Search PubMed⌕ Search

Biomedical subjects

F Palma

Publications and source records attributed to F Palma.

At least 37 records · Page 2Linked to original sources
PubMed1994

Comparative aspects of Na(+)-K+ and Ca(2+)-Mg2+ ATPase in erythrocyte membranes of various mammals.

This work is a comparative study of Na(+)-K+ and Ca(2+)-Mg2+ ATPase associated with the erythrocyte plasma membranes in different mammals. The method used to test the activity of these enzymes is based on quantitative measurements of ADP released during the reaction with HPLC: the chromatographic type is an Ion-Pair Reversed Phase. We have found that the levels of Ca2+ stimulated ATPase are higher than those of Na(+)-K+ ATPase in red blood cells of all the different mammalian species, with the only exception being lamb erythrocytes where the values of both the ATPase activities are almost equal. The results obtained have shown different levels of the Na(+)-K+ ATPase as well as of the Ca2+ stimulated ATPase activity. Furthermore, we have made a comparative study of the ATPase activities in different red blood cells with specific reference to the optimum pH, thermostability, kinetic characteristics and the inhibitory effects of ouabain and vanadate.

Adenosine Diphosphate↗
PubMed1992

Liver transplantation in malignant primary hepatic neoplasms.

Between April 1986 and August 1990, 151 liver transplantations were performed at our institution, 16 (11%) of them in 14 patients with primary hepatic tumors. There were 12 hepatocellular carcinomas, 1 angiosarcoma, and 1 Klatskin tumor. None of the tumors was resectable, and there was no preoperative evidence of extrahepatic tumoral extension. Exploratory laparotomy was performed prior to transplantation in three patients and selective embolization of the tumor in six patients. There was no difference in the intraoperative requirements for blood or plasma in the patients with hepatic tumors when compared with other transplant recipients (28.6 +/- 23.6 units packed red blood cells [PRBC] versus 20.1 +/- 17.8 units PRBC, and 17.9 +/- 12.2 units plasma versus 17.1 +/- 10.5 units plasma, respectively). Extracorporeal venovenous bypass was used in all but one patient. There was no significant differences in the incidence of acute rejection or in the length of hospitalization in these patients when compared with other transplant recipients. All patients received triple immunosuppressive therapy (corticosteroids, azathioprine, and cyclosporin A). Intraoperative mortality was zero. At a mean of 13.3 months' follow-up (range: 1 to 47 months), 2 of 14 patients had died of sepsis and 1 of terminal cirrhosis (autopsies revealed no evidence of tumor recurrence); 3 patients (21%) had recurrences of the tumor (1 in the central nervous system and liver, and the other 2 in the lung). One of the three patients with a recurrent tumor is still alive after 16 months. The remaining nine patients (64%) are still alive.

Adult↗
PubMed1992

High resolution of multiple forms of red blood cell enzymes using a Toyopearl DEAE 650 S.

We have investigated a new anion exchange chromatographic support (Toyopearl DEAE 650 S) which simultaneously allows easily to remove hemoglobin from hemolysates and to obtain a very high resolution of enzymes present in multiple forms. The results obtained are better than those obtainable using an anion-exchange HPLC column. The data obtained at analytical level suggest a wider use of this new matrix also for preparative purposes without significant changes in the resolution.

Animals↗
PubMed1992

Preparative purification of pig red blood cell hexokinase type III using a new efficient chromatographic support.

In this paper we report the purification of pig erythrocyte hexokinase type III, at preparative level, using 52 liters of starting material (hemolysate). This was possible using a new efficient anion exchanger support, the Toyopearl DEAE 650 M which allows completely to change the strategy of removing hemoglobin from hemolysates, permitting to handle large amounts of starting material and reducing work would have required months using conventional anion exchanger supports, to only 2-3 days. Furthermore, we have tested the binding of other red blood cell enzymes to the Toyopearl DEAE 650 M, showing a wider potential use of this chromatographic support for their purification at a preparative level.

