stress and poor management. A common subject of betaine research is the idea of betaine saving some methionine. Although research on betaine may not always come to the same results, some discoveries repeat themselves. Because of their effectiveness in increasing growth performance, feed utilization, meat quality, and alleviating heat stress in chicken farms, betaine and methionine are extensively used as feed supplements in poultry diets. This review highlights the influences of betaine on poultry performance, meat quality, carcass characteristics, antioxidant activity, in addition to its role in mitigating heat stress.

Dietary Intake Adequacy and Food Sources of Nutrients Involved in the Methionine-Methylation Cycle in Women of Childbearing Age from the ANIBES Spanish Population.

Growing evidence confirms choline as a critical perinatal nutrient. However, intake levels of choline and betaine among the Spanish fertile population remain unknown. Given their role in one-carbon metabolism with potential epigenetic effects, the aim of the present study was to evaluate the dietary intakes, their adequacy to existing guidelines and the main food sources together with other micronutrients involved in the methylation-methionine cycle (vitamin B6, folates and vitamin B12) in women of childbearing age. The ANIBES study, a cross-sectional study of a representative sample of women of childbearing age (18-45 years, n = 641) resident in Spain, was used. The sample was divided into younger women (18-30 years, n = 251) and older women (31-45 years, n = 390). Dietary intake was assessed by a three-day dietary record by using a tablet device. Total median intakes for the total sample were 303.9 mg/d for choline; 122.6 mg/d for betaine; 1.3 mg/d for vitamin B6; 140.8 µg/d for folates, and 3.8 µg/d for vitamin B12. The older subgroup showed significantly higher choline (p < 0.05), betaine (p < 0.001) and folates (p < 0.05) intakes than younger women. Main food sources for the whole sample were meat and meat products for choline (28.3%), vitamin B6 (25.7%) and vitamin B12 (22.8%); cereals and derivatives (79.9%) for betaine; vegetables (20.0%) for folates. Overall intake adequacy was only observed for vitamin B12, with a very limited number of participants showing adequate intakes for all the other micronutrients. These results illustrate there is a relevant need to raise awareness about optimizing the status of the methionine cycle-related vitamins and cofactors in this potentially vulnerable population.

Association of dietary intakes of vitamin B12, vitamin B6, folate, and methionine with the risk of esophageal cancer: the Japan Public Health Center-based (JPHC) prospective study.

BACKGROUND: B vitamins and methionine are essential substrates in the one-carbon metabolism pathway involved in DNA synthesis and methylation. They may have essential roles in cancer development. We aimed to evaluate the associations of dietary intakes of vitamin B12, vitamin B6, folate, and methionine with the risk of esophageal cancer (EC) using data from the Japan Public Health Center-based Prospective Study. METHODS: We included 87,053 Japanese individuals who completed a food frequency questionnaire and were followed up from 1995-1998 to 2013 and 2015. Hazard ratios (HRs) and 95% confidence intervals (CIs) were calculated by Cox proportional-hazard regression across quintiles of dietary intakes of B vitamins and methionine. RESULTS: After 1,456,678 person-years of follow-up, 427 EC cases were documented. The multivariable HR (95% CI) of incident EC in the highest versus lowest quintile of dietary intake of vitamin B12 was 1.75 (1.13-2.71; p-trend=0.01). Stratification analysis based on alcohol consumption showed that higher dietary intakes of vitamin B12 and methionine were associated with an increased risk of EC among never-drinkers; HRs (95% CIs) were 2.82 (1.18-6.74; p-trend=0.009; p-interaction=0.18) and 3.45 (1.32-9.06; p-trend=0.003; p-interaction 0.02) for vitamin B12 and methionine, respectively. Meanwhile, there was no association between vitamin B12 and methionine intake with the risk of EC among drinkers. There were no associations between dietary intake of folate or vitamin B6 and the risk of EC. CONCLUSION: Dietary intake of vitamin B12 was positively associated with the risk of EC in the Japanese population.

