Methaemoglobin reductase deficiency in dogs

Erythrocyte methaemoglobin reductase deficiency is described in a toy Alaskan Eskimo dog, a miniature poodle dog and a cocker/poodle cross dog. Blood methaemoglobin contents ranged from 19% to 36% of total haemoglobin, with methaemoglobin reductase values between 13% and 33% of normal. There appeared to be a negative linear correlation between erythrocyte methaemoglobin content and methaemoglobin reductase activity. A single intravenous injection of methylene blue (1 mg/kg body weight) resulted in a dramatic decrease in methaemoglobin content within 1 hour when given to two of the deficient dogs in the present study. Following the disappearance of methylene blue from blood, methaemoglobin content increased linearly at 3.2% and 2.5% per day in these dogs, suggesting that about 3.0% of erythrocyte haemoglobin is normally oxidised to methaemoglobin each day in dogs, as has been estimated in humans. Oral riboflavin therapy was not effective in reducing blood methaemoglobin content in deficient dogs.     

Elemental selenium and glutathione reductase

Selenium is an essential trace element important to several metabolic processes, although selenium in the chemical form of elemental selenium (Se degree) is commonly believed to be biologically inert. Recent data shows that colloidal suspensions of red amorphous elemental selenium are more easily reduced than previously thought, and that such reductions can potentially take place under biological conditions. The enzyme glutathione reductase, already known to be involved in selenium metabolism, is a good candidate to mediate the reduction of colloidal selenium to hydrogen selenide (H2Se), a compound of established biological activity. The implications of this hypothesis include the possibility that elemental selenium may be biologically active at least under some conditions, and that glutathione reductase may function in selenium metabolism primarily by maintaining the glutathione concentration in a reduced state.     

Effects of development and ageing on pulmonary NADPH-cytochrome c reductase, glutathione peroxidase, glutathione reductase and thioredoxin reductase activities in male and female rats

The behaviour of the principal NADPH-consuming detoxification enzymes (NADPH-cytochrome c reductase, glutathione peroxidase-glutathione reductase system, and thioredoxin reductase) was studied during development and senescence of the rat lung. We have also studied the influence of sex on the development and senescent values. The NADPH-cytochrome c reductase activity increases at birth and afterwards remains constant until the 25th day after birth, at which age there is a maximum activity. Its activity decreases during the ageing period in both sexes. The glutathione reductase and thioredoxin reductase activities show significant differences with respect to sex during the adult stage, however during ageing these differences disappear. These enzymes show maximum activity at 25 days after birth, and afterwards the activity decreases continuously until the adult levels are reached. The activity of glutathione reductase is increased during the ageing period, especially in the female rats, however, in senescence the levels of thioredoxin reductase are lower than in the adult stage. The glutathione peroxidase shows a significant difference between both sexes during senescence and in the male its activity in this stage is higher than during development and adulthood.     

Polymorphisms in the methylenetetrahydrofolate reductase and methionine synthase reductase genes and homocysteine levels in Brazilian children

Hyperhomocysteinemia is a risk factor for thrombosis, and methylenetetrahydrofolate reductase (MTHFR) and methionine synthase reductase (MTRR) polymorphisms, folate, and B12 levels could contribute to plasma homocysteine (Hcy) variation. Although well established in adults, few studies have been performed in childhood. In this study, we investigated association of polymorphisms C677T and A1298C in the MTHFR gene and A66G in the MTRR gene with Hcy levels in children. These polymorphisms, as well as Hcy, folate, and vitamin B12 levels were investigated in 220 normal children with ages ranging from 1 to 8 years. Plasma Hcy, folate, and vitamin B12 levels were normal in all children. None of the polymorphisms could be considered an independent risk factor for hyperhomocysteinemia during childhood. The median Hcy levels in 37 children (17%) doubly heterozygous for C677T and A1298C mutations in the MTHFR gene were not different from the other genotypes. However, the association of the different genotypes with Hcy, folate, and vitamin B12 levels demonstrated significant P-values. The folate levels demonstrated a statistically significant decrease (P = 0.0477) from the C677T mutation in the MTHFR gene (TT genotype) when compared to the other groups. Folate was the only independent risk factor for hyperhomocysteinemia. Thus, monitoring the concentrations of folate would be more helpful for evaluating hyperhomocysteinemia and for preventing cardiovascular disease.     

