Biomedical subjects

Maria Angeles Martínez

Publications and source records attributed to Maria Angeles Martínez.

6 recordsLinked to original sources

PubMed2005

Genetic analysis of three genes causing isolated methylmalonic acidemia: identification of 21 novel allelic variants.

Isolated methylmalonic aciduria (MMA) is an inborn error of metabolism due to the impaired isomerization of l-methylmalonyl-CoA to succinyl-CoA. This reaction is catalyzed by the mitochondrial protein methylmalonyl-CoA mutase (MCM, EC 5.4.99.2), an adenosylcobalamin-dependent enzyme. Four different forms of isolated MMA have been described: mut MMA associated with defects in the MCM apoenzyme, and phenotypically divided into two subtypes mut- and mut0 MMA, and three different defects involved in the synthesis of the active form of the cofactor adenosylcobalamin, termed cbl MMA, and classified into three different complementation groups cblA, cblB, and cblH associated with defects in the MMAA and MMAB genes and with an unidentified protein, respectively. In this work we describe the genetic analysis of 25 MMA patients, mainly from Spain. Using biochemical and cellular approaches our patients have been classified, identifying 13 mut MMA, 7 cblA, 2 cblB, and 3 noncblA, noncblB deficient patients. cDNA and genomic DNA sequence analysis of the MUT, MMAA, and MMAB genes have allowed us to identify 27 different changes, 21 novel ones. Among the missense mutations identified in the MUT gene only one, the c.970G>A (p.A324T) variant located in the substrate binding domain is likely a mut- mutation. The remaining missense mutations c.326A>G (p.Q109R), c.983T>C (p.L328P), c.1846C>T (p.R616C), and c.1850T>G (p.L617R) are probably mut0. In the MMAA patients analyzed, frameshift mutations are prevalent. We have explored the genotype-phenotype correlation for this clinically heterogeneous disease.

Acidosis↗

PubMed2004

Vasopressin effects on the coronary circulation after a short ischemia in anesthetized goats: role of nitric oxide and prostanoids.

To examine the coronary effects of arginine-vasopressin during reperfusion after a short ischemia, left circumflex coronary artery flow was electromagnetically measured, and 15 min total occlusion of this artery followed by reperfusion was induced in anesthetized goats (five nontreated, five treated with the inhibitor of nitric oxide synthesis Nomega-nitro-L-arginine methyl ester (L-NAME) and five treated with the inhibitor of cyclooxygenase meclofenamate). The vasoactive drugs and L-NAME were intracoronarily injected, and meclofenamate by i.v. route. At 60 min of reperfusion, coronary vascular conductance was not changed significantly in nontreated and was decreased by 35% (P<0.01) in L-NAME-treated and by 30% (P<0.01) in meclofenamate-treated animals. During reperfusion, the coronary vasodilatation with acetylcholine (3-100 ng) and sodium nitroprusside (1-10 microg) was not altered in nontreated animals, and the vasodilatation with acetylcholine but not with sodium nitroprusside was partially decreased in L-NAME--but not in meclofenamate-treated animals. The vasoconstriction in response to arginine-vasopressin (0.03-0.3 microg) was increased during reperfusion in nontreated, was not changed in L-NAME-treated and was decreased in meclofenamate-treated animals. Therefore, it is suggested that during reperfusion after a short ischemia: (1) the coronary vasodilator reserve is preserved; (2) the coronary vasodilatation with acetylcholine is also preserved, but in this vasodilatation, the role of nitric oxide may be attenuated and prostanoids may be not involved; and (3) the coronary vasoconstriction with arginine-vasopressin is increased, probably due to both attenuation of the modulatory role of nitric oxide and the release of vasoconstrictor prostanoids.

Acetylcholine↗

PubMed2004

Mechanisms underlying responsiveness to tetrahydrobiopterin in mild phenylketonuria mutations.

A subtype of phenylalanine hydroxylase (PAH) deficiency that responds to cofactor (tetrahydrobiopterin, BH4) supplementation has been associated with phenylketonuria (PKU) mutations. The underlying molecular mechanism of this responsiveness is as yet unknown and requires a detailed in vitro expression analysis of the associated mutations. With this aim, we optimized the analysis of the kinetic and cofactor binding properties in recombinant human PAH and in seven mild PKU mutations, i.e., c.194T>C (p.I65T), c.204A>T (p.R68S), c.731C>T (p.P244L), c.782G>A (p.R261Q), c.926C>T (p.A309V), c.1162G>A (p.V388M), and c.1162G>A (p.Y414C) expressed in E. coli. For p.I65T, p.R68S, and p.R261Q, we could in addition study the equilibrium binding of BH4 to the tetrameric forms by isothermal titration calorimetry (ITC). All the mutations resulted in catalytic defects, and p.I65T, p.R68S, p.P244L, and most probably p.A309V, showed reduced binding affinity for BH4. The possible stabilizing effect of the cofactor was explored using a cell-free in vitro synthesis assay combined with pulse-chase methodology. BH4 prevents the degradation of the proteins of folding variants p.A309V, p.V388M, and p.Y414C, acting as a chemical chaperone. In addition, for wild-type PAH and all mild PKU mutants analyzed in this study, BH4 increases the PAH activity of the synthesized protein and protects from the rapid inactivation observed in vitro. Catalase and superoxide dismutase partially mimic this protection. All together, our results indicate that the response to BH4 substitution therapy by PKU mutations may have a multifactorial basis. Both effects of BH4 on PAH, i.e., the chemical chaperone effect preventing protein misfolding and the protection from inactivation, may be relevant mechanisms of the responsive phenotype.

