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Biomedical subjects

G Moro

Publications and source records attributed to G Moro.

At least 19 recordsLinked to original sources
PubMed2006

A mixture of prebiotic oligosaccharides reduces the incidence of atopic dermatitis during the first six months of age.

BACKGROUND: Oligosaccharides may alter postnatal immune development by influencing the constitution of gastrointestinal bacterial flora. AIMS: To investigate the effect of a prebiotic mixture of galacto- and long chain fructo-oligosaccharides on the incidence of atopic dermatitis (AD) during the first six months of life in formula fed infants at high risk of atopy. METHODS: Prospective, double-blind, randomised, placebo controlled trial; 259 infants at risk for atopy were enrolled. A total of 102 infants in the prebiotic group and 104 infants in the placebo group completed the study. If bottle feeding was started, the infant was randomly assigned to one of two hydrolysed protein formula groups (0.8 g/100 ml prebiotics or maltodextrine as placebo). All infants were examined for clinical evidence of atopic dermatitis. In a subgroup of 98 infants, faecal flora was analysed. RESULTS: Ten infants (9.8%; 95 CI 5.4-17.1%) in the intervention group and 24 infants (23.1%; 95 CI 16.0-32.1%) in the control group developed AD. The severity of the dermatitis was not affected by diet. Prebiotic supplements were associated with a significantly higher number of faecal bifidobacteria compared with controls but there was no significant difference in lactobacilli counts. CONCLUSION: Results show for the first time a beneficial effect of prebiotics on the development of atopic dermatitis in a high risk population of infants. Although the mechanism of this effect requires further investigation, it appears likely that oligosaccharides modulate postnatal immune development by altering bowel flora and have a potential role in primary allergy prevention during infancy.

Bifidobacterium↗
PubMed2004

Prebiotics in infant formulas.

BACKGROUND: The intestinal flora of breast-fed infants is an important physiologic factor in the function of the gut and in the development of the immune system. The current research is part of a group of studies performed to answer the question whether a bovine milk formula supplemented with a prebiotic mixture from galactooligosaccharides and fructooligosaccharides can stimulate an intestinal flora similar to that of breast-fed infants. METHODS: The prebiotic effect of the oligosaccharide mixture was tested in preterm and term infants by measuring fecal flora using plating as well as fluorescent in situ hybridization techniques. The effect of the oligosaccharides on the bacterial metabolism was studied by measuring short-chain fatty acid production in vitro and the short-chain fatty acid pattern in the stools of a group of term infants. RESULTS: The oligosaccharide mixture increases significantly the number of bifidobacteria and reduces the number of pathogens in term as well as in preterm infants when compared with a group of infants fed an unsupplemented formula. Using a concentration of 0.8 g oligosacchrides/100 mL formula, the amount of bifidobacteria is similar to that typical of breast-fed infants. In vitro, the short-chain fatty acids produced by the mixture of oligosaccharides under study were similar to those produced by the human milk oligosaccharides fraction. In clinical trials the pattern of fecal short-chain fatty acids in infants fed the oligosaccharide mixture was similar to that of breast-fed infants but was significantly different from that of a group of infants fed with an unsupplemented formula. Additionally, the fecal pH was significantly higher in the group fed an unsupplemented formula than in the groups fed either breast milk or a supplemented formula. CONCLUSION: The data obtained indicate that the prebiotic mixture under study is able to stimulate the development of a microbial flora similar to that of breast-fed infants. Several biota, whose growth is enhanced by this prebiotic mixture, represent important factors in the postnatal development of the immune system. On this evidence it can be suggested that prebiotics may play a role as modulators of the postnatal development of the immune system.

Animals↗
PubMed2003

[Use of platelet-rich plasma in major maxillary sinus augmentation].

