Modulation of Methyl Group Metabolism by Streptozotocin-induced Diabetes and All-trans-retinoic Acid

Abstract

The hepatic enzyme glycine N-methyltransferase (GNMT) plays a major role in the control of methyl group and homocysteine metabolism. Because disruption of these vital pathways is associated with numerous pathologies, understanding GNMT control is important for evaluating methyl group regulation. Recently, gluconeogenic conditions have been shown to modulate homocysteine metabolism and treatment with glucocorticoids and/or all-trans-retinoic acid (RA)-induced active GNMT protein, thereby leading to methyl group loss. This study was conducted to determine the effect of diabetes, alone and in combination with RA, on GNMT regulation. Diabetes and RA increased GNMT activity 87 and 148%, respectively. Moreover, the induction of GNMT activity by diabetes and RA was reflected in its abundance. Cell culture studies demonstrated that pretreatment with insulin prevented GNMT induction by both RA and dexamethasone. There was a significant decline in homocysteine concentrations in diabetic rats, owing in part to a 38% increase in the abundance of the transsulfuration enzyme cystathionine β-synthase; treatment of diabetic rats with RA prevented cystathionine β-synthase induction. A diabetic state also increased the activity of the folate-independent homocysteine remethylation enzyme betaine-homocysteine S-methyltransferase, whereas the activity of the folate-dependent enzyme methionine synthase was diminished 52%. In contrast, RA treatment attenuated the streptozotocin-mediated increase in betaine-homocysteine S-methyltransferase, whereas methionine synthase activity remained diminished. These results indicate that both a diabetic condition and RA treatment have marked effects on the metabolism of methyl groups and homocysteine, a finding that may have significant implications for diabetics and their potential sensitivity to retinoids. The hepatic enzyme glycine N-methyltransferase (GNMT) plays a major role in the control of methyl group and homocysteine metabolism. Because disruption of these vital pathways is associated with numerous pathologies, understanding GNMT control is important for evaluating methyl group regulation. Recently, gluconeogenic conditions have been shown to modulate homocysteine metabolism and treatment with glucocorticoids and/or all-trans-retinoic acid (RA)-induced active GNMT protein, thereby leading to methyl group loss. This study was conducted to determine the effect of diabetes, alone and in combination with RA, on GNMT regulation. Diabetes and RA increased GNMT activity 87 and 148%, respectively. Moreover, the induction of GNMT activity by diabetes and RA was reflected in its abundance. Cell culture studies demonstrated that pretreatment with insulin prevented GNMT induction by both RA and dexamethasone. There was a significant decline in homocysteine concentrations in diabetic rats, owing in part to a 38% increase in the abundance of the transsulfuration enzyme cystathionine β-synthase; treatment of diabetic rats with RA prevented cystathionine β-synthase induction. A diabetic state also increased the activity of the folate-independent homocysteine remethylation enzyme betaine-homocysteine S-methyltransferase, whereas the activity of the folate-dependent enzyme methionine synthase was diminished 52%. In contrast, RA treatment attenuated the streptozotocin-mediated increase in betaine-homocysteine S-methyltransferase, whereas methionine synthase activity remained diminished. These results indicate that both a diabetic condition and RA treatment have marked effects on the metabolism of methyl groups and homocysteine, a finding that may have significant implications for diabetics and their potential sensitivity to retinoids. Methyl group and folate-dependent one-carbon metabolism are interrelated pathways that provide for the subsequent transmethylation of various molecules using S-adenosylmethionine (AdoMet) 1The abbreviations used are: AdoMet, S-adenosylmethionine; GNMT, glycine N-methyltransferase; AdoHcy, S-adenosylhomocysteine; 5-CH3-THF, 5-methyltetrahydrofolate; MS, methionine synthase; BHMT, betaine-homocysteine S-methyltransferase; CBS, cystathionine β-synthase; RA, all-trans-retinoic acid; DEX, dexamethasone; STZ, streptozotocin; HPLC, high