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Biomedical subjects

M D Fernández-Moreno

Publications and source records attributed to M D Fernández-Moreno.

9 recordsLinked to original sources

Thiopurine methyltransferase activity in a Spanish population sample: decrease of enzymatic activity in multiple sclerosis patients.

The present study was performed in order to obtain the thiopurine methyltransferase (TPMT) activity frequency distribution histogram in a Spanish population. A total of 3640 Spanish clinical laboratory samples were evaluated, which included 1249 patients with Crohn's disease, 589 with ulcerative colitis, 348 with multiple sclerosis (MS), 487 with several autoimmune diseases different from the above-mentioned diseases and 967 a donor group. We have measured the TPMT activity in red blood cells (RBCs) by a radiochemical method, using S-adenosyl-L-[methyl-3H]methionine as methyl donor. The different groups present in their entirety a normal distribution histogram and a wide range of TPMT activity from 0 to 41 U/ml RBCs. The differences found between the Spanish population TPMT activity frequency distribution histogram and the pattern previously described in a North American population were not due to azathioprine treatment or gender. The effect of autoimmune diseases on TPMT activity was evaluated: the enzymatic activity was similar in the donor group (19.9 +/- 6.3 U/ml RBCs) and in the patients with Crohn's disease (20.0 +/- 5.8 U/ml RBCs) and ulcerative colitis (19.7 +/- 6.1 U/ml RBCs); however, it decreased significantly (p<0.0001) in MS patients (17.1 +/- 6.1 U/ml RBCs) with respect to the donor group. In conclusion, our results show that the Spanish population TPMT distribution is closer to that of the Jewish population of Israel than to North American populations, and that in MS the enzymatic activity of TPMT decreases significantly. This observation may take into account the usage of azathioprine as therapeutic agent in Spanish MS patients.

Azathioprine↗

Determination of thiopurine methyltransferase activity in human erythrocytes by high-performance liquid chromatography: comparison with the radiochemical method.

The current article describes a new assay to measure thiopurine methyltransferase (TPMT) activity from red blood cells. This method is based on the measurement of the reaction product 6-methylmercaptopurine (6-MMP) by high-performance liquid chromatography (HPLC). 6-MMP is extracted by ethyl acetate with recoveries of 85%, 80%, 80%, and 92% for 50, 250, 500, and 1,000 ng/100 microL packed red blood cells, respectively. 6-MMP was identified and measured by a Zorbax CN column installed in an HPLC system. The chromatograms were resolved using a mobile phase consisting of 40 mmol/L sodium phosphate buffer (pH 3) and methanol in a gradient from 1% to 20% of methanol. Under these conditions 6-MMP is well resolved from substrates (6-mercaptopurine and S-adenosyl-L-methionine) and endogenous peaks. When the TPMT activity from 20 patients was measured by the HPLC-linked assay and the classic radiochemical method, a linear correlation was obtained between both procedures ( y = 0.99x + 0.33; x-axis, radiochemical assay; y-axis, HPLC-linked assay; r = 0.98). In conclusion, the current report describes a new, reliable, safe, and nonradioactive method to measure TPMT activity that is shorter and simpler than the previously described ones.

Antimetabolites, Antineoplastic↗

Expression of insulin-like growth factor-I (IGF-I) receptor gene in rat brain and liver during development and in regenerating adult rat liver.

Insulin-like growth factor-I (IGF-I) is involved in the growth and development of liver and brain during fetal life by acting through specific plasma membrane receptors. In an attempt to determine how the changes in IGF-I receptor number are regulated during development, we have compared [125I]IGF-I binding to membrane fractions with the concentration of IGF-I receptor mRNA in rat liver and brain. IGF-I binding to liver membranes was 4.5 times higher in 20-day-old fetuses than in adult rats. After partial hepatectomy (70%) in adult rats a transient 2-fold increase of IGF-I binding to liver membranes was observed. In fetal and regenerating liver increases similar to those observed for IGF-I binding were observed in IGF-I receptor mRNA concentrations. In brain microsomes IGF-I binding was 3.5 times higher in 20-day-old fetuses than in adults. This difference reflects a similar change in the number of IGF-I receptors without modifications in the affinity of the receptor for the ligand. In contrast to the liver, no significant changes in the concentration of IGF-I receptor mRNA were observed in the developing brain when determined by hybridization solution, Northern blot or RNase protection analysis. These findings suggest that in the liver, the IGF-I receptor gene is regulated at pretranslational level during development and regeneration, while in brain it is preferentially regulated at translational or posttranslational level.