Dihydrofolate reductase from Kaposi's sarcoma-associated herpesvirus

Kaposi's sarcoma-associated herpesvirus (KSHV) is the first human virus known to encode dihydrofolate reductase (DHFR), an enzyme required for nucleotide and methionine biosynthesis. We have studied the purified KSHV-DHFR enzyme in vitro and analyzed its expression in cultured B-cell lines derived from primary effusion lymphoma (PEL), an AIDS-associated malignancy. The amino acid sequence of KSHV-DHFR is most similar to human DHFR (hDHFR), but the viral enzyme contains an additional 23 amino acids at the carboxyl-terminus. The viral DHFR, overexpressed and purified from E. coli, was catalytically active in vitro. The K(m) of KSHV-DHFR for dihydrofolate (FH(2)) was 2.4 microM, which is significantly higher than the K(m) of recombinant hDHFR (rhDHFR) for FH(2) (390 nM). K(m) values for NADPH were similar for the two enzymes, about 1 microM. KSHV-DHFR was inhibited by folate antagonists such as methotrexate (K(i): 200 pM), aminopterin (K(i): 610 pM), pyrimethamine (K(i): 29 nM), trimethoprim (K(i): 2.3 microM), and piritrexim (K(i): 3.9 nM). In all cases, K(i) values for these folate antagonists were higher for KSHV-DHFR than for rhDHFR. The viral enzyme was expressed at levels two- to tenfold higher than hDHFR in PEL cell lines as an early lytic cycle gene. KSHV-DHFR mRNA and protein appeared from 6 to 24 h after chemical induction of the KSHV lytic cycle. Epitope-tagged KSHV-DHFR and rhDHFR both localized to the nucleus of transfected cells, while other KSHV nucleotide metabolism genes localized to the cytoplasm. DHFR activity was not essential for viral replication in cultured PEL cells. Since hDHFR was not detectable in peripheral blood mononuclear cells (PBMCs), KSHV-DHFR may function to provide increased DHFR activity in vivo in infected cells that have little or none of their own enzyme.     

S-nitrosoglutathione reductase in human lung cancer

S-Nitrosoglutathione (GSNO) reductase regulates cell signaling pathways relevant to asthma and protects cells from nitrosative stress. Recent evidence suggests that this enzyme may prevent human hepatocellular carcinoma arising in the setting of chronic hepatitis. We hypothesized that GSNO reductase may also protect the lung against potentially carcinogenic reactions associated with nitrosative stress. We report that wild-type Ras is S-nitrosylated and activated by nitrosative stress and that it is denitrosylated by GSNO reductase. In human lung cancer, the activity and expression of GSNO reductase are decreased. Further, the distribution of the enzyme (including its colocalization with wild-type Ras) is abnormal. We conclude that decreased activity of GSNO reductase could leave the human lung vulnerable to the oncogenic effects of nitrosative stress, as is the case in the liver. This potential should be considered when developing therapies that inhibit pulmonary GSNO reductase to treat asthma and other conditions.     

Methylenetetrahydrofolate reductase polymorphism and pre-eclampsia.

A common missense mutation in the methylenetetrahydrofolate reductase (MTHFR) gene, a C to T substitution at nucleotide 677, is responsible for reduced MTHFR activity and associated with modestly increased plasma homocysteine concentrations. Since underlying maternal vascular disease increases the risk of pre-eclampsia, we had the working hypothesis that pre-eclampsia patients would have an increased T677 allele frequency compared with controls. The MTHFR genotypes were determined in 67 pre-eclampsia patients, 98 normal pregnant women, and 260 healthy adults by the PCR/RFLP method. The T677 allele and the genotype homozygous for the T677 allele were significantly increased in the pre-eclamptic group compared with the controls (p < 0.02 and p < 0.004, respectively). The data indicate that the T677 variant of the MTHFR gene is one of the genetic risk factors for pre-eclampsia.     

Dopamine agonists in dihydropteridine reductase deficiency

The traditional treatment of severe disorders of tetrahydrobiopterin (BH4) metabolism is based on the replacement therapy with BH4, 5-hydroxytryptophan, and L-dopa. Major problems are encountered with L-dopa therapy, especially with increasing age when higher doses are necessary, because of its short half-life and adverse effects. Consequently, different L-dopa-sparing strategies have been successively introduced, with partial reduction of L-dopa dosage and amelioration of the clinical outcome. Recently, we demonstrated that the dopamine agonist pramipexole improves the therapeutic effect of L-dopa in 6-pyruvoyl tetrahydropterin synthase (PTPS) deficiency, the most common disorder of BH4 metabolism. Here we report its effectiveness in two patients (males, 7 and 22 years) with dihydropteridine reductase (DHPR) deficiency, the second most frequent cause of BH4 deficiency. Both patients experienced residual symptoms of dopamine deficiency, movement and behavioral disability, and complications of L-dopa therapy, associated with fluctuating hyperprolactinemia. They had full clinical and biochemical assessment, by an adapted Unified Parkinson's Disease Rating Scale (UPDRS) and measurement of diurnal plasma prolactin (PRL) profile before and after a trial with pramipexole. Besides allowing the reduction of L-dopa daily dosage (-58%) and administrations (from three to two) in one patient and to stop L-dopa therapy in the other, the introduction of pramipexole markedly improved and stabilized clinical and biochemical picture in both patients, as revealed by reduction of UPDRS scores and normalization of diurnal plasma prolactin profiles. Dopamine agonists can improve or even replace L-dopa therapy in disorders of synthesis and regeneration of BH4.     