AIM: Platelet-rich plasma (PRP) is a solution, usually in the form of a gel, with a high concentration of platelets (equal to double or three-fold normal reference values). This preparation also contains a high concentration of fibrinogen, a fundamental element for the release of numerous polypeptides (complementary factors) and above all for the induction of platelet degranulation and the consequent release of growth factors, which interact in a highly positive fashion with the bone metabolism at the host site. The biological benefit that PRP can give to tissues treated with the technique of major maxillary sinus augmentation thus appears undoubted. METHODS: The series reported consists of 3 patients who were treated by major augmentation of the maxillary sinus associated with the use of a mixture consisting of PRP, beta-tricalcium phosphate (Cerasorb ) and autologous bone tissue. RESULTS: All the patients were successfully treated and it was possible to detect tissue regeneration both horizontally and vertically (approximately 4 mm), with formation of high quality bone tissue, which subsequently enabled optimal osseointegration of the implants inserted. CONCLUSION: The introduction of PRP into the field of regenerative techniques has opened new horizons, especially in the branch of implantology. This mixture is able to reduce the quantity of autologous tissue harvested. The gel-like consistency also avoids the application and subsequent removal of a membrane, and in consequence the surgical trauma undergone by the patient is markedly reduced.

Blood Platelets↗
PubMed2002

Dosage-related bifidogenic effects of galacto- and fructooligosaccharides in formula-fed term infants.

BACKGROUND: Human milk oligosaccharides have been shown to stimulate selectively the growth of Bifidobacteria and Lactobacilli in the intestine. In this study, the bifidogenic effect of an experimental prebiotic oligosaccharide mixture consisting of low-molecular-weight galactooligosaccharides and high-molecular-weight fructooligosaccharides was analyzed in 90 term infants. METHODS: Two test formulas were supplemented with either 0.4 g/dL or with 0.8 g/dL oligosaccharides. In the control formula, maltodextrin was used as placebo. At study day 1 and study day 28, the fecal species, colony forming units (cfu) and pH were measured and stool characteristics, growth, and side effects were recorded. RESULTS: At study day 1, the median number of Bifidobacteria did not differ among the groups (0.4 g/dL group, mean [interquartile range] 8.5 [1.9] cfu/g; 0.8 g/dL group, 7.7 [6.1] cfu/g; and the placebo group, 8.8 [6.1] cfu/g) (figures in square brackets are interquartile range). At the end of the 28-day feeding period, the number of Bifidobacteria was significantly increased for both groups receiving supplemented formulas (the 0.4 g/dL group, 9.3 [4.9] cfu/g; the 0.8 g/dL group, 9.7 [0.8] cfu/g) versus the placebo group (7.2 [4.9] cfu/g, P < 0.001). This effect was dose dependent (0.4 g/dL versus 0.8 g/dL, P < 0.01). The number of Lactobacilli also increased significantly in both groups fed the supplemented formulas (versus placebo, P < 0.001), but there was no statistically significant difference between the group fed formula with 0.4 g/dL oligosaccharides and the group fed formula with 0.8 g/dL oligosaccharides. The dosage of supplement significantly influenced the change in fecal pH (P < 0.05) (placebo, pH 5.5-6.1; 0.4 g/dL formula, pH 5.48-5.44; 0.8 g/dL formula, pH 5.54-5.19). Slight changes in the stool frequency resulted in a significant difference between the placebo group and the group fed the 0.8 g/dL formula at day 28 (P < 0.01). Supplementation had a significant dose-dependent influence on stool consistency (0.8 g/dL versus placebo, P < 0.0001; 0.8 g/dL versus 0.4 g/dL, P < 0.01). Supplementation had no influence on the incidence of side effects (crying, regurgitation, vomiting) or growth. CONCLUSIONS: These data indicate that supplementation of a term infant's formula with a mixture of galacto- and fructooligosaccharides has a dose-dependent stimulating effect on the growth of Bifidobacteria and Lactobacilli in the intestine and results in softer stool with increasing dosage of supplementation.

Bifidobacterium↗
PubMed2001

Paclitaxel administration on days 1 and 8 every 21 days in anthracycline-pretreated metastatic breast cancer patients. A multicenter phase II trial.

