Simone Parvez

Simple purification of aromatic l-amino acid decarboxylase from human pheochromocytoma using high-performance liquid chromatography☆

We purified aromatic L-amino acid decarboxylase (AADC) homogeneously and rapidly from human pheochromocytoma using high-performance liquid chromatography. HPLC with gel permeation and hydrophobic columns was highly effective, and the entire purification could be finished within 3 days. Purified AADC showed a single band with an Mr of 50,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and decarboxylated L-3,4-dihydroxyphenylalanine, L-5-hydroxytryptophan, and L-threo-3,4-dihydroxyphenylserine (a synthetic precursor of natural norepinephrine). Amino acid analysis of purified AADC was performed.

VARIATION IN ACTIVITY OF MONOAMINE METABOLIZING ENZYMES IN RAT LIVER DURING PREGNANCY

1 Catechol‐O‐methyltransferase (COMT) and monoamine oxidase (MAO) activities in rat liver were measured during pregnancy, parturition and postpartum. Compared with activity in non‐pregnant controls, both enzymes showed a significant decrease in activity which was most pronounced at day 18. 2 The metabolism of intravenously infused [3 H]‐adrenaline to [3 H]‐metanephrine and to [3 H]‐acidic metabolites was also significantly depressed during pregnancy but had returned to control values by the 21st day. 3 The effects of reserpine and/or nialamide on hepatic COMT and MAO were studied in control and 20‐day‐pregnant rats. Their action on COMT activity differed in the two groups. MAO was inhibited to a similar extent in these groups whether the drugs were given separately or in combination. 4 It seems possible that the changes in endocrine function which occur during pregnancy are responsible for the observed alterations in enzyme activity.

Placental Transfer of 3H-Epinephrine and Its Metabolites to the Fetal Heart during Variable Hormonal Treatments

The possible transfer of high-specific activity 3H-epinphrine from maternal circulation to the fetal heart was studied. Different hormones were administered to pregnant rats to determine the effects of endocrine function in permeability of catecholaminesthrough the placenta. The perfusion of 60 µCi3H-epinephrine to ratspregnant for 21 days resulted in a significant accumulation of the tracer in the fetal heart. This accumulation was markedly declined by pre-administration of progesterone, estrogens, and oxytocin in the pregnant rats and by maternal adrenalectomy or hypophysectomy. Similar treatment produced highly significant increases in the transformation of 3H-epinephrine to 3H-metanephrine and 3H-acid metabolites in the fetal heart. The maximum increases in catecholamine metabolite formation in the fetal heart were observed following maternal adrenalectomy and hypophysectomy. The results suggest that there is an appreciable transfer of catecholamines and their metabolites from the maternal circulation to the fetal heart on the 21st day of pregnancy. The treatment with different hormones modifies this transfer to a significant extent and could be attributed to the direct action of these hormones on catecholamine regulation.

Activity of catechol-o-methyltransferase in brain regions and adrenal gland during the oestrus cycle.

The activity of the enzyme catechol-O-methyltransferase (COMT) during four different phases of the oestrus cycle were determined. Brain and hypothalamus had highest level of COMT activity during oestrus phase, while at pro-oestrus it was at its lowest level. COMT activity in the adrenal gland was also modified during the four phases with the maximum level at met-oestrus and the minimum at pro-oestrus phase. The results provide evidence to show that COMT in the brain, hypothalamus and the adrenals is markedly affected by changes in physiological status of the female rats and its evolution in brain is different from adrenal gland during the oestrus cycle. The variations in the activity of COMT may be the consequence of modified endocrine activity which takes place naturally during oestrus cycle.

Progesterone mediated increase in monoamine stores and the regulation of enzymes of biosynthesis and metabolism in the adrenal gland during late pregnancy in the rat.

The influence of repeated injections of progesterone to pregnant rats upon monoamine storage and regulation of enzymes phenylethanolamine-N-methyltransferase (PNMT), monoamine oxidase (MAO) and catechol-O-methyltransferase (COMT) was studied. All the pregnant females received progesterone (4 mg/100 g body weight) on 19, 20 and 21 days post-coitum but one group was killed at 21 days of pregnancy and the other one at 0 h parturition. Adrenal epinephrine demonstrated highly significant increase in progesterone treated rats. At the same time norepinephrine content declined significantly from the control value. The activity of enzyme PNMT also showed marked increase in the adrenals of progesterone treated females. Activity of enzyme MAO showed a slight decline after progesterone treatment to pregnant rats. Enzyme COMT in progesterone treateed animals showed decline at 0 h parturition but at 21 days post-coitum it was significantly higher from non-injected females. All the increases and decreases in monoamines and the three enzymes were significant when the results were expressed per adrenal gland or per gram of adrenal. The results suggest that exogenous progesterone administration during late pregnancy increases epinephrine stores by declining monoamine metabolism by MAO and COMT and increasing their synthesis by PNMT which is responsible for N-methylation of norepinephrine to epinephrine.

The neurochemical markers in cerebrospinal fluid to differentiate between aseptic and tuberculous meningitis.

