Serum Methionine Research Articles

Ethanol feeding of micropigs alters methionine metabolism and increases hepatocellular apoptosis and proliferation

Chronic alcoholism is associated with increased cancer risk that may be related to ethanol-induced alterations in methionine and deoxynucleotide metabolism. These metabolic relationships were studied in micropigs fed diets for 12 months that contained 40% ethanol or cornstarch control with adequate folate. Ethanol feeding altered methionine metabolism without changing mean terminal liver folate levels. After initial equilibration to diet, ethanol feeding significantly increased monthly serum homocysteine levels while reducing serum methionine levels over the time course of the experiment. After 12 months, hepatic methionine synthase activity and the ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH) were significantly reduced in ethanol-fed animals, whereas the ratio of liver deoxyuridine triphosphate (dUTP) to deoxythymidine triphosphate (dTTP) was increased and correlated inversely with methionine synthase activity. These findings were associated with increased frequency of hepatocytes with apoptotic bodies and positivity for proliferating cell nuclear antigen (PCNA) in livers from ethanol-fed minipigs. These studies suggest that chronic ethanol feeding perturbs methionine metabolism by impairment of methionine synthase activity, resulting in deoxynucleoside triphosphate (dNTP) imbalance, increased apoptosis, and regenerative proliferation. These biochemical alterations may provide a promoting environment for carcinogenesis during long-term ethanol exposure. (Hepatology 1996 Mar;23(3):497-505)

Ethanol feeding of micropigs alters methionine metabolism and increases hepatocellular apoptosis and proliferation.

Chronic alcoholism is associated with increased cancer risk that may be related to ethanol-induced alterations in methionine and deoxynucleotide metabolism. These metabolic relationships were studied in micropigs fed diets for 12 months that contained 40% ethanol or cornstarch control with adequate folate. Ethanol feeding altered methionine metabolism without changing mean terminal liver folate levels. After initial equilibration to diet, ethanol feeding significantly increased monthly serum homocysteine levels while reducing serum methionine levels over the time course of the experiment. After 12 months, hepatic methionine synthase activity and the ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH) were significantly reduced in ethanol-fed animals, whereas the ratio of liver deoxyuridine triphosphate (dUTP) to deoxythymidine triphosphate (dTTP) was increased and correlated inversely with methionine synthase activity. These findings were associated with increased frequency of hepatocytes with apoptotic bodies and positivity for proliferating cell nuclear antigen (PCNA) in livers from ethanol-fed minipigs. These studies suggest that chronic ethanol feeding perturbs methionine metabolism by impairment of methionine synthase activity, resulting in deoxynucleoside triphosphate (dNTP) imbalance, increased apoptosis, and regenerative proliferation. These biochemical alterations may provide a promoting environment for carcinogenesis during long-term ethanol exposure.

Carbon tetrachloride-induced hepatic injury is associated with global dna hypomethylation and homocysteinemia: Effect of s-adenosylmethionine treatment