Paclitaxel is now included in second- and even first-line regimens in advanced breast cancer. The optimal dose and schedule of this drug, however, still remain a matter of investigation. A group of 57 consecutive patients with advanced breast cancer previously treated with anthracycline-containing regimens were submitted to treatment with single-agent paclitaxel administered at 130 mg/m2 on days 1 and 8 every 21 days. Of the 57 patients, 56 were fully evaluable, and of these 25 had an absolute anthracycline resistance, 14 a relative resistance and 17 were potentially sensitive. The median age of the patients was 57 years (range 33-71 years), their median performance status was 1 (0-3), and 27 (47%) had liver involvement, 17 (30%) lung involvement, 30 (53%) bone involvement and 15 (26%) skin/lymph node involvement. Toxicity was recorded in 295 cycles. This scheme was well tolerated, the dose-limiting toxicities being hematological and neurological. Grade 3/4 leukopenia was observed in 20% of patients at nadir, while grade 3 leukopenia was observed in 3% of patients at recycle. Only one patient experienced febrile neutropenia. Grade 2/3 neurotoxicity was observed in 26% of patients, leading to drug withdrawal in three. The treatment was given on an outpatient basis in all patients and the median relative dose intensity of 86.6 mg/m2 per week was 100% of the planned dose (range 75-100%). Three patients (5%) attained a complete clinical response and 12 (21%) a partial response for an overall response rate of 26% (95% confidence interval 18-38%), while 30 (53%) attained disease stabilization and 11 progressed (19%). Time to progression in responding patients was 10.3 months, and the median overall survival of the entire population was 15.4 months. To conclude, paclitaxel administration on days 1 and 8 every 21 days was active and manageable in advanced breast cancer patients previously treated with anthracyclines. The response obtained was durable.

Adult↗
PubMed2001

Inducible IL-2 production by dendritic cells revealed by global gene expression analysis.

Dendritic cells (DCs) are strong activators of primary T cell responses. Their priming ability is acquired upon encounter with maturation stimuli. To identify the genes that are differentially expressed upon maturation induced by exposure to Gram-negative bacteria, a kinetic study of DC gene expression was done with microarrays representing 11,000 genes and ESTs (expressed sequence tags). Approximately 3000 differentially expressed transcripts were identified. We found that functional interleukin 2 (IL-2) mRNA, which gave rise to IL-2 production, was transiently up-regulated at early time-points after bacterial encounter. In contrast, macrophages did not produce IL-2 upon bacterial stimulation. Thus, IL-2 is an additional key cytokine that confers unique T cell stimulatory capacity to DCs.

Animals↗
PubMed2000

Fat intake and metabolism in Swedish and Italian infants.

The purpose of the study was to compare fat intake and metabolism between two infant populations from Sweden and Italy given breast milk or similar infant formulas, but different weaning foods. Nutrient intake and fat metabolism were studied prospectively from 3-12 mo in 68 Swedish and 46 Italian healthy infants, breastfed or given similar infant formulas in combination with Swedish or Mediterranean weaning foods. Although nutrient intake and fat metabolism were similar at 6 mo, fat intake was lower at 12 mo in the Italian than in the Swedish formula group (p < 0.001). At 6 and 12 mo, higher dietary ratios of monounsaturated to saturated fatty acids (p < 0.01 and p < 0.001, respectively), and monounsaturated to polyunsaturated fatty acids (p < 0.05, p < 0.001) were found in the Italian than in the Swedish formula group. Total cholesterol and apolipoprotein B were lower at 6 mo (p < 0.01) in Italian breastfed infants than in Swedish ones. Lower concentrations at 6 and 12 mo of total cholesterol (p < 0.05, p < 0.05, respectively), apolipoprotein B (p < 0.05, p < 0.01) and triglycerides (p < 0.001, p < 0.01), and of apolipoprotein A1 (p < 0.01) at 12 mo, were found in the Italian formula group than in the Swedish one. In conclusion, plasma total cholesterol, apolipoprotein B and triglycerides were found to be lower in Italian infants than in Swedish infants during the second half of infancy. These findings may partly result from differences in fat compositions between Swedish and Mediterranean weaning diets and in total fat intake in late infancy. Differences in duration of breastfeeding and possibly in breast milk composition may also have influenced our results.