pressure liquid chromatography.1The abbreviations used are: AdoMet, S-adenosylmethionine; GNMT, glycine N-methyltransferase; AdoHcy, S-adenosylhomocysteine; 5-CH3-THF, 5-methyltetrahydrofolate; MS, methionine synthase; BHMT, betaine-homocysteine S-methyltransferase; CBS, cystathionine β-synthase; RA, all-trans-retinoic acid; DEX, dexamethasone; STZ, streptozotocin; HPLC, high pressure liquid chromatography. (1Wagner C. Bailey L.B. Folate in Health and Disease. Marcel Dekker, New York1995: 23-42Google Scholar). An adequate supply of methyl groups via the diet or the folate-dependent one-carbon pool and regulation of these pathways are essential in preventing associated pathologies such as cancer, vascular disease, and neural tube defects (2Ross S.A. Ann. N. Y. Acad. Sci. 2003; 983: 197-207Crossref PubMed Scopus (144) Google Scholar, 3Kang S.-S. Wong P.W.K. Malinow M.R. Annu. Rev. Nutr. 1992; 12: 279-298Crossref PubMed Scopus (711) Google Scholar, 4Scott J.M. Kirkie P.N. Weir D.G. Annu. Rev. Nutr. 1990; 10: 277-295Crossref PubMed Scopus (56) Google Scholar). Glycine N-methyltransferase (GNMT) (EC 2.1.1.20) is an abundant, tissue-specific protein that plays a key role in the regulation of hepatic methyl group metabolism by the enzymatic conversion of glycine and AdoMet to S-adenosylhomocysteine (AdoHcy) and sarcosine (1Wagner C. Bailey L.B. Folate in Health and Disease. Marcel Dekker, New York1995: 23-42Google Scholar, 5Kerr S.J. J. Biol. Chem. 1972; 247: 4248-4252Abstract Full Text PDF PubMed Google Scholar, 6Yeo E.-J. Wagner C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 210-214Crossref PubMed Scopus (112) Google Scholar). GNMT functions to optimize the AdoMet/AdoHcy ratio, an indicator of transmethylation potential, because AdoHcy is a potent inhibitor of most AdoMet-dependent methyltransferases (5Kerr S.J. J. Biol. Chem. 1972; 247: 4248-4252Abstract Full Text PDF PubMed Google Scholar, 7Cantoni G.L. Chiang P.K. Cavallini D. Gaull G.E. Zappia V. Natural Sulfur Compounds. Plenum Press, New York1980: 67-80Crossref Google Scholar). Following hydrolysis of AdoHcy, the resulting homocysteine can undergo remethylation to methionine or be irreversibly catabolized by the transsulfuration pathway. Folate-dependent remethylation occurs with the donation of a methyl group by 5-methyltetrahydrofolate (5-CH3-THF) through the action of B12-dependent methionine synthase (MS) (8Shane B. Stokstad E.L.R. Annu. Rev. Nutr. 1985; 5: 115-141Crossref PubMed Scopus (268) Google Scholar). In hepatic tissue, betaine derived from the oxidation of choline can also serve as a folate-independent source of methyl groups for homocysteine remethylation via the enzyme betaine-homocysteine S-methyltransferase (BHMT). Transsulfuration to cysteine occurs through the activity of two vitamin B6-dependent enzymes, cystathionine β-synthase (CBS) and γ-cystathionase. The regulation of hepatic GNMT represents an important mechanism for controlling both the folate-dependent supply of methyl groups as well as their utilization in AdoMet-dependent transmethylation reactions. GNMT activity is regulated in response to changes in methyl group status as the result of allosteric inhibition of 5,10-methylenetetrahydrofolate reductase by AdoMet (9Jencks D.A. Matthews R.G. J. Biol. Chem. 1987; 262: 2485-2493Abstract Full Text PDF PubMed Google Scholar, 10Kutzbach C. Stokstad E.L.R. Biochim. Biophys. Acta. 1967; 250: 459-477Crossref Scopus (272) Google Scholar) and inhibition of GNMT by 5-CH3-THF (11Wagner C. Briggs W.T. Cook R.J. Biochem. Biophys. Res. Commun. 1985; 127: 746-752Crossref PubMed Scopus (140) Google Scholar, 12Wagner C. Decha-Umphai W. Corbin J. J. Biol. Chem. 1989; 264: 9638-9642Abstract Full Text PDF PubMed Google Scholar), the enzymatic product of 5,10-methylenetetrahydrofolate reductase. This allows methyl groups to be conserved under conditions of deficiency by decreasing GNMT activity, whereas elevations in its activity function to dispose of excess methyl groups. In addition to allosteric control, phosphorylation of GNMT represents another posttranslational mechanism to increase the activity of GNMT and regulate methyl group metabolism (12Wagner C. Decha-Umphai W. Corbin J. J. Biol. Chem. 1989; 264: 9638-9642Abstract Full Text PDF PubMed Google Scholar). Recent studies have identified both nutritional and hormonal factors that alter methyl group metabolism by targeting the key enzymes involved. All-trans-retinoic acid (RA) has been shown to induce hepatic GNMT activity and protein abundance, resulting in the loss of methyl groups for subsequent transmethylation reactions (13McMullen M.H. Rowling M.J. Ozias M.K. Schalinske K.L. Arch. Biochem. Biophys. 