Carbon tetrachloride (CC1 4) administration to rats produces hepatic cirrhosis and supplementation with S-adenosylmethionine (SAM) can partially prevent CCl 4-induced liver injury. These effects are thought to be caused by oxidative stress and the subsequent formation of free radicals, but the mechanism whereby this occurs and the accurate nature of the mechanisms by which SAM exerts its protective action are not well understood. The effect of short-term administration of CC1 4on hepatic DNA methylation and on SAM and S-adenosylhomocysteine (SAH) were assessed. CCl 4 administration to rats for 3 weeks resulted in hypomethylation of liver DNA, determined by comparing the extent to which DNA from livers of control or treated animals could be methylated in vitro using [ 3H-methyl] SAM as methyl donor. This CC1 4 effect on DNA methylation was corrected by the administration of SAM (10 mg/kg/d, intramuscularly), with values of methyl groups incorporation comparable with those observed in the control animals. Hepatic SAM was decreased by CCl 4 (65.3 ± 5.27 vs. 102.2 ± 4.89 μmol/g; P < .05) and SAH was increased (69.5± 14.6 vs. 29.4± 3.83 nmol/g; P <.05). This led to a marked reduction of the SAM/SAH ratio (the methylation ratio) from 3.47 in control rats to 0.94 in CC1 4-treated animals ( P < .05). SAM treatment partially prevented ( P < .05) the reduction of the ratio SAM/SAH induced by CC1 4. CC1 4 also induced a marked elevation of serum homocysteine levels (more than 20-fold; P < .001), which was partially prevented by SAM administration. A decrease in serum methionine concentration was also observed (20.87 ± 1.76 vs. 31.25 ± 2.37 μmol/L; P < .05) in response to CCl 4, whereas cystathionine levels remained unchanged. Hepatic folate was reduced by CC1 4 (11.2 ± 2.1 vs. 17.6 ± 1.8 μmol/L; P < .05) and SAM treatment prevented this effect ( P < .05). Hepatic glutathione (reduced form) and the activity of the enzyme SAM synthetase, which have been found to be reduced after the administration of CC1 4 to rats for longer periods, were unchanged after treatment with the hepatotoxic agent for only 3 weeks. It is proposed that CCl 4 disrupts the distribution of homocysteine between remethylation and its degradation via the transsulphuration pathway and that SAM, by resetting the methylation ratio, restores this equilibrium.

Levels of amino acids in human hepatocellular carcinoma and adjacent liver tissue.

Total parenteral nutrition can be used to overcome amino acid imbalance in cancer patients. Because there is little documentation of treatment for amino acid imbalance in patients with hepatocellular carcinoma (HCC), we designed a study 1) to compare tissue levels of amino acids between HCC and the adjacent liver and 2) to determine which serum amino acids correlate to tumor volume. A significant elevation of methionine and a significant decrease of glycine and cystine were observed in HCC compared with adjacent liver tissue, and a significant correlation was found between tumor volume and serum methionine levels (r = -0.636, p < 0.01). Thus the tumor tissue competes successfully with host tissue for nitrogen substrates, particularly methionine, and an accelerated protein synthesis in HCC consumes large amounts of these amino acids. The possibility of methionine-depleted treatment could be considered for patients with HCC.

Total sulfur amino acid requirement and cysteine replacement value for juvenile hybrid striped bass (Morone saxatilis × M. chrysops)

Graded levels of l-methionine and l-cysteine were fed to juvenile hybrid striped bass (Morone saxatilis × M. chrysops) to determine the requirement for total sulfur amino acids and the ability of cysteine to spare the requirement. Reproducible mortality occurred in fish fed 0.4% sulfur amino acids (0.37% methionine + 0.03% cysteine), but not in other treatment groups in the same experimental system. Based on weight gain, feed efficiency, and serum methionine concentrations, the minimum requirement appeared to be between 0.68 and 0.73% of the dry diet. Dietary cysteine spared up to 40% of the total sulfur amino acid requirement. Histopathological evaluations did not reveal the causes of mortality in fish fed 0.4% sulfur amino acids. This requirement estimate is on the low end of the established ranges for fish. The ability of cysteine to spare the sulfur amino acid requirement is similar to other fish.

Anesthetic implications of the child with homocystinuria

We present a 7 1 2 - year-old girl with type I homocystinuria who required anesthetic care during repair of bilateral ectopia lentis. Homocystinuria is an autosomal recessive disorder and represents the second most common error of amino acid metabolism. Defective function of the enzyme cystathionine synthetase results in elevated serum levels of methionine and homocysteine. Physiologic disturbances include an increased risk of thromboembolic events and hypoglycemia. Perioperative measures to prevent thromboembolic events include diet therapy to lower serum methionine and homocysteine levels, adequate preoperative hydration, maintenance of intraoperative cardiac output, and the use of pneumatic stockings to prevent peripheral stagnation of blood. Hypoglycemia is thought to be due to alterations in insulin release associated with high levels of circulating sulfur-containing amino acids such as methionine. Perioperative measures include normalization of serum methionine and administration of exogenous glucose during periods of fasting.