Age Factors↗
PubMed2000

Protein intake and metabolism in formula-fed infants given Swedish or Italian weaning foods.

The aim of the study was to compare protein intake and metabolism between infants from two countries given similar infant formulae but different weaning foods. Healthy Swedish and Italian infants were studied between 3 and 12 mo. Infants in both populations were assigned to 1 of 3 infant formulae, containing 13, 15 or 18/20 g l(-1) of protein, given in addition to Swedish or Italian weaning foods. Protein intake from weaning foods was higher in Italian than in Swedish infants at 6 and 12 mo, whereas protein intake from formula at 6 mo and from formula/milk at 12 mo was similar in both populations. Plasma isoleucine, leucine, lysine, histidine and valine at 6 mo were lower in Italian than in Swedish infants fed formula with 13 g l(-1) of protein. All essential plasma amino acids were similar in Italian and Swedish groups at 12 mo. Serum urea was similar at 6 mo in corresponding formula groups, but was higher at 12 mo in the Italian than in the Swedish formula group. Serum albumin and growth were normal in both populations throughout infancy. In conclusion, formula with protein content of 13 g l(-1) seems to provide sufficient protein intake when combined with Swedish or Italian weaning foods during the second half of infancy, as indicated by normal serum albumin and normal growth. However, the bioavailability of protein and amino acids from weaning foods, in addition to their protein content, should be considered, as indicated by some indices of protein metabolism in the Italian infants.

Albumins↗
PubMed1999

Postprandial plasma amino acids in preterm infants: influence of the protein source.

Preprandial plasma amino acid concentrations have been used extensively as a marker of the nutritional value of dietary proteins in preterm infants. This study investigated the postprandial plasma amino acid profiles of preterm infants fed with different dietary proteins at similar protein intakes during the first weeks of life. In 12 preterm infants, pre- and postprandial plasma amino acid concentrations were measured before the removal of an indwelling central venous catheter placed for parenteral nutrition. All infants received breast milk until the time of study. At the start day of the study, infants were randomized to receive a test meal of 10 ml/kg, either of breast milk fortified with breast milk protein to reach a protein content of 2.0 g/dl or of a bovine milk preterm formula with a protein content of 2.0 g/dl (whey/casein ratio 60/40). Five samples of 100 microl blood were obtained immediately before and 15, 30, 45 and 60 min after the test meal. The plasma amino acid analysis was performed by a reversed-phase high-performance liquid chromatography based on o-phthaldialdehyde/2-mercaptoethanol pre-column derivatization. In both groups, the plasma amino acid concentrations increased within the first 30 min and the levels did not return to the preprandial baseline during the observation period. Fifteen minutes after the test meal, the plasma levels of all essential amino acids with the exception of histidine were higher in the bovine milk formula fed infants than in the fortified breast milk fed infants. The sum of plasma essential amino acid levels found in the formula fed infants were significantly (p < 0.05) higher than the levels found in the fortified breast milk fed infants at 15, 30 and 45 min. The kinetics of individual amino acids were influenced by the different quality of the protein even when the intakes in the groups were similar, as demonstrated for histidine and phenylalanine. The data indicate that postprandial plasma amino acid concentrations depend significantly on the dietary amino acid source and cannot simply be calculated from the amino acid composition of dietary proteins. Therefore, postprandial plasma amino acid concentrations should be included in the nutritional evaluation of dietary proteins in preterm infants.

Amino Acids↗
PubMed1999

Study of calcium metabolism in idiopathic hypercalciuria by strontium oral load test.