Immunomodulatory effects of HMG-CoA reductase inhibitors

3-Hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase inhibitors, or statins, are competitive inhibitors of the rate-limiting enzyme in cholesterol synthesis. Several clinical trials have shown a marked reduction in cholesterol levels associated with decreased cardiovascular mortality in patients treated with statins. However, more recent observations have suggested that the clinical benefits of statins may be, at least in part, independent of the effect of statins on cholesterol synthesis. These so-called pleiotropic or cholesterol-independent effects of statins could be the result of reduction in the formation of intermediaries in the mevalonate pathway as statins, by inhibiting L-mevalonic acid synthesis, also prevent the production of isoprenoids in the cholesterol biosynthetic pathway. Isoprenoids serve as important lipid attachments for the posttranslational modification of a variety of proteins such as small GTP-binding proteins of the Ras superfamily implicated in intracellular signaling. The list of different pleitropic effects of statins is still growing and includes, among others, direct effects of statins on modulating endothelial function, decreasing oxidative stress and, more recently, anti-inflammatory and immunomodulatory actions of statins. For instance, statins decrease T cell activation, the recruitment of inflammatory cells into atherosclerotic lesions, and inhibit IFN-gamma expression of MHC II on antigen-presenting cells. This review article summarizes the anti-inflammatory and immunomodulatory effects of statins and thus provides a new rationale to use statins as a new class of immunosuppressive agents.     

Methylenetetrahydrofolate reductase thermolabile variant and oral clefts

Folic acid can prevent neural tube defects; in some cases the mechanism is probably a correction of a metabolic defect caused by thermolabile methylenetetrahydrofolate reductase (MTHFR) found in increased frequency in cases. It is less clear whether folic acid can prevent oral clefts, in part because it is not known whether thermolabile MTHFR is more common in those with oral clefts. This study examined the prevalence of the mutation (677 C→T) that causes thermolabile MTHFR in subjects with oral clefts from a national Irish support group, and an anonymous control group randomly selected from a neonatal screening program covering all births in Ireland. Eighty-three of 848 control subjects were homozygous (TT) thermolabile MTHFR (9.8%). This defect was almost three times as common in the 27 subjects (25.9%) with isolated cleft palate (odds ratio 3.23, 95% confidence interval 1.32 –7.86, P = 0.02) and somewhat more common in the 66 subjects with cleft lip with or without cleft palate (15.2%, odds ratio 1.65, 95% confidence interval 0.81–3.35, P = 0.20). When the two groups with different etiologies were combined, the overall odds ratio was 2.06 (95% confidence interval 1.16–3.66, P = 0.02). In the Irish population homozygosity for the common folate-related polymorphism associated with thermolabile MTHFR is significantly more frequent in those with isolated cleft palate, and could be etiologically important. Am. J. Med. Genet. 86:71–74, 1999. Published 1999 Wiley-Liss, Inc.     

Methylenetetrahydrofolate reductase variant and schizophrenia/depression

Patients with methylenetetrahydrofolate reductase (MTHFR) deficiency often show psychiatric manifestations. Since a common variant of the MTHFR gene, T677(Ala), responsible for the thermolabile MTHFR with less than 50% specific MTHFR activity, has been reported, we examined whether the T677 allele is associated with psychiatric disorders in an unrelated Japanese population consisting of 297 schizophrenics, 32 patients with major depression, 40 patients with bipolar disorder, and 419 controls. The genotype homozygous for the T677 allele was significantly frequently observed in schizophrenics with an odds ratio of 1.9 (P = 0.0006), and in patients with major depression with an odds ratio of 2.8 (P = 0.005). Our data suggest associations of the MTHFR gene variant with schizophrenia and depression in the Japanese. Am. J. Med. Genet. 74:526–528, 1997. © 1997 Wiley-Liss, Inc.     

A population study of alpha-keto acid reductase

An electrophoretic survey of 509 individuals of Japanese, Mexican American, American Negro, Eskimo, Amerindian and Anglo-American origin failed to reveal genetically determined variation at the alpha-keto acid reductase locus by starch gel electrophoresis. Additional screening of 232 individuals by thin layer isoelectric focusing in polyacrylamide gels detected a single phenotype. Interspecific variation at the alpha-keto acid reductase locus is clearly resolved using either method. These results indicate that the alpha-keto acid reductase locus is monomorphic in most human population groups despite earlier results suggesting the existence of genetic polymorphism.