In this study, the use of neurochemical markers in patients with aseptic and tuberculous meningitis has been investigated. The cerebrospinal fluid levels of amino acids, nitrite (a metabolite of nitric oxide), vitamin B12 and homocysteine were quantitated in both groups of patients. Among the amino acids, aspartic acid and glutamic acid both excitatory amino acid, GABA, glycine and tryptophan were all significantly increased in both patient groups whereas decreased level of taurine and increased level of phenylalanine were only found in patients with tuberculous meningitis. The levels of nitrite and its precursor arginine were significantly higher in patients with tuberculous meningitis whereas unchanged levels were found in patients with aseptic meningitis. A significantly increased homocysteine level and a decreased level of vitamin B12 were found only in patients with tuberculous meningitis whereas unchanged levels were found in patients with aseptic meningitis. This indicates that patients with tuberculous meningitis are particularly prone to vitamin B12 deficiency resulting into increased level of HC, and involvement of free radical showing the importance of these biological markers for promoting the possibility for the design of therapeutic approach.

CHARACTERIZATION OF ENZYMES OF CATECHOLAMINE SYNTHESIS AND METABOLISM IN HUMAN FETAL MEMBRANES AT BIRTH

We looked for the presence of the enzymes monoamine oxidase, catechol-O-methyltransferase, and phenylethanolamine-N-methyltransferase in human fetal membranes at term. The activity of all three enzymes was detected via highly sensitive and selective radiometric enzyme assays. The most novel finding was the extremely high level of monoamine oxidase activity in the chorion compared with that in the amnion. The other enzyme of catecholamine metabolism, catechol-O-methyltransferase, did not show any difference in activity between the two layers. In addition, we observed that the enzyme phenylethanolamine-N-methyltransferase, which is primarily located in the adrenal medulla, was also present in appreciable levels in the two layers of fetal membranes. These results suggest that fetal membranes, like the placenta, possess the enzymatic machinery to metabolize catecholamines and have the capacity to synthesize epinephrine.

Decreased phenylethanolamine-N-methyltransferase and catechol-O-methyltransferase activity in rabbit adrenal glands during pregnancy.

1 Phenylethanolamine‐N‐methyltransferase (PNMT) activity in adrenal gland was lower than the control values at day 24 of rabbit pregnancy and fell throughout the gestational period. 2 Catechol‐O‐methyltransferase (COMT) activity during pregnancy and parturition followed a similar pattern to PNMT. 3 Adrenaline content of the adrenal gland declined significantly during the last few days of pregnancy but at parturition showed a tendency to return towards normal. 4 The biochemical mechanisms which result in a decline in PNMT and COMT activities and adrenaline concentration appear to be the consequence of modifications in endocrine gland secretion during pregnancy.

Chapter 30 – Role of Selenium, Iron, Copper, and Zinc in Parkinsonism

A number of essential trace elements play a major role in various metabolic pathways. Selenium (Se), copper (Cu), zinc (Zn), and iron (Fe) are essential trace elements that have been studied in many diseases, including autoimmune, neurological, and psychiatric disorders. The observations from these studies suggest that alterations in essential trace elements Se, Cu, Zn, and Fe may play a role in the pathogenesis of Parkinsons disease (PD) patients. However, the findings from plasma levels of the trace elements show a variety of results that are difficult to interpret. The extracellular concentrations of transition metals such as Cu and Fe are substantially elevated during aging and in some neurodegenerative disorders. Increases in the extracellular redox capacity can potentially generate neurotoxic free radicals from reduction of Cu(II) or Fe(III), thereby resulting in neuronal cell death. Although observations support the role of iron as a neurotoxin, it remains to be established whether accumulation of iron in PD is primary or secondary to other known events such as glutathione (GSH) depletion and the mitochondrial transport chain complex I inhibition. Cu and Zn have significant antiatherogenic effects influencing the activity of antioxidant enzymes (glutathione-peroxidase and superoxide-dismutase), mechanism of apoptosis, and other mechanisms. In addition to the alterations in the homeostasis, redox activity, and localization of transition metals, it is also important to realize that the alterations in specific zinc-, copper-, and iron-containing metalloenzymes appear to play a crucial role in the neurodegenerative process.

Alterations in activity of enzyme catechol-O-methyltransferase in central and peripheral regions of the rat and rabbit during pregnancy, parturition, and the postpartum period

Natural variations in the activity of the enzyme catechol-O-methyltransferase in central and peripheral regions of the rat and rabbit during pregnancy and parturition were observed. All of the organs of the rat showed a decrease in enzyme activity up to the twenty-first day of pregnancy. Rabbit tissues also demonstrated a decrease in catechol-O-methyltransferase activity during Days 24 to 30 after coitus. Increases in activity of the rat ovary, brain, and hypothalamus took place from during Day 21 after coitus to the onset of parturition. The enzyme activity in the rat hypothalamus reached its maximum level at four hours post partum from the Day 21 value, but the rabbit hypothalamus showed the maximum activity at 0 hour parturition. During post partum hours, rat brain catechol-O-methyltransferase activity was very high but the rabbit brain showed a very low level of activity. Rabbit uterine activity reached its minimum value at Day 31 after coitus, but at the comparable intervals the situation was inversed in the rat. Results show an important modification of monoamine degradation by 3-O-methylation during midgestation in the central and peripheral regions. These modifications may be attributed to the natural variations in endocrine activity taking place during pregnancy.