Calcium excretion and absorption were evaluated in hypercalciuric calcium stone formers by the study of Sr2+ excretion and absorption after an oral load. Ca2+ stone formers (n = 140) were studied, and the results were compared in the 83 of them who had idiopathic hypercalciuria and in the 57 who had Ca2+ excretion within reference values. Hypercalciuric patients showed increased renal Sr2+ clearance (CRE; 5.26 +/- 0.358 vs 3.29 +/- 0.277 mL/min; P <0.001), whereas Sr2+ absorption [assessed as the area under the serum concentration-time curve (AUC)] was increased at 30 and 60 min (1.53 +/- 0.087 vs 1.21 +/- 0.071 mmol. L-1. min; P <0.05), but not at 240 min after the load. In hypercalciuric patients, the AUCs were positively correlated with urinary Sr2+ fractional excretion (P <0. 001). Conversely, in normocalciuric patients plasma parathyroid hormone (PTH) was negatively correlated with the AUCs (P <0.01) and CRE (P <0.05), whereas 1,25-dihydroxyvitamin D plasma concentrations normalized to PTH were positively correlated with the AUCs (P <0.05). The results of Sr2+ load tests suggest that in the hypercalciuric population, Ca2+ absorption is altered predominantly in the duodenum and that the normal regulation exerted by calciotropic hormones on tubular and enteral Ca2+ handling is lost.

Administration, Oral↗
PubMed1998

Cisplatin, epirubicin, and lonidamine combination regimen as first-line chemotherapy for metastatic breast cancer: a pilot study.

We assessed the activity and tolerability of a cisplatin, epirubicin, and lonidamine combination regimen as first-line chemotherapy in 28 advanced breast cancer patients. The schedule of treatment was as follows: 60 mg/ m2 epirubicin followed by 40 mg/m2 cisplatin given on days 1 and 2 every 21 days, with 450 mg lonidamine being given per os (three tablets) on days of chemotherapy administration and in the period intervening between one cycle and the next. Patients received a median of 5 (range 1-6) cycles. Overall, 22 patients were evaluable for response and 28, for toxicity. Four patients refused to continue the treatment after the first course, one was lost to follow-up, and one died due to toxicity (septic shock). The incidence of grade 3/4 nausea and vomiting was found to be greater than that expected with epirubicin and lonidamine alone. The addition of cisplatin resulted in an increase in platelet and hemoglobin toxicities, whereas the WBC toxicity did not differ from that expected with epirubicin and lonidamine. The hematological toxicity was found to be cumulative, leading to treatment delay in about 50% of patients at the fifth and sixth courses. The activity of this cytotoxic regimen was noteworthy, with the overall response rate being 81.8% (31.8% complete responses and 50.0% partial responses) in evaluable patients. This response rate decreased to 64.2% when all registered patients were included according to an intent-to-treat analysis. In conclusion, the association of cisplatin, epirubicin, and lonidamine given on the schedule described herein, appears to be very active but substantially toxic. We are now testing this combination in a randomized comparison, with the cisplatin dose being reduced to 30 mg/m2 given on days 1 and 2.

Administration, Oral↗
PubMed1998

Usefulness of short-term urine collection in the nutritional monitoring of low birthweight infants.