2002; 401: 73-80Crossref PubMed Scopus (27) Google Scholar, 14Ozias M.K. Schalinske K.L. J. Nutr. 2003; 133: 4090-4094Crossref PubMed Scopus (30) Google Scholar, 15Rowling M.J. McMullen M.H. Schalinske K.L. J. Nutr. 2002; 132: 365-369Crossref PubMed Scopus (77) Google Scholar, 16Rowling M.J. Schalinske K.L. J. Nutr. 2001; 131: 1914-1917Crossref PubMed Scopus (29) Google Scholar). Moreover, dexamethasone (DEX) was just as effective as RA in the induction of GNMT activity in rat liver and hepatoma cells, and the co-administration of both DEX and RA induced GNMT in an additive fashion (17Rowling M.J. Schalinske K.L. J. Nutr. 2003; 133: 3392-3398Crossref PubMed Scopus (41) Google Scholar). A diabetic state characterized by insufficient circulating concentrations of insulin and elevated levels of the counter-regulatory hormones glucagon and glucocorticoids has also been reported to alter enzymes involved in methyl group metabolism. An elevation in the activity of GNMT has been reported in alloxan-induced diabetic sheep and rats (6Yeo E.-J. Wagner C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 210-214Crossref PubMed Scopus (112) Google Scholar, 18Xue G.-P. Snoswell A.M. Biochem. Int. 1985; 10: 897-905PubMed Google Scholar); however, little is known about the mechanistic basis for this increase. Brosnan and co-workers have shown that under diabetic conditions, the catabolism of homocysteine was enhanced by transcriptional regulation of CBS, and these changes were prevented by treatment with insulin (19Jacobs R.L. House J.D. Brosnan M.E. Brosnan J.T. Diabetes. 1998; 47: 1967-1970Crossref PubMed Scopus (170) Google Scholar, 20Jacobs R.L. Stead L.M. Brosnan M.E. Brosnan J.T. J. Biol. Chem. 2001; 276: 43740-43747Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar, 21Ratnam S. Maclean K.N. Jacobs R.L. Brosnan M.E. Kraus J.P. Brosnan J.T. J. Biol. Chem. 2002; 277: 42912-42918Abstract Full Text Full Text PDF PubMed Scopus (124) Google Scholar). Although we have shown that DEX can significantly alter methyl group and homocysteine metabolism (17Rowling M.J. Schalinske K.L. J. Nutr. 2003; 133: 3392-3398Crossref PubMed Scopus (41) Google Scholar), the focus in this study was to evaluate these pathways in a diabetic rat model, alone and in combination with RA, as well as determine the effect of insulin on preventing these alterations. Chemicals—Reagents were obtained as follows: S-adenosyl-l-[methyl-3H]methionine, PerkinElmer Life Sciences; chemiluminescence Western blotting detection reagents, Amersham Biosciences; S-adenosyl-l-methionine and streptozotocin, Sigma; protease inhibitors and RA, Calbiochem; goat anti-mouse IgG horseradish peroxidase, goat anti-rabbit IgG horseradish peroxidase, and rabbit anti-sheep IgG horseradish peroxidase, Southern Biotechnology (Birmingham, AL). GNMT and CBS antibodies were kindly provided by Yi-Ming Chen, National Yang-Ming University, Taipei, Taiwan (22Liu H.-H. Dhen K.-H. Shin Y.-P. Lui W.-Y. Wong F.-H. Dhen Y.-M.A. J. Biomed. Sci. 2003; 10: 87-97Crossref PubMed Google Scholar), and Jan Kraus, University of Colorado Health Sciences Center, Denver, CO, respectively. All other chemicals were of analytical grade. Animals—All animal experiments were approved by and conducted in accordance with Iowa State University Laboratory Animal Resources Guidelines. Male Sprague-Dawley rats (Harlan Sprague-Dawley, Indianapolis, IN) (125–149 g) were housed in individual plastic cages in a room with a 12-h light/dark cycle and allowed free access to food and water. Rats were randomly assigned to treatment groups (6 rats per group) and acclimated to the control diet (17Rowling M.J. Schalinske K.L. J. Nutr. 2003; 133: 3392-3398Crossref PubMed Scopus (41) Google Scholar) for 5 days. Following the acclimation period, rats received a single intraperitoneal injection of either streptozotocin (STZ, 60 mg/kg body weight) in the vehicle (10 mm citrate buffer, pH 