Animals↗
PubMed1991

Glucose-6-phosphate dehydrogenase activity and protein turnover in erythroblasts separated by velocity sedimentation at unit gravity and Percoll gradient centrifugation.

This work was undertaken to improve a separation method for preparation of large amounts of erythroid cells of different age with homogeneous and minimal contamination of myeloid cells. Our method was suitably employed in the study of the decay mechanism of glucose-6-phosphate dehydrogenase (G6PDH) during the erythroid cell maturation. Twenty fractions of erythroid cells at different advancing stages of maturation were prepared by fractionating, at unit gravity, bone marrow cells from anaemic rabbit. The specific activity of the G6PDH was assayed and plotted vs the fraction number and the typical sigmoid curve of the activity decay was drawn. The separated cells were then grouped in three sets of fractions following the three phases of the sigmoid curve and the fractions of each set were combined. From the cytochemical analysis of the three main fractions so obtained, we found a 25-30% myeloid cell contamination in the first fraction, while in the other two fractions the myeloid contamination was 10% or less. For this reason we performed a rapid separation of the first fraction on a discontinuous percoll gradient. By this method, the myeloid cell contamination of the first fraction was levelled down to the other two. The fractions, so obtained, (I, II and III in order of increasing cell maturation) showed a four fold decrease of glucose-6-phosphate dehydrogenase activity expressed both per cell number and on protein base. On the contrary the concentration of the total soluble proteins did not change significantly in the three fractions. The three purified cellular populations were used to provide information on the protein turnover of the erythroid cells during their development. We measured, in intact cells, the rate of synthesis and degradation of total proteins and then, in cell lysates, we determined the rate of degradation of G6PDH, purified from rabbit RBC and radiolabeled by reductive methylation with C14-formaldehyde. The rates of proteolysis obtained with total proteins and methyl-G6PDH clearly indicate that the proteolytic machinery of the erythroblasts reduces its activity during the cell maturation.

Anemia↗
PubMed1991

Free radicals promote "in vitro" a different intracellular decay of rabbit reticulocyte and erythrocyte glycolytic enzymes.

Rabbit red blood cells (RBC) were exposed in vitro to an oxygen-radical-generating system represented by iron and ascorbic acid. Under these experimental conditions we have investigated the effect of this system on some intracellular rabbit reticulocyte and erythrocyte enzymes. The results obtained have shown a pronounced decay of hexokinase activity both in the erythrocytes and reticulocytes when exposed to these radical species. We have found that the amount of hexokinase inactivated is at least three times higher in a blood sample with a percentage of reticulocytes of 50-60%. This different behaviour of the hexokinase decay in the erythrocytes and reticulocytes could be due to its different intracellular distribution related to the two distinct cells. In addition we have evaluated some important intracellular compounds involved in maintaining the redox and the energetic state of the cell such as the reduced glutathione and the adenine nucleotides and their degradation products, in order to understand if there is any correlation between the hexokinase decay and a change concerning the metabolic conditions of the rabbit reticulocytes and erythrocytes exposed to free radicals.

Animals↗
PubMed1991

[HPLC method for measuring (Na(+)-K(+)) ATPase and (Ca(++)-Mg(++)) ATPase in erythrocytes from different species of mammals].