Biopterins↗

PubMed2002

Coronary reactivity to endothelin-1 during partial ischemia and reperfusion in anesthetized goats. Role of nitric oxide and prostanoids.

To examine the coronary reactivity to endothelin-1 and its interaction with nitric oxide or prostanoids during partial coronary ischemia and reperfusion, left circumflex coronary artery flow was electromagnetically measured, and partial occlusion of this artery was induced for 60 min, followed by reperfusion in anesthetized goats (eight non-treated, six treated with N(w)-nitro-L-arginine methyl esther (L-NAME) and five treated with meclofenamate). During partial occlusion, coronary vascular conductance was reduced by 24-37% (P<0.01), and the coronary vasodilatation in response to acetylcholine (3-100 ng) and sodium nitroprusside (1-10 microg) was much decreased in every case; the vasoconstriction in response to endothelin-1 (1-10 microg) was depressed in non-treated animals, and this depression was reversed by L-NAME and was accentuated by meclofenamate. At 30 min of reperfusion coronary vascular conductance remained decreased by 22-27% (P<0.01), and the vasodilatation in response to acetylcholine (3-100 ng) and sodium nitroprusside (1-10 microg), as well as the vasoconstriction with endothelin-1 (1-10 microg), were as in the control and comparable in the three groups of animals. These results suggest: (a) that during ischemia, the coronary vasodilator reserve is greatly reduced, and the vasoconstriction with endothelin-1 is blunted, with preservation of the modulatory role of nitric oxide and involvement of vasoconstrictor prostanoids in this vasoconstriction, and (2) that during reperfusion, the coronary vasodilator reserve and the coronary reactivity to acetylcholine and endothelin-1 recover, but the modulatory role of nitric oxide in this reactivity may be attenuated.

Acetylcholine↗

PubMed2002

Functional characterization of PCCA mutations causing propionic acidemia.

Propionic acidemia (PA, MIM 232000 and 232050) is caused by a deficiency of mitochondrial biotin-dependent propionyl-CoA carboxylase (PCC, EC 6.4.1.3), a heteropolymeric enzyme composed of alpha and beta subunits, which are encoded by the PCCA and PCCB genes, respectively. The PCCA protein (alpha subunit) is responsible for the formation of carboxybiotin upon hydrolysis of ATP and contains a C-terminal biotin-binding domain and a biotin carboxylase domain, defined by homology with other biotin-dependent carboxylases, some of them characterized structurally. More than 24 mutations have been found in the PCCA gene in patients with PA, among them 14 missense mutations and one in-frame deletion, for which the precise molecular effect is unknown. In this study, we have established the pathogenicity of 11 PCCA mutations (10 missense and an in-frame deletion) by expression studies in deficient fibroblasts and in a cell-free in vitro system, and analyzed the effect of each mutation on PCC activity, protein stability and domain structure. The results show that most mutant proteins show an increased turnover and are functionally deficient, suggesting that the structural alterations they cause are incompatible with normal assembly to produce a stable, functional PCC oligomer. These results are discussed in the context of the genotype-phenotype correlations in PCCA-deficient PA patients.

Animals↗

PubMed2002

Coronary responses to endothelin-1 and acetylcholine during partial coronary ischaemia and reperfusion in anaesthetized goats.

To examine coronary reactivity to acetylcholine and endothelin-1 (ET-1) during partial ischaemia and reperfusion, flow in the left circumflex coronary artery was measured electromagnetically, and coronary partial ischaemia was induced by stenosis of this artery in anaesthetized goats. In eight animals not treated with N(G)-nitro-l-arginine methyl ester (l-NAME), coronary stenosis reduced coronary flow by 45%, mean arterial pressure by 16% and coronary vascular conductance by 34%. During this ischaemia, coronary vasodilatation to acetylcholine (0.003-0.1 microg) and sodium nitroprusside (SNP; 1-10 microg) was markedly reduced, and coronary vasoconstriction to ET-1 (0.01-0.3 nmol) was attenuated. After 30 min of reperfusion, coronary flow, mean arterial pressure and coronary vascular conductance remained decreased, and the effects of acetylcholine, SNP and ET-1 were as in control animals. In six goats treated with N(G)-nitro-l-arginine methyl ester, coronary stenosis reduced coronary flow by 26% and coronary vascular conductance by 24%, but did not affect mean arterial pressure. During this ischaemia, coronary vasodilatation to acetylcholine and SNP was also markedly reduced, but vasoconstriction to ET-1 was unaffected. After 30 min of reperfusion, coronary flow and coronary vascular conductance remained decreased and mean arterial pressure was normal; in addition, the effects of acetylcholine were lower, those of SNP were similar and those of ET-1 were higher than in control animals. Therefore partial ischaemia reduces the coronary vasodilator reserve and blunts coronary vasoconstriction to ET-1, and reperfusion does not alter the endothelium-dependent and -independent coronary vasodilatation or vasoconstriction to ET-1.

Acetylcholine↗