4.5) or the vehicle alone. The following day animals received a daily oral dose of RA (30 μmol/kg body weight) or vehicle (corn oil) alone. Following a 5-day treatment period with RA, rats were anesthetized and whole blood samples were collected via cardiac puncture. An aliquot of whole blood was used to assess blood glucose concentrations using a commercial kit (Sigma), whereas the remaining blood was centrifuged at 4,000 × g for 6 min, and the resulting plasma fraction was stored at –20 °C for subsequent analysis of homocysteine concentrations. Liver samples were removed and homogenized in 4 volumes of ice-cold phosphate-buffered (10 mm, pH 7.0) sucrose (0.25 m) containing 1 mm EDTA, 1 mm and mm Following at × g for min, were stored at °C with 1 mm for subsequent enzymatic and protein abundance protein concentrations were on the of Biochem. PubMed Scopus Google Scholar) using a commercial kit and as a liver samples were homogenized in volumes of acid for the of AdoMet and Cell and hepatoma were obtained from the All were in in a with and a of °C were were in essential containing and were in containing as well as and Cell received to the of and were with either a vehicle of RA, DEX, or both RA and DEX as (17Rowling M.J. Schalinske K.L. J. Nutr. 2003; 133: 3392-3398Crossref PubMed Scopus (41) Google Scholar). A group of and was with insulin for to the addition of RA and/or Following a period, were using mm EDTA, with and on in a containing mm mm mm mm 5 mm EDTA, 1 mm mm and mm and Following at × g for min, were stored at °C for analysis of GNMT protein abundance. GNMT enzymatic activity of GNMT was using the by Cook and Wagner R.J. Wagner C. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar) with and in The of 1 mm 5 mm 1 and was with of protein by at °C for acid was to the by the addition of and × g) to of the were removed for liquid of GNMT and CBS protein abundance was using as (17Rowling M.J. Schalinske K.L. J. Nutr. 2003; 133: 3392-3398Crossref PubMed Scopus (41) Google Scholar). A was used to the of Following were to a and at 4 °C with a of the GNMT (22Liu H.-H. Dhen K.-H. Shin Y.-P. Lui W.-Y. Wong F.-H. Dhen Y.-M.A. J. Biomed. Sci. 2003; 10: 87-97Crossref PubMed Google Scholar). The was with goat anti-mouse IgG horseradish for 1 at room CBS protein abundance was using a as to the CBS A of the CBS was was by with a goat anti-rabbit IgG horseradish GNMT and CBS protein abundance were with chemiluminescence and to analysis was using both GNMT and CBS, samples from treatment group were randomly for analysis on a single of AdoMet and AdoHcy acid were centrifuged at × g for min, and the resulting were and to a to AdoMet and AdoHcy D. J. 1985; PubMed Scopus Google Scholar, K.L. PubMed Scopus Google Scholar). AdoMet and AdoHcy were and by and detection using a containing in 5 mm acid at of and activity was as by J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The 5 mm betaine and of The activity of was as Weir J.M. Sci. 1985; PubMed Scopus Google Scholar). The containing mm, pH AdoMet (10 homocysteine and mm, was to liver and for 1 at was to the and the was to a of were collected for subsequent liquid both the and homocysteine was daily from a plasma homocysteine concentrations were using and detection S. J. PubMed Scopus Google Scholar). in was to of plasma samples and at 4 °C for The was with ice-cold acid containing 1 mm Following at × g for 5 min, were to a containing pH and was to the plasma samples to as an were a in a of in was used for The of treatment group were to a analysis of the analysis of was significant were using significant in and of RA for diabetic rats was and of whereas RA treatment was effect induced a diabetic state in rats as blood glucose concentrations were elevated with control Rats with RA significant effect on circulating levels of glucose with control and RA attenuated the by diabetic and blood glucose concentrations in rats with and/or in a Diabetes and RA rats a increase in hepatic GNMT activity with the control RA increased GNMT activity with control rats and was significantly the of induction in GNMT activity was in the rats both and RA this increase was significantly from RA treatment alone. The changes in GNMT activity in diabetic and rats were also reflected in the abundance of the protein, a and increase