Animals↗

Effect of bilateral adrenalectomy on VIP receptor/effector system in rat intestinal epithelial cells.

The number of vasoactive intestinal peptide (VIP) receptors and the efficiency of VIP in the stimulation of cyclic AMP accumulation in rat jejunal epithelial cells increased after bilateral adrenalectomy. However, this condition increased neither receptor affinity nor VIP potency. In addition, jejunal VIP levels followed a parallel increase. These changes reversed to control conditions after glucocorticoid replacement with dexamethasone indicating that adrenalectomy modifies the intestinal VIP receptor/effector system and suggest a relationship between corticosteroids and VIP in the functions of intestinal epithelium.

Adrenalectomy↗

Lindane effect upon the vasoactive intestinal peptide receptor/effector system in rat enterocytes.

Isolated rat enterocytes exposed to the insecticide lindane (the gamma-isomer of hexachlorocyclohexane, HCCH) showed an important decrease in the efficiency of the neuropeptide vasoactive intestinal peptide (VIP) upon the stimulation of cyclic AMP accumulation. The effect of lindane was time- and dose-dependent, optimal conditions being reached after 5 min incubation of cells at 25 degrees C with 0.5 mM of this organochlorine compound. Lindane action exhibited an important degree of specificity since the isomer alpha-HCCH and endrin reproduced the same inhibitory pattern but beta-HCCH and dieldrin were inactive. The inhibition of VIP-induced cyclic AMP accumulation could not be explained by a lindane-dependent reduction in the binding of VIP to its specific receptors. Among various possibilities, the results suggest the modification of membrane fluidity by lindane and/or the activation of Ca2+-dependent protein kinase C by this compound leading to phosphorylation of Gs/adenylate cyclase.

Animals↗

The effects of experimental uremia in rats on duodenal VIP levels and the interaction of VIP with duodenal epithelial cells.

The effects of experimental uremia on the concentration of vasoactive intestinal peptide (VIP) in duodenum as well as on the interaction of this neuropeptide with the corresponding epithelial cells were studied in rats. Duodenal VIP concentration was significantly decreased in uremic rats as compared to control animals. The specific binding of VIP to duodenal epithelial cells increased in rats with uremia due to an increase in the number of VIP receptors rather than a change in the binding affinity or in the extent of VIP degradation. On the other hand, the efficacy but not the potency of VIP upon cyclic AMP generation varied in parallel to that observed at the receptor level.

Animals↗

Effect of intestinal ischaemia on intestinal VIP levels and VIP interaction with intestinal epithelial cells from rat.

1. The number (but not the affinity) of vasoactive intestinal peptide (VIP) receptors in small intestinal epithelial cells decreased following intestinal ischaemia in rats as compared to sham-operated animals. 2. There was a parallel decrease of the efficiency (but not the potency) of the neuropeptide upon cyclic AMP formation at the same level after intestinal ischaemia. 3. The surgical manipulation did not modify the level of VIP immunoreactivity in the gut segment studied. 4. These results suggest that the VIPergic system is not directly involved in the high loss of water and electrolytes that appears following intestinal ischaemia.

Animals↗

Interaction of insulin with small intestinal epithelial cells from developing rats.

The interaction of insulin with its specific receptors was studied in jejunoileal epithelial cells from 14-, 20-, and 60-day-old rats. Characteristics such as potency and specificity of the insulin receptor system did not vary during development. However, insulin binding decreased with age due to a diminished concentration of insulin receptors, whereas the circulating levels of the hormone followed an inverse evolution. These results support previous suggestions on the potential role of insulin on the mechanisms of differentiation and proliferation of small intestinal mucosa.

Animals↗

Effect of resection of small intestine on the interaction of vasoactive intestinal peptide with rat colonic epithelial cells.

Vasoactive intestinal peptide (VIP) receptors and VIP-dependent cyclic AMP production were studied in rat colonic epithelial cells 3 days after a 60% resection of the small intestine. Basal cyclic AMP levels were similar in both control and resected animals. The potency, but not the efficiency, of the peptide on the stimulation of cyclic AMP production was diminished in cells from resected rats. Accordingly, the affinity of VIP receptors, but not the binding capacity, decreased as a consequence of the loss of a part of the small intestinal mucosa. These observations are consistent with the known inhibitory role of cyclic AMP on cell proliferation in colonic epithelium and other tissues and suggest a participation of VIP acting through the cyclic nucleotide in the compensatory hyperproliferative response of the colon following massive resection of the small intestine.

Animals↗