To establish adequacy of urine collection times shorter than 24h in the metabolic monitoring of low birthweight infants, we collected urine for 24 h in 39 LBW infants during the third and fourth week of life. All urine voidings over the 24-h period were separately collected, the volume of each sampling and the time of voiding were recorded, and 20% of the volume was removed for pooling. All individual and pooled samples were analysed for total nitrogen, urea and ammonia, alpha-amino nitrogen, creatinine, sodium, potassium, calcium and phosphorus, and for each compound the ratio to 1 mol creatinine was established. Individual sample results were "pooled" to obtain 3-, 6- and 12-h period excretion and than related to the 24-h excretion as measured in the pooled 24-h sample. As the volume of urine obtained in any 6-h collecting period depended on the time of sampling (06:00-12:00 h, 17.5+/-3.1% of total; 12:00-18:00 h, 31.6+/-5.1% of total; 18:00-24:00 h, 25.6+/-3.1% of total; and 0:00-06:00h, 25.3+/-2.9% of total), calculations were based on samples obtained from 18:00 to 06:00 h. The correlation between results of 3- and 24 h-collection periods was weakest, while results of the 6-h collection correlated highly with the total daily excretion (r = between 0.82 and 0.93 for the different compounds) and the correlation was only slightly better when the 12-h collection period was considered. The correlation between the mean molar substrate/creatinine ratio of all individual samples of a 24-h collecting period and the and total daily excretion of the respective substrate was weaker (r = between 0.46 and 0.76 for the different compounds) than the correlation between the results of a 6-h collecting period and the daily excretion is not as stable than in later life. The data indicate that 6-h urine sampling may be sufficient for metabolic monitoring of LBW infants. By contrast, urinary substrate/creatinine ratios are not good markers of the daily excretions of the respective substrate during the first weeks of life.

Ammonia↗
PubMed1998

Postnatal development of urea synthesis capacity in preterm infants with intrauterine growth retardation.

The postnatal development of the urea-synthesizing capacity was studied in 21 preterm infants with intrauterine growth retardation (IUGR) and compared with results found in 12 infants without IUGR as controls. The urea-synthesizing capacity was estimated by the ratio Q of 15N abundance of ammonia and urea in 6-hour urine samples collected after enteral administration of 3 mg [15N]H4Cl/kg body weight. The measurements were performed on the first day when a protein intake of 3.0-3.5 g/kg/day and an energy intake of 120 kcal/kg/day were tolerated (study day 1: postnatal 14-21 days) and on the day of discharge from the hospital (study day 2: postnatal age 39-56 days). The group of infants with IUGR was subdivided in one group of infants who developed catch-up growth (n = 12) and one group who did not demonstrate catch-up growth (n = 9). On study day 1, the Q values of the IUGR infants without catch-up growth were significantly higher than those of the IUGR infants with catch-up growth (13.4 +/- 2.3 vs. 9.2 +/- 2.2) or of the control infants without IUGR. During the time period from study day 1 to study day 2 the Q values of the IUGR infants with catch-up growth decreased significantly (9.2 +/- 2.2 vs. 4.8 +/- 2.0; p < 0.001) and were in the range of the control infants without IUGR. In contrast, the Q values of the IUGR infants without catch-up growth did not significantly change during the study period (13.4 +/- 2.3 vs. 11.3 +/- 2.8; p = 0.097). On both study days there was a significant correlation between the Q values and the degree of IUGR (study day 1: r = 0.652, p < 0.01; study day 2: r = 0.842, p < 0.001). The data indicate that the urea-synthesizing capacity of preterm infants increases during early postnatal life and that severe IUGR may impair this development. Metabolic investigations using urea as marker for evaluation of optimal quantity or quality of dietary proteins should carefully be interpreted when infants with severe IUGR are studied.

Child Development↗
PubMed1998

Responsiveness of urinary markers of bone resorption to orchiectomy and clodronate treatment in mature rats: a comparative study.

OBJECTIVE: The aim of this study was to assess the responsiveness of two classes of bone resorption markers to the enhancement of osteoclastic activity induced by orchiectomy and to its inhibition by clodronate treatment in mature rats. DESIGN: Bone mineral density (BMD) at femural metaphysis, femural diaphysis, lumbar vertebrae, and the urinary excretion of pyridinoline (Pyr), deoxypyridinoline (D-Pyr), galactosylhydroxylysine (GHyl) and glucosylgalactosylhydroxylysine (GGHyl) were monitored at regular intervals for 30 days prior to and for 60 days following orchiectomy in eleven rats, divided into two groups: five rats untreated and the other six treated with clodronate. RESULTS: Prior to orchiectomy, a significant (P < 0.01) decrease in BMD was observed only at the distal femural metaphysis. This decrease appeared to be associated with a time-dependent increase in the urinary excretion of all markers. Following orchiectomy, the BMD of the untreated group decreased significantly (P < 0.01) at all bone sites. The bone loss was accompanied by a significant (P < 0.01) increase in Pyr and D-Pyr concentrations in urine, whereas urinary GHyl and GGHyl did not change significantly. In the clodronate-treated group, the BMD of the three skeletal sites did not change significantly, while the urinary excretion of all urinary biochemical markers decreased significantly (P < 0.001). CONCLUSION: This study showed that pyridinolines are able to monitor the bone response to orchiectomy and to clodronate treatment response in androgen-deficient mature male rat. whereas glycosides appear prone to confounding factors.