(Na(+)-K+)ATPase and (Ca(++)-Mg++)ATPase are enzymes located in erythrocyte plasma membranes, driving back ions against the electrochemical gradient; (Na(+)-K+)ATPase transports 3 Na+ ions out of the cell, and 2 K+ ions into it for each hydrolyzed ATP molecule, whereas the Ca(2+)-pump transports Ca2+ ions out of the cells, by utilizing still the ATP hydrolysis. The method used to test the activity of the above mentioned enzymes is based on the measuring of the ADP quantity released during the reaction by HPLC, that is High Performance Liquid Chromatography; the chromatographic type is a Ion-Pair Reversed-Phase. This method presents the following important advantages for the assay of the enzymes we analysed: 1) It is reproducible through time; 2) It is perfectly linear; 3) It is extremely sensitive. This method allowed us to carry out a comparative study of (Na(+)-K+)ATPase and (Ca(++)-Mg++)ATPase in erythrocyte plasma membranes of several species of mammalia: man, horse, rabbit, lamb, rat. We recovered different values in ATPase activity; (Ca(++)-Mg++)ATPase shows a higher activity than Na(+)-K+)ATPase; moreover, some differences exist in the various Mammalia considered, with relation to each pump: the lamb shows the lowest activity for both pumps, whereas the rabbit shows the highest one. At present, the different values obtained are being interpreted and analysed. This method is also very versatile, since it allowed us to assess the Km value for Ca++ of the (Ca(++)-Mg++)ATPase in erythrocyte plasma membranes of rabbit. The value resulted to be 100 microMs, thus 10 times higher than the human Km value for the Ca++.

Adenosine Diphosphate↗
PubMed1990

Comparative study on glucose-6-phosphate dehydrogenase from rabbit tissues.

The activity of glucose-6-phosphate dehydrogenase (G6PD) was measured in bone marrow, spleen, lung, liver, kidney, adipose tissue, brain, heart, muscle, and in the erythroid cell line of rabbit. In tissues, the activity ranged from 6.87 to 0.09 U/g wet tissue, found in bone marrow and muscles, respectively, whereas in the erythroid cell line it ranged from 14.3 to 2.4 U/g cells for erythroblasts and erythrocytes, respectively. The electrophoretic patterns of the tissue crude extracts showed an identical set of three activity bands, and the immunotitration curves obtained with rat antirabbit erythrocyte G6PD antibodies shared the same equivalence point. The enzyme, purified to homogeneity from different tissues, showed no significant differences among the Km values for NADP and G6P. The results give a picture of the variability of the G6PD activity in rabbit tissues and suggest the presence of the same enzyme molecule in each tissue.

Animals↗
PubMed1989

Reversed-phase high-performance liquid chromatography separation of dimethylaminoazobenzene sulfonyl- and dimethylaminoazobenzene thiohydantoin-amino acid derivatives for amino acid analysis and microsequencing studies at the picomole level.

A simple and fast reversed-phase high-performance liquid chromatographic method has been developed for the complete separation of 35 dimethylaminoazobenzene sulfonyl (DABS)-amino acids and by-products. This method allows simultaneous determination of primary and secondary amino acids which can be present in protein and peptide hydrolysates and also detects the presence of cysteic acid, S-sulfocysteine, hydroxyproline, taurine, norleucine, cystine, and delta-hydroxylysine. The precolumn derivatization of amino acids with dimethylaminoazobenzene sulfonyl chloride (DABS-Cl) is simple and quick (10 min at 70 degrees C) and allows the complete reaction of primary and secondary amino acids. The separation of the compounds under investigation is achieved in 25 min using a reversed-phase 3-microns Supelcosil LC-18 column at room temperature. The versatility of the proposed method is documented by amino acid determination on protein samples obtained using different hydrolysis techniques (HCl, methane-sulfonic acid, and NaOH), with attention given to the detection of tryptophan in protein samples with high sugar concentration. Furthermore, we have reported the experimental conditions necessary to apply this method to the amino acid analysis of very low amount of proteins (1 to 5 micrograms) electroeluted from a stained band after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The stability of DABS-derivatives, the short time of analysis, the high reproducibility and sensitivity of the system, and the complete resolution of all compounds of interest make this method suitable for routine analysis. Furthermore, we have also developed a fast reversed-phase high-performance liquid chromatographic method for the complete separation of dimethylaminoazobenzene thiohydantoin (DABTH)-amino acids. The separation of the compounds under investigation is obtained, at room temperature, in less than 18 min using a reversed-phase Supelcosil LC-18 DB column, 3-micron particles, and also allows the complete separation of DABTH-Ile, DABTH-Leu, and DABTH-Norleu. The short time of analysis, together with the high reproducibility of the system and its sensitivity at picomole levels, make this method very suitable for the identification of DABTH-amino acids released during microsequencing studies of proteins and peptides with the dimethylaminoazobenzene isothiocyanate reagent. In addition, we have shown that it is possible to obtain complete separation of DABTH-amino acids also under isocratic conditions.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acids↗
PubMed1989