Effect of chromium-methionine supplementation on meat quality of broilers reared under heat stress / [Efeito da suplementação de cromo-metionina na qualidade da carne de frangos de corte criados em estresse por calor]

Objetivou-se avaliar o efeito da suplementação de cromo-metionina em dietas para frangos de corte criados em estresse por calor, no período de 22 a 43 dias de idade, nos parâmetros de qualidade da carne. Foram utilizados 336 frangos de corte, machos, da linhagem Cobb 500, com 21 dias de idade, distribuídos em delineamento em blocos inteiramente ao acaso, com quatro blocos (cada câmara climática), seis tratamentos (0; 0,10; 0,20; 0,40; 0,80 e 1,20mgkg-1 de Cr na forma de Cr-metionina), oito repetições e sete aves por unidade experimental. Aos 43 dias de idade, duas aves por unidade experimental foram selecionadas e abatidas para avaliação da qualidade da carne de peito, por meio dos parâmetros de pH15min, pH24h, luminosidade (L*), teor de vermelho (a*), teor de amarelo (b*), croma (C*), ângulo hue (Hº), capacidade de retenção de água, perda de peso por cozimento e força de cisalhamento. Houve efeito quadrático (P=0,0070) na capacidade de retenção de água da carne de peito. A suplementação de CrMet não afetou (P>0,05) os demais parâmetros de qualidade da carne. Assim, recomenda-se a suplementação de 0,59mgkg-1 de CrMet para frangos de corte para melhoria da capacidade de retenção de água do peito.(AU)

Dietary intake of sulfur amino acids and risk of kwashiorkor malnutrition in eastern Democratic Republic of the Congo.

BACKGROUND: Kwashiorkor is an often-fatal type of severe acute malnutrition affecting hundreds of thousands of children annually, but whose etiology is still unknown. Evidence suggests inadequate sulfur amino acid (SAA) status may explain many signs of the condition but studies evaluating dietary protein intake in relation to the genesis of kwashiorkor have been conflicting. We know of no studies of kwashiorkor that have measured dietary SAAs. OBJECTIVES: We aimed to determine whether children in a population previously determined to have high prevalence of kwashiorkor [high-prevalence population (HPP)] have lower dietary intakes of SAAs than children in a low-prevalence population (LPP). METHODS: A cross-sectional census survey design of 358 children compared 2 previously identified adjacent populations of children 36-59 mo old in North Kivu Province of the Democratic Republic of the Congo. Data collected included urinary thiocyanate (SCN), cyanogens in cassava-based food products, recent history of illness, and a 24-h quantitative diet recall for the child. RESULTS: The HPP and LPP had kwashiorkor prevalence of 4.5% and 1.7%, respectively. A total of 170 children from 141 households in the LPP and 169 children from 138 households in the HPP completed the study. A higher proportion of HPP children had measurable urinary SCN (44.8% compared with 29.4%, P < 0.01). LPP children were less likely to have been ill recently (26.8% compared with 13.6%, P < 0.01). Median [IQR] intake of SAAs was 32.4 [22.9-49.3] mg/kg for the LPP and 29.6 [18.1-44.3] mg/kg for the HPP (P < 0.05). Methionine was the first limiting amino acid in both populations, with the highest risk of inadequate intake found among HPP children (35.1% compared with 23.6%, P < 0.05). CONCLUSIONS: Children in a population with a higher prevalence of kwashiorkor have lower dietary intake of SAAs than children in a population with a lower prevalence. Trial interventions to reduce incidence of kwashiorkor should consider increasing SAA intake, paying particular attention to methionine.

Optimal methionine plus cystine requirements in diets supplemented with L-methionine in starter, grower, and finisher broilers.

Correct supplementation of dietary amino acids, such as methionine (Met) and cystine (Cys), is crucial to support the exponential growth of broilers. Historically, most available recommendations with regard to the optimal amount of Met plus Cys are based on studies wherein DL-Met was used as the Met source. Nowadays, L-Met is available as a registered feed additive, urging the need to establish the optimal L-Met plus Cys supplementation. The objective of this trial was to investigate these optimal L-Met plus Cys requirements of broilers in the starter (0-10 d), grower (11-23 d), and finisher (24-35 d) phase of life separately. A basal diet deficient in L-Met plus Cys was created along with 6 other diets with increasing L-Met concentrations for each phase. Birds were only included in one life phase and fed with a commercial diet before inclusion. The BW, daily weight gain, daily feed intake, and feed conversion ratio (gain-to-feed ratio) were measured for all birds. Slaughter parameters were determined for birds included in the finisher phase. At the end of each study period, significant differences (P < 0.05) were observed in all measured performance parameters. Birds fed with the deficient diets were characterized by a lower performance, whereas from some point, no gain in performance could be observed. Correct supplementation of L-Met plus Cys seemed more crucial in the starter and grower phase, which was characterized by bigger differences in performance between test diets compared with the finisher birds. The optimal L-Met plus Cys requirements were determined using linear broken line and exponential asymptotic models. The linear broken line model showed overall the best fit. The optimal L-Met plus Cys level was found to be 0.69, 0.66, and 0.62% for birds in the starter, grower, and finisher phase, respectively. From this study, it could be concluded that broilers have lower L-Met plus Cys requirements based on L-Met supplementation than the conventional requirements based on DL-Met. Nevertheless, further research is required to confirm these findings.