was Moreover, co-administration of and RA significantly induced GNMT protein abundance to a either treatment alone. both RA treatment and a diabetic condition modulate the of active GNMT in rat of GNMT by RA and/or DEX was by reported (17Rowling M.J. Schalinske K.L. J. Nutr. 2003; 133: 3392-3398Crossref PubMed Scopus (41) Google Scholar), both RA and DEX alone or in co-administration in induction of the GNMT protein in rat hepatoma with insulin for to treatment with RA and/or DEX prevented GNMT induction. induction of GNMT in was also prevented by however, RA was an effective to induce GNMT in these and CBS homocysteine concentrations were significantly in both groups of rats of RA Although we have that RA alone was effective at circulating homocysteine levels M.K. Schalinske K.L. J. Nutr. 2003; 133: 4090-4094Crossref PubMed Scopus (30) Google Scholar), the under these conditions The by diabetic rats to be in to a 38% increase in the abundance of CBS, the enzyme in the catabolism of homocysteine CBS activity is regulated and studies conducted in be to evaluate the transsulfuration pathway. RA treatment of diabetic rats prevented CBS induction plasma homocysteine concentrations remained diminished abundance of CBS was elevated in diabetic rats and following treatment with samples from the rats as for 1 were used to determine CBS abundance by Western analysis as under A is shown the are with are significantly and RA addition to the plasma homocysteine levels also to changes in the activity of enzymes involved in homocysteine activity was significantly increased in diabetic rats, whereas the activity of was RA significantly alter either was a for increased activity, to M.K. Schalinske K.L. J. Nutr. 2003; 133: 4090-4094Crossref PubMed Scopus (30) Google Scholar). RA was effective at the activity of in diabetic rats, the in of remethylation enzymes methionine synthase and in the of rats with and/or of of in a in the of AdoMet and AdoHcy by RA diabetic condition in a significant increase in the hepatic concentrations of AdoMet and AdoHcy Although RA treatment significantly alter AdoMet and AdoHcy concentrations in rats, their in diabetic Because AdoMet and AdoHcy are allosteric of a of enzymes involved in methyl and homocysteine is to determine the effect of these changes in diabetic rats in the of using concentrations of AdoMet and AdoHcy from rats with and/or of in a Methyl homocysteine, and the folate-dependent one-carbon pool interrelated that are important in and Because GNMT is a vital protein that functions to control the supply and utilization of methyl is to nutritional and/or hormonal factors modulate its demonstrated that RA and DEX were to induce active GNMT in rat liver and in hepatoma (17Rowling M.J. Schalinske K.L. J. Nutr. 2003; 133: 3392-3398Crossref PubMed Scopus (41) Google Scholar). we have these by that a diabetic by treatment with STZ, also has a on the activity of hepatic rat studies using alloxan-induced diabetes reported in GNMT activity (6Yeo E.-J. Wagner C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 210-214Crossref PubMed Scopus (112) Google Scholar), whereas treatment with glucagon were R.L. Stead L.M. Brosnan M.E. Brosnan J.T. J. Biol. Chem. 2001; 276: 43740-43747Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar); however, the mechanistic basis for this increase in GNMT activity has been A of studies that for both diabetes and RA, the abundance of GNMT protein was that GNMT was regulated by transcriptional and/or or at the of of the is known regulation of GNMT in a diabetic state is the result of an increase in levels or a of The culture studies shown that pretreatment with insulin prevented GNMT induction by both DEX and be important in studies to this diabetic condition with in methyl group metabolism. the of GNMT to RA and DEX to be have in other studies that DEX was a RA for GNMT induction a of and that GNMT, such as the hepatoma B. J. and is RA, DEX, and/or a diabetic state alter the of studies on the regulation of GNMT activity have to be at the posttranslational by phosphorylation and allosteric inhibition by 5-CH3-THF (11Wagner C. Briggs W.T. Cook R.J. Biochem. Biophys. Res. Commun. 