DNA cleavage by metabolites of butylated hydroxytoluene.

The effect of butylated hydroxytoluene (BHT) and its metabolites on DNA cleavage in vitro was studied with supercoiled plasmid DNA, pUC18, by agarose gel electrophoresis. Among several BHT metabolites, 2,6-di-t-butyl-p-benzoquinone (BHT-quinone) caused cleavage of supercoiled DNA (form I) at a concentration as low as 1 x 10(-6) M. The relative amount of linear form (form III) was increased with increasing concentration of BHT-quinone. 2,6-Di-t-butyl-4-hydroperoxy-4-methyl-2,5-cyclohexadienone (BHT-peroxyquinol) and 3,5-di-t-butyl-4-hydroxybenzaldehyde (BHT-CHO) also cleaved DNA, but to a lesser extent than BHT-quinone. No DNA cleavage was detected by BHT, 2,6-di-t-butyl-4-hydroxymethyl phenol (BHT-OH), 3,5-di-t-butyl-4-hydroxybenzoic acid (BHT-COOH), 2,6-di-t-butyl-4-hydroxy-4-methyl-2,5-cyclohexadienone (BHT-quinol) or 2,6-di-t-butyl-4-methylene-2,5-cyclohexadienone (BHT-quinone methide). The DNA cleavage by BHT-quinone was inhibited by oxygen radical scavengers including superoxide dismutase (SOD), catalase, polyethylene glycol, t-butyl alcohol, dimethyl sulfoxide, sodium azide, sodium benzoate, bovine serum albumin and methionine, while it was enhanced by the addition of FeCl2. The production of superoxide radical in a solution of BHT-quinone was confirmed by cytochrome c reduction assay. Superoxide was not produced by BHT or other BHT metabolites except for BHT-quinone. These results suggest that BHT-quinone, one of the principal metabolites of BHT, cleaves DNA strands via its generation of oxygen radicals.(ABSTRACT TRUNCATED AT 250 WORDS)

A comparison of fasting serum amino acid profiles of young and elderly subjects.

The fasting serum amino acid profile in 37 healthy young women and men (30-35 years) was compared with the fasting profile in 30 institutionalized elderly women and men (80-89 years), an ambulatory, self-fed senior-residence group. Levels of serum lysine, leucine, methionine, valine, and total essential amino acids were significantly lower in the older group than in the younger group; however, citrulline and hydroxyproline were significantly higher in the older group compared to the younger group. Histidine, threonine, tryptophan, and the ratio of tryptophan to large neutral amino acids (isoleucine, leucine, phenylalanine, tyrosine, and valine) were also generally lower in the older group than in the younger group, while the difference due to age was more pronounced in the females compared to males. The essential/nonessential amino acid ratio was lower in females compared to males.

Structural and Biochemical Changes in a Spinal Myelinopathy in Twelve English Foxhounds and Two Harriers

Degenerative lesions in the spinal cord white matter of 12 English Foxhounds and two Harriers between 3 and 6 years old were associated with a diet composed mainly of ruminant stomachs. Lesions were present throughout the length of the spinal cord and were more severe in ventral and lateral columns than in dorsal columns. Degenerate fibers were accompanied by astrocytic proliferation. Changes suggestive of a primary myelinopathy included vacuolated myelin sheaths around apparently intact axons and thick-thin transitions in myelin sheath thickness. Mixed sensory and motor peripheral nerves and muscle histochemical fiber type profiles appeared normal. Similarities were noted with the changes described in subacute combined degeneration of the spinal cord in human beings, a neuropathy caused by methionine and methylation deficiency in patients with vitamin B12 deficiency. Mean serum methionine levels were significantly lower (P greater than 0.01) and mean liver methionine synthetase levels were significantly greater (P greater than 0.01) in affected dogs restored to a balanced diet than in age-matched controls maintained on the balanced diet. The elevated methionine synthetase levels possibly reflected compensatory reactions to the associated dietary change.