Reduced hepatic global hydroxymethylation in mice treated with non-genotoxic carcinogens is transiently reversible with a methyl supplemented diet.

Non-genotoxic carcinogens (NGCs) are known to cause perturbations in DNA methylation, which can be an early event leading to changes in gene expression and the onset of carcinogenicity. Phenobarbital (PB) has been shown to alter liver DNA methylation and hydroxymethylation patterns in mice in a time dependent manner. The goals of this study were to assess if clofibrate (CFB), a well-studied rodent NGC, would produce epigenetic changes in mice similar to PB, and if a methyl donor supplementation (MDS) would modulate epigenetic and gene expression changes induced by phenobarbital. CByB6F1 mice were treated with 0.5% clofibrate or 0.14% phenobarbital for 7 and 28 days. A subgroup of PB treated and control mice were also fed MDS diet. Liquid Chromatography-Ionization Mass Spectrometry (LC-MS) was used to quantify global liver 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) levels. Gene expression analysis was conducted using Affymetrix microarrays. A decrease in liver 5hmC but not 5mC levels was observed upon treatment with both CFB and PB with varying time of onset. We observed moderate increases in 5hmC levels in PB-treated mice when exposed to MDS diet and lower expression levels of several phenobarbital induced genes involved in cell proliferation, growth, and invasion, suggesting an early modulating effect of methyl donor supplementation. Overall, epigenetic profiling can aid in identifying early mechanism-based biomarkers of non-genotoxic carcinogenicity and increases the quality of cancer risk assessment for candidate drugs. Global DNA methylation assessment by LC-MS is an informative first step toward understanding the risk of carcinogenicity.

Dietary supplementations of methionine improve growth performances, innate immunity, digestive enzymes, and antioxidant activities of rohu (Labeo rohita).

The dietary composition of fish constitutes numerous immune and growth-promoting substance; during the current study, methionine was amended in the diet of Labeo rohita (L. rohita), in order to appraise the diverse modulatory effects of methionine supplementation. Fish were grouped into three on the basis of methionine as methionine supplemented group 1 (MSG1), methionine supplemented group 2 (MSG2), and CTRL (control group). We assessed the effects of methionine at the end. Captivating results were obtained from the methionine supplementing groups. The results depict significantly high (p < 0.05) innate immunity parameters, including myeloperoxidase, IgG concentrations, lysozyme, respiratory burst, and total bacteriolytic activity of complement in MSG2. Furthermore, ameliorate growth performances were also noticed such as high weight gain, FCE%, and body length in MSG2, respectively. We also measured high level of antioxidant digestive enzymes, gut length, and survival rate in MSG2 compared to the rest of the groups. The results were further validated by measuring the stress parameters, such as aspartate aminotransferase (AST), glucose, and hematocrit (HCT%), and the concentration remains low in MSG2. Lastly, after 90 days, fish were challenged with Aeromonas hydrophila, the highest survival was observed in the MSG2 and MSG1 compared to CTRL. These results ultimately give the baseline to the credible effects of methionine on L. rohita.

Onset of Marine-Lenhart syndrome and Graves' ophthalmopathy in a female patient treated with alemtuzumab for multiple sclerosis.