Aging↗
PubMed1998

Strontium absorption and excretion in normocalciuric subjects: relation to calcium metabolism.

The relationships of Sr intestinal absorption and renal excretion with biohumoral factors regulating Ca metabolism were studied in 47 normocalciuric subjects with Ca kidney stones. Sr concentrations were measured in serum and urine after an oral load of stable Sr (30.2 mumol/kg body wt). Enteral absorption of the ion (9.77 +/- 0.438 mmol.L-1.min, 240 min after Sr administration), expressed as the area under the plasma concentration-time curve (AUC), and renal clearance (CRE) in these subjects during the test (2.80 +/- 0.336 mL/min) were not different from values for 27 controls. CRE was not correlated with AUCs. Plasma concentrations of parathyroid hormone (PTH) negatively correlated with AUCs (P < 0.01) and correlated with CRE after one outlier was excluded (P < 0.05). Plasma concentrations of 1,25-dihydroxyvitamin D correlated positively with AUCs (P < 0.01) when normalized to the plasma concentration of PTH. Multiple stepwise regression showed that PTH and phosphatemia were significantly related to AUC values at 240 min (P < 0.01). These findings suggest that Sr absorption and excretion reflect the regulation of Ca metabolism, but some differences in renal handling of the two ions may exist.

Administration, Oral↗
PubMed1997

T-tubule membranes from chicken skeletal muscle possess an enzymic cascade for degradation of extracellular ATP.

The chicken T-tubule Mg2+-ATPase is an integral membrane glycoprotein that presents properties different from those of other ATPases located in skeletal muscle cells and exhibits ATP-hydrolysing activity on the extracellular side of the transverse tubule (TT) membranes. In this study we demonstrate that TT vesicles purified from chicken skeletal muscle possess ecto-ADPase and ecto-5'-nucleotidase activities that, along with ecto-ATPase, are able to sequentially degrade extracellular ATP to ADP, AMP and adenosine. Characterization studies of these TT ectonucleotidases revealed remarkable differences between ecto-ATPase and ecto-ADPase activities with respect to thermal stability, temperature dependence of the hydrolytic activity, effect of ionic strength, kinetic behaviour, divalent cation preference and responses to azide, N-ethylmaleimide, NaSCN, Triton X-100 and concanavalin A. Ecto-ATPase, but not ecto-ADPase, was inhibited by a polyclonal antibody against the chicken TT ecto-ATPase. On the basis of these results we propose that ATP and ADP hydrolysis are accomplished by two distinct enzymes and therefore the TT ecto-ATPase is not an apyrase. 5'-Nucleotidase activity was inhibited by adenosine 5'-[alpha,beta-methylene]diphosphate and concanavalin A, followed simple Michaelis-Menten kinetics and was released from the membranes by treatment with phosphatidylinositol-specific phospholipase C, indicating that AMP hydrolysis in T-tubules is catalysed by a typical ecto-5'-nucleotidase. Results obtained from electrophoresis experiments under native conditions suggest that ecto-ATPase, ecto-ADPase and 5'-nucleotidase might be associated, forming functional complexes in the T-tubule membranes. The TT ectonucleotidases constitute an enzymic cascade for the degradation of extracellular ATP that might be involved in the regulation of purinergic signalling in the muscle fibre.

5'-Nucleotidase↗