[Benignancy of the Delorme operation in the treatment of rectal prolapse. Indications and results].

In five patients, three male and two female (average age 74 years), with complete rectal prolapse, Delorme's operation was performed, realizing resection of the entire prolapsed rectal mucosa and the suturing the plicature of the rectal muscular layer, which permits its invagination into the muscular funnel formed by the outer sphincter. This technique, which also tenses the longitudinal muscle of the rectum that is lax in every total prolapse, partially remedies the dislocation of the hiatal ligament and improves the function of the elevator muscle, which is disordered in these patients. The five patients were followed-up for 3 to 36 months. There was no postoperative morbi-mortality in patients of advanced age or high risk had to be operated for rectal prolapse.

Aged↗
PubMed1987

Rabbit bone marrow glucose-6-phosphate dehydrogenase during erythroid cell development.

Studies were carried out on glucose-6-phosphate dehydrogenase (G6P-DH) during the differentiation of rabbit bone marrow erythroid cells. It was found that G6P-DH, although displaying a 7-fold activity decrease, did not change the relative amounts of its three dimeric forms. Using homogeneous enzyme preparations, we observed that from dividing to non-dividing erythroblasts the following properties remained constant: V max dependence on pH and temperature, Km for G6P dependence on pH, heat stability, 2-deoxy glucose-6-phosphate utilization, molecular weight, while the Km for NADP significantly increased in non-dividing erythroblasts. These results indicate that no shift towards the oxidized form of the enzyme and no substantial modifications of the protein take place during cell differentiation.

Animals↗
PubMed1987

Action of acetaldehyde on glucose metabolism of newborn and adult erythrocytes.

This paper reports the effect of acetaldehyde on erythrocytes (RBC) of human adults and newborns. Acetaldehyde increases glucose consumption in adult RBC, but has no effect on that of newborn RBC. The compound stimulates the hexose monophosphate shunt and decreases the pyruvate production of the two RBC suspensions. In newborn RBC, acetaldehyde slightly modifies triose-P and fructose-1,6-bisphosphate but has no effect on the ATP/ADP ratio and glucose-1,6-bisphosphate content, which change markedly in adult RBC. Analysis of aldehyde dehydrogenase reveals nearly one half of enzyme activity in newborn RBC. The data indicate that in both adult and newborn RBC, acetaldehyde causes an intracellular reduced state, but the newborn cells take advantage of their greater pyruvate production for complete removal of the exceeding NADH equivalents.

2,3-Diphosphoglycerate↗
PubMed1985

Glucose-1,6-P2 synthesis, phosphoglucomutase and phosphoribomutase correlate with glucose-1,6-P2 concentration in mammals red blood cells.

Glucose 1,6-biphosphate (G1,6P2) was measured in human, pig, cow, rabbit, rat and sheep red blood cells. Mean values are variable among the species and range from 33 to 122 nmol/ml RBC for pig and rabbit erythrocytes, respectively. The activities of G1,6P2 synthase, phosphoglucomutase (PGM) and phosphoribomutase (PRM) have also been assayed in red cell haemolysates of the same species. The correlations between the biphosphate content and the occurrence of the three enzymatic activities have been studied in order to gain an insight into the regulation of the G1,6P2 turnover in mammalian erythrocytes.

Animals↗