BACKGROUND: Immune checkpoint blockade therapy may lead to thyroid dysfunction in 3-7% of treated patients. Alemtuzumab is a CD52 inhibitor leading to thyroid dysfunction in approximately 40% of patients. A female patient was affected by multiple sclerosis (MS) and subclinical hyperthyroidism due to an autonomously functioning thyroid nodule (AFTN). After alemtuzumab treatment, she developed aggressive clinical hyperthyroidism consistent with Marine-Lenhart syndrome. CASE PRESENTATION: A 36-year-old woman presented in July 2019 with symptoms of hyperthyroidism and eye complaints. Three years earlier, she was diagnosed with MS. Subclinical hyperthyroidism was diagnosed in April 2017. Thyroid scintigraphy showed an intranodular distribution of 99mTc-pertechnatate consisting of an AFTN in the right lobe of the thyroid. In June 2018, because of the MS, she was treated with alemtuzumab. In November 2018, she was started on methimazole treatment because of the symptoms of hyperthyroidism. In December 2018, thyroid function was normal under methimazole treatment. In June 2019, the patient received a second round of alemtuzumab administration. One month later, she developed symptoms of hyperthyroidism. These symptoms were accompanied by diplopia. Blood tests showed severe hyperthyroidism. Thyroid scintigraphy showed a diffuse distribution of 99mTc-pertechnatate and the presence of a "cool" area in the right lobe of the thyroid, confirmed by ultrasonography. The nodule was diagnosed as a low-risk indeterminate lesion. CONCLUSION: We present a case of Graves' disease with active, moderate-to-severe Graves' ophthalmopathy in a patient with pre-existing AFTN presenting with a coexisting, rare case of Marine-Lenhart syndrome associated with immune reconstitution after alemtuzumab treatment.

Effect of methionine deficiency on the apoptosis and cell cycle of kidney in broilers.

Abundant evidence now supports the idea that methionine deficiency has negative effects on chicken healthy especially in the aspect of cell cycle regulation and apoptosis. But, lacking of knowledge is the evaluation on metabolic organs. To test the effect of methionine deficiency on the kidney, we assessed the apoptosis and the cell cycle of kidney induced by methionine deficiency by the methods of TdT-mediated dUTP Nick-End Labeling (TUNEL), flow cytometry (FCM), quantitative real-time PCR (qRT-PCR) and Enzyme-Linked ImmunoSorbent Assay (ELISA) of chickens for 42 days of age. Our results showed that the number of the apoptotic cells was increased (p < .05 or p < .01), while bcl-2 mRNA expression levels were decreased (p < .05 or p < .01) and bax and caspase-3 mRNA expression levels were higher (p < .01) in methionine deficiency group. Furthermore, the cell cycle results showed a time-dependent increase in G2M phase cells and a corresponding decrease in cells at G2M and S stages, the mRNA expression of p53 and p21 was increased (P < .05 or P < .01) and cyclin B and PCNA was significantly lower (P < .05 or P < .01) in the methionine deficiency group than that of the control group. These findings suggested that methionine deficiency could induce renal apoptosis and cell cycle arrest.

Alterations in immune and antioxidant gene networks by gamma-d-glutamyl-meso-diaminopimelic acid in bovine mammary epithelial cells are attenuated by in vitro supply of methionine and arginine.

Nucleotide-binding oligomerization domain (NOD)-like receptor 1 (NOD1) is a cytosolic pattern recognition receptor with a crucial role in the innate immune response of cells triggered by the presence of compounds such as gamma-d-glutamyl-meso-diaminopimelic acid (iE-DAP) present in the peptidoglycan of all gram-negative and certain gram-positive bacteria. Methionine (Met) and arginine (Arg) are functional AA with immunomodulatory properties. In the present study, we aimed to assess the effect of increased Met and Arg supply on mRNA abundance of genes associated with innate immune response, antioxidant function, and AA metabolism during iE-DAP challenge in bovine mammary epithelial cells (BMEC). Primary BMEC (n = 4 per treatment) were precultured in modified medium for 12 h with the following AA formulations: ideal profile of AA (control), increased Met supply (incMet), increased Arg supply (incArg), or increased supply of Met plus Arg (incMetArg). Subsequently, cells were challenged with or without iE-DAP (10 µg/mL) for 6 h. Data were analyzed as a 2 × 2 × 2 factorial using the MIXED procedure of SAS 9.4. Greater mRNA abundance of NOD1, the antioxidant enzyme SOD1, and AA transporters (SLC7A1 and SLC3A2) was observed in the incMet cells after iE-DAP stimulation. Although increased Met alone had no effect, incMetArg led to greater abundance of the inflammatory cytokine IL-6, and the antioxidant enzyme GPX1 after iE-DAP stimulation. The increased Arg alone downregulated NOD1 after iE-DAP stimulation, coupled with a downregulation in the AA transporters mRNA abundance (SLC7A1, SLC7A5, SLC3A2, and SLC38A9), and upregulation in GSS and KEAP1 mRNA abundance. Overall, the data indicated that increased supply of both Met and Arg in the culture medium were more effective in modulating the innate immune response and antioxidant capacity of BMEC during in vitro iE-DAP stimulation.