Malnutrition in Hemodialysis Patients

Twenty-nine hemodialysis patients were studied to evaluate different laboratory measures in assessing muscle protein stores and detecting protein malnutrition. Arm muscle circumference (AMC) was used as a reference for somatic protein stores. AMC correlated with serum complement C3, plasma histidine, isoleucine, leucine, methionine and threonine concentrations as well as with body mass index. The lowest quartile of AMC was found most reliably by measuring plasma methionine, histidine, leucine and isoleucine concentrations. Protein malnutrition was detected in five patients (17%). They had significantly lower serum prealbumin and plasma leucine concentrations than the others. In the follow-up of 3 years every malnourished patient died, three from septic infection. Of the others only seven died, none from infection. The analysis of plasma essential amino acid and serum prealbumin concentrations had an important role in assessing muscle protein stores as well as the protein nutrition status in hemodialysis patients. Patients with malnutrition should be detected in view of their unfavourable prognosis.

Effect of Short-Term Essential Amino Acid-Containing Dialysate in Young Children on CAPD

In young children on CAPD, hypoproteinemia and malnutrition are often observed. We used essential amino acid-containing dialysate (EAAD) to assess short-term effectiveness on serum amino acid concentrations in young children undergoing CAPD. EAAD consisted of a 540 ml, 1.5% glucose-containing dialysate and 100 ml of 7.4% essential amino acid (EAA) solution. Aside from methionine, all serum EAA rose during the 6 hour peritoneal dialysis cycle using EAAD, peaking at about 200% of pre-treatment level one hour after start of treatment. They then returned to near pre-treatment levels at the end of the cycle. However, serum methionine increased 680% of pre-treatment level, one hour after start and 390% at the end of the cycle. In the serum non-EAA tyrosine, which showed low levels in patients with chronic renal failure, increased after EAAD treatment. Other non-EAA, most of which showed increased levels in patients with chronic renal failure, decreased after EAAD treatment. These changes in serum amino acids suggest that EAA, absorbed from EAAD, may have increased uptake of non-EAA in protein synthesis. This may improve the nutritional status of young children on CAPD.

Taurine concentrations in serum of critically injured patients and age- and sex-matched healthy control subjects

Sulfur-containing amino acid concentrations were measured in serum of nine previously healthy adult trauma patients after 7 d of taurine-free parenteral nutrition and were compared with fasting serum concentrations in age- and sex-matched healthy subjects. Serum taurine concentrations in the patient group were depressed by 60% relative to control values (P less than 0.05) whereas serum methionine concentrations were significantly higher in the patient group. Serum cysteine concentrations were not significantly different between the two groups. This early decline in serum taurine concentrations in unsupplemented trauma patients suggests that taurine may be essential in the post-injury state.

Effects of intraperitoneal transplantation of microcarrier-attached hepatocytes on D-galactosamine-induced acute liver failure in rats

A study was conducted to investigate morphologic as well as metabolic characteristics of microcarrier-attached hepatocytes in culture, and also to evaluate the effect of intraperitoneal transplantation of the microcarrier-attached hepatocytes on acute hepatic failure in rats induced by D-galactosamine (GalN). Rat hepatocytes were isolated by collagenase perfusion, and cultured on collagen-coated microcarriers. Protein synthesis estimated by [14C] leucine incorporation was four-fold higher in microcarrier culture than in cell suspension. The rates of albumin, transthyretin and bile acid syntheses in hepatocytes cultured on microcarriers were similar to those in monolayer culture. When microcarrier-attached hepatocytes were intraperitoneally transplanted into rats with Galn-induced acute liver failure, a marked improvement in survival rate was observed as compared with control rats which received injections of microcarriers alone (80% vs 0% beyond 6 days of transplantation). Mean serum glutamate oxaloacetate transaminase (SGOT), serum glutamate pyruvate transaminase (SGPT), methionine and glucose levels were similar in both groups, while serum bilirubin and ammonia levels were lower (P less than 0.1, P less than 0.05) in rats transplanted with the microcarrier-attached hepatocytes. Immunohistochemical examinations revealed that the transplanted hepatocytes around microcarriers had albumin synthesis activity, whereas almost no albumin synthesis was demonstrated in recipient liver. In conclusion, intraperitoneal transplantation of the microcarrier-attached hepatocytes will provide sufficient metabolic support, representing detoxication of ammonia (and presumably bilirubin) and synthesis of albumin, to allow GalN-damaged liver function to restore. Microcarrier culture of isolated hepatocytes seems to be one of the most appropriate tools for an artificial liver support.