1985; 127: 746-752Crossref PubMed Scopus (140) Google Scholar, 12Wagner C. Decha-Umphai W. Corbin J. J. Biol. Chem. 1989; 264: 9638-9642Abstract Full Text PDF PubMed Google Scholar, Wagner C. Biochem. J. PubMed Scopus Google Scholar, R.J. Wagner C. J. Nutr. 1989; PubMed Scopus Google Scholar). on the known of glucocorticoids and RA with to regulation of is that the induction of GNMT is the result of increased In of this we have that pretreatment of hepatoma with prevented GNMT induction by DEX and RA, and abundance of the protein to changes in its J. and GNMT has been reported to a a in its Y. J. Biochem. 1998; Scopus Google Scholar). is that are to regulation of GNMT we the that in phosphorylation of the protein have a role in GNMT activity, as glucagon and glucocorticoids can their action via in protein concentrations the from AdoMet-dependent transmethylation remethylation to and catabolism via the transsulfuration pathway. has been reported (19Jacobs R.L. House J.D. Brosnan M.E. Brosnan J.T. Diabetes. 1998; 47: 1967-1970Crossref PubMed Scopus (170) Google Scholar, 20Jacobs R.L. Stead L.M. Brosnan M.E. Brosnan J.T. J. Biol. Chem. 2001; 276: 43740-43747Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar), we also that a diabetic state was characterized by a in circulating homocysteine This in homocysteine to an increase in the activity of and the abundance of CBS, the activity of was studies have the to of CBS and whereas the activity of hepatic and were significantly (19Jacobs R.L. House J.D. Brosnan M.E. Brosnan J.T. Diabetes. 1998; 47: 1967-1970Crossref PubMed Scopus (170) Google Scholar, 20Jacobs R.L. Stead L.M. Brosnan M.E. Brosnan J.T. J. Biol. Chem. 2001; 276: 43740-43747Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar, 21Ratnam S. Maclean K.N. Jacobs R.L. Brosnan M.E. Kraus J.P. Brosnan J.T. J. Biol. Chem. 2002; 277: 42912-42918Abstract Full Text Full Text PDF PubMed Scopus (124) Google Scholar). in the of studies is to determine homocysteine catabolism because CBS is regulated by other factors such as AdoMet, a known allosteric of CBS activity J.D. J. Biol. Chem. Full Text PDF PubMed Google Scholar). In of the increase in CBS abundance that we we also that the hepatic of AdoMet was elevated in with studies using rats R.L. Stead L.M. Brosnan M.E. Brosnan J.T. J. Biol. Chem. 2001; 276: 43740-43747Abstract Full Text Full Text PDF PubMed Scopus (57) Google Scholar). in hepatic AdoMet levels may be the result of increased of methionine as is by glucocorticoids B. J.M. PubMed Scopus Google Scholar). An increase in hepatic AdoMet concentrations also be to 5-CH3-THF levels as a result of allosteric inhibition of 5,10-methylenetetrahydrofolate reductase (9Jencks D.A. Matthews R.G. J. Biol. Chem. 1987; 262: 2485-2493Abstract Full Text PDF PubMed Google Scholar, 10Kutzbach C. Stokstad E.L.R. Biochim. Biophys. Acta. 1967; 250: 459-477Crossref Scopus (272) Google Scholar). The of 5-CH3-THF as an for GNMT (11Wagner C. Briggs W.T. Cook R.J. Biochem. Biophys. Res. Commun. 1985; 127: 746-752Crossref PubMed Scopus (140) Google Scholar), may be a for the increase in its activity in diabetic An of was that the of RA to diabetic rats prevented CBS induction and AdoMet and the activity of BHMT, whereas activity was In to diabetes, RA alone on CBS abundance or the activity of and MS, we have in studies that RA increase provided for a period of M.K. Schalinske K.L. J. Nutr. 2003; 133: 4090-4094Crossref PubMed Scopus (30) Google Scholar). be that the activity and abundance of GNMT remained elevated in rats with This the for under in conditions to determine the of these The implications of these results for are as diabetes and its are a significant and the of has in J. A. J. Acad. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). In addition to metabolism of methyl groups and homocysteine is on a of and as well as an adequate source of methyl groups such as methionine and Moreover, is well known that the has a significant of enzymes important in methyl homocysteine, and folate-dependent one-carbon metabolism A. J. Nutr. Biochem. 2003; PubMed Scopus Google Scholar). these and hormonal factors the for understanding the these pathways and most diabetics control their by the of are their Moreover, of the changes in study were demonstrated in a diabetes diabetic model, an increase in hepatic CBS, BHMT, and AdoMet S. Brosnan M.E. Brosnan J.T. J. Scholar). for the GNMT studies using and