Plasma amino acid patterns in very low birth weight infants during parenteral nutrition

Plasma amino acids were measured in eight very low birth weight infants (less than or equal to 1000 gm) before and after infusion of parenteral alimentations with Freamine III. Significant elevation in serum threonine, valine, isoleucine, methionine, serine, proline, glycine and ornithine was noted after twenty four hours of infusion. On the other hand, significant decreases in taurine and tyrosine levels were noted. Our study suggests that current solution is not optimal for premature neonates and the amount of protein administered during the first week in infants weighing less than or equal to 1000 gm should be decreased from the recommended 2.5-3.0 gm/kg/day.

DNA cleavage by phenylhydroquinone: The major metabolite of a fungicide o-phenylphenol

The interaction of o-phenylphenol (OPP) and its metabolites with DNA was examined. As a model system, the reactivities of OPP and its metabolites with DNA were studied by using pUC18 DNA. The major metabolite formed in vitro from OPP by mixed function oxidase was phenylhydroquinone (PHQ). This result corresponds to the findings that PHQ in the form of glucuronide conjugate was the main product detected in bladder of OPP fed rats in vivo. When supercoiled pUC18 DNA (form I) was incubated with PHQ at concentrations from 1 × 10 −6 M to 2 × 10 −1 M, DNA strand scission by PHQ was observed at a concentration as low as 1 × 10 −5 M and the amount of linear form (form III) increased with increasing PHQ concentration. PHQ causes DNA strand scission. The DNA cleavage by OPP and phenyl- p-benzoquinone (PBQ) was barely detectable. The DNA cleavage by PHQ was inhibited by superoxide dismutase (SOD), catalase and several oxygen radical scavengers such as polyethylene glycol, tert-butanol, dimethyl sulfoxide, sodium azide, sodium benzoate, bovine serum albumin and methionine. The production of superoxide radical from PHQ was confirmed by cytochrome c reduction assay. These results indicate that the oxygen radicals such as superoxide, hydroxyl radicals and some others generated in the process of oxidation of PHQ in aqueous solution are responsible for the DNA cleavage. In order to identify the sites of cleavage of DNA by PHQ, a 5′-end 32P-labeled 206 base-pair EcoRI- BglI fragment of pUC18 DNA was incubated with PHQ. The DNA was then analyzed by sequencing gel electrophoresis followed by autoradiography. When the DNA was incubated with PHQ and further treated with piperidine, cleavage was detected relatively more frequently at guanine residues. The attack seemed to occur at guanine residues in general, but was not restricted to guanines with specific residues in the vicinity.

Whole Rat Embryos Require Methionine for Neural Tube Closure when Cultured on Cow Serum

Headfold-stage rat embryos, when cultured on cow serum without supplemental methionine, failed to close their neural tubes, lacked eyes and branchial arches, were abnormally shaped and were reduced in protein content compared to methionine-supplemented embryos. Methionine was essential during the first 18 h of culture, a period in which neural tube closure was initiated in supplemented cultures. All cow serum samples tested were found to require methionine addition, and the methionine was not replaced by other amino acids or vitamins, including folate. Methionine was not toxic to cultured rat embryos at concentrations up to at least 500 micrograms/ml. Analyses of serum free amino acids revealed lower levels of free methionine in cow serum compared to rat serum, and cow serum proteins contained less methionine relative to other amino acids than did rat serum proteins. Dialysis of cow serum reduced but did not eliminate the requirement for methionine. This suggested either that the free amino acids of cow serum were imbalanced or that a dialyzable component in serum interfered with the availability/utilization of methionine. Dietary supplementation of cows with rumen-protected DL-methionine increased the serum methionine level, and serum drawn from supplemented cows supported normal rat embryo development without additional methionine.

Der Einfluß von Dl-Methionin, N-Hydroxymethyl-DL- Methionin-Calciumsalz (Mepron(R)) und DL-Methionin- Hydroxyanalog-Calciumsalz (MHA(R)) auf den N-Ansatz von Kälbern

Digestibility and N balance experiments with calves showed that the methionine requirement of the host animal is covered by microbial protein synthesis even in the already ruminant animal. A supplement of the ration with DL methionine is only effective when it is provided in milk replacer. The MHA substitution of the ration has only little influence on N retention, independent of its supply in milk replacer or concentrated feed. MEPRON proved to be the most effective methionine derivative. As supplement both with the fluid feed and with the concentrated feed this compound shows exclusively positive results even in already ruminant calves with regard to live weight gain, N retention and the content of free methionine in the blood serum of the test animals.

Quantitation of total homocysteine, total cysteine, and methionine in normal serum and urine using capillary gas chromatography-mass spectrometry

Total homocysteine, total cysteine, and methionine have been extracted and partially purified from serum and urine using reduction with 2-mercaptoethanol followed by cation-exchange chromatography and anion-exchange chromatography. The t-butyldimethylsilyl derivatives were prepared and analyzed using capillary gas chromatography-mass spectrometry with selected ion monitoring. The addition of dl-[3,3,3′,3′,4,4,4′,4′- 2H 8]homocystine, dl-[3,3,3′,3′- 2H 4]cystine, and l-[ methyl- 2H 3]methionine to the starting samples prior to the reduction of all disulfides, including the deuterated internal standards, with 2-mercaptoethanol makes it possible to quantitate all three amino acids. Normal ranges for total homocysteine, total cysteine, and methionine have been determined in human and rat serum and in human urine.

Lactational and Systemic Responses to the Supplementation of Protected Methionine in Soybean Meal Diets

Twenty-seven Holstein cows (14 primiparous and 13 multiparous) were randomly assigned to diets containing soybean meal without or with 15g of added DL-methionine daily, provided as 50g of ruminally protected methionine product, during wk 4 through 16 postpartum. Cows were fed a 15.3% crude protein total mixed diet of (dry matter basis) 30% corn silage, 15% alfalfa hay, and 55% concentrate mix. Yields of milk (32.9 and 35.2 kg/d), 4% fat-corrected milk (27.8 and 29.5 kg/d), and solids-corrected milk (28.5 and 30.1 kg/d) were higher for cows fed supplemental methionine. Milk protein percentage (2.99 and 3.06) was increased with supplemental methionine, while the percentage of fat (2.96 and 3.00), solids-not-fat (8.69 and 8.73), and total solids (11.67 and 11.71) were similar among diets. Dry matter intake (19.3 and 21.3 kg/d) was higher with methionine supplementation. Ruminal pH, volatile fatty acids, ammonia, and serum urea were generally unaffected by methionine supplementation. Concentrations of methionine in arterial and venous serum were elevated slightly by methionine supplementation, but the first-limiting amino acid for milk production, as calculated by several methods, was not changed.

A Method for the Determination of Methionine in Human Serum by High-Performance Liquid Chromatography with Electrochemical Detection

Abstract We have found remarkable enhancement of electrochemical response for the analysis of methionine by use of a glassy carbon electrode preanodized in a 0.2 M phosphate buffer (KH2PO4−KOH, pH 6.5) at +1.9 V vs. Ag/AgCl for 2 min. On the basis of this finding, we have developed a method for the determination of methionine in human serum by reversed-phase high-performance liquid chromatography with electrochemical detection using the electrochemically pretreated glassy carbon electrode. The minimum detectable quantity of methionine